Regulation of Cell Cycle Progression

What are the key regulators of cell cycle progression in mitosis?

Transcriptional Regulation of Cyclin/CDK Complexes

Transcriptional regulation plays a critical role in the control of cyclin/CDK complexes, which are key regulators of the cell cycle. Several mechanisms have been identified that affect the activity of these complexes.ref.9.4 ref.8.30 ref.7.2

1. Transcriptional Regulation by E2F Transcription Factor 1 (E2F) E2F transcription factor 1 (E2F) is a major regulator of cell cycle progression. It controls the expression of genes involved in DNA synthesis and cell division.ref.9.4 ref.8.30 ref.9.5 E2F promotes the transcription of genes encoding cyclin D, cyclin E, cyclin A, and cyclin B, which are essential for the G1/S and G2/M transitions of the cell cycle. Activation of E2F leads to the increased expression of these cyclins, promoting cell cycle progression. On the other hand, inhibition of E2F activity results in the downregulation of cyclin expression, leading to cell cycle arrest.ref.9.9 ref.9.18 ref.9.5

2. Transcriptional Regulation by NFκB Signaling NFκB signaling is a crucial pathway involved in various cellular processes, including inflammation, immunity, and cell survival. Recent studies have shown that NFκB also plays a role in the regulation of cyclin/CDK complexes.ref.8.30 ref.77.30 ref.8.24 NFκB can directly bind to the promoters of cyclin B1, cyclin B2, Plk-1, and cdc25 genes and activate their transcription. This leads to the increased expression of these cyclins and cdc25, which in turn promotes cell cycle progression.ref.8.30 ref.8.24 ref.8.24

3. Transcriptional Regulation by Cdh1 Cdh1 is a substrate recognition subunit of the anaphase-promoting complex/cyclosome (APC/C), a ubiquitin ligase that regulates cell cycle progression. Cdh1 targets various proteins for degradation, including cyclin B and Cdc20, leading to their proteasomal degradation and cell cycle arrest.ref.9.1 ref.9.1 ref.9.6 However, recent studies have also shown that Cdh1 can regulate the transcription of cyclin B1. It acts as a transcriptional repressor by binding to the promoter region of cyclin B1 and inhibiting its expression. This provides an additional level of regulation for cyclin B1 expression and cell cycle progression.ref.9.6 ref.9.1 ref.9.1

4. Transcriptional Regulation of Cyclin B1, Cyclin B2, Plk-1, and Cdc25 by NFκB In addition to direct regulation of cyclin B1, cyclin B2, Plk-1, and cdc25 genes by NFκB, there is evidence to suggest that NFκB can also indirectly regulate the expression of these genes. NFκB can activate the transcription of other factors, such as Myc and Jun, which in turn regulate the expression of cyclin B1, cyclin B2, Plk-1, and cdc25.ref.8.24 ref.8.30 ref.8.30 This indirect regulation provides a more complex and interconnected network of transcriptional regulation that controls the activity of cyclin/CDK complexes.ref.9.4 ref.9.8 ref.8.30

5. Transcriptional Regulation of Cyclins by Cytokine- and Growth Factor-Induced Pathways Cytokines and growth factors are key regulators of cell proliferation and differentiation. They activate signaling pathways that lead to the transcriptional regulation of cyclins, which are essential for cell cycle progression.ref.9.31 ref.9.9 ref.9.5 These pathways can activate transcription factors, such as STATs and AP-1, which in turn regulate the expression of cyclins. For example, cytokine-induced activation of STAT3 can lead to the upregulation of cyclin D1 expression, promoting cell cycle progression.ref.9.31 ref.9.9 ref.9.5

Protein Inhibitors and the Regulation of Cyclin/CDK Complexes

Protein inhibitors play a crucial role in the regulation of cyclin/CDK complexes and cell cycle progression. They control the activity of CDKs by inhibitory phosphorylation and dephosphorylation events.ref.2.4 ref.9.4 ref.15.1

1. Inhibitory Phosphorylation and CDK Inactivation CDKs are kept in an inactive state by inhibitory phosphorylation at specific sites, such as Thr 14 and Tyr 15. These phosphorylation events prevent the binding of cyclins to CDKs and inhibit their activity.ref.17.2 ref.17.2 ref.4.1 Several kinases, including Wee1, contribute to this inhibitory phosphorylation. Wee1 kinase phosphorylates CDKs at these inhibitory sites, keeping them in an inactive state and preventing premature cell cycle progression.ref.108.10 ref.17.2 ref.11.26

2. Cdc25 Phosphatases and CDK Activation Cdc25 phosphatases are key regulators of CDK activity and cell cycle progression. They remove the inhibitory phosphates from CDKs, leading to their activation and cell cycle progression.ref.14.5 ref.4.6 ref.4.2 Cdc25 phosphatases are regulated by various mechanisms, including phosphorylation events mediated by CHK1/2 kinases. CHK1/2-mediated phosphorylation of Cdc25 proteins results in their functional inactivation, preventing CDKs dephosphorylation and activation. This mechanism is important for the activation of the cell cycle checkpoint in response to DNA damage.ref.14.5 ref.4.2 ref.4.2

3. Wee1 Kinase and Negative Feedback Loop Wee1 kinase is another important regulator of CDK activity. It inhibits CDK activity by phosphorylating CDKs, contributing to the negative feedback loop that controls Cyclin B activity.ref.9.10 ref.4.6 ref.4.2 The balance between Wee1/Myt1 kinases and Cdc25 phosphatases regulates the inhibitory phosphorylation of CDKs. When Cyclin B levels are high, Wee1 kinase is activated, leading to the inhibitory phosphorylation of CDKs and preventing premature cell cycle progression. This negative feedback loop ensures proper cell cycle control and the maintenance of genomic integrity.ref.9.10 ref.11.26 ref.4.2

4. Regulation of CDK Activity for Proper Cell Cycle Progression The regulation of CDK activity by protein inhibitors is crucial for proper cell cycle progression and the maintenance of genomic integrity. The balance between inhibitory phosphorylation and dephosphorylation events ensures that CDKs are activated at the appropriate time and in the correct order.ref.2.4 ref.14.5 ref.9.4 Dysregulation of this balance can result in cell cycle defects and genomic instability, which are associated with various diseases, including cancer.ref.7.2 ref.2.4 ref.7.2

Positive Feedback Regulation and Mitotic Phosphatases in the Control of Mitosis

Proper control of the duration of mitosis is essential for the fidelity of cell division and cell survival. Positive feedback regulation and mitotic phosphatases play important roles in achieving this control.ref.7.37 ref.7.22 ref.7.1

1. Positive Feedback Regulation of Cdk1 Positive feedback regulation helps achieve proper activation thresholds for Cdk1, which is necessary for progression and exit from mitosis. Cdk1 is activated by cyclin B, and once activated, it can phosphorylate and activate more cyclin B, leading to further Cdk1 activation.ref.4.6 ref.7.22 ref.7.37 This positive feedback loop ensures that Cdk1 activity reaches a critical threshold to initiate and maintain mitosis. It also helps ensure a short and constant duration of mitosis.ref.7.37 ref.7.22 ref.7.19

2. Mitotic Phosphatases and Cdk1 Substrate Dephosphorylation Mitotic phosphatases, specifically members of the PP1 and PP2A families, are involved in regulating the entry and exit from mitosis by reversing Cdk1 substrate phosphorylation. These phosphatases dephosphorylate Cdk1 substrates, leading to their inactivation and promoting mitotic exit.ref.1.17 ref.1.4 ref.1.6 The activity of these phosphatases is tightly regulated and coordinated with other mitotic regulators, such as the spindle assembly checkpoint, to ensure proper timing and control of mitosis.ref.1.4 ref.1.6 ref.17.28

3. Temporal Insulation of Mitosis through Positive Feedback Regulation Computational modeling studies have shown that positive feedback regulation can account for the observed temporal insulation of mitosis. Temporal insulation refers to the short, constant duration of mitosis that is uncoupled from the variability in earlier cell cycle phases.ref.7.1 ref.7.10 ref.7.0 Positive feedback regulation ensures that mitosis remains a short and constant fraction of the cell cycle, despite variations in the duration of other cell cycle phases. This temporal insulation is crucial for the fidelity of cell division and the maintenance of genomic integrity.ref.7.1 ref.7.0 ref.7.37

4. Positive Feedback Regulation as a Regulatory Strategy Positive feedback regulation is a commonly used regulatory strategy in biological systems to create modularity and ensure proper control of cellular processes. By incorporating positive feedback loops, cells can achieve robust and precise control of critical events, such as cell cycle progression.ref.7.37 ref.7.37 Positive feedback regulation helps maintain a short and constant duration of mitosis, preventing the accumulation of errors and ensuring the fidelity of cell division.ref.7.37 ref.7.37

In conclusion, transcriptional regulation and protein inhibitors play crucial roles in the control of cyclin/CDK complexes and cell cycle progression. Transcriptional regulation by factors such as E2F and NFκB affects the expression of cyclins and other cell cycle regulators. Protein inhibitors, such as Wee1 and Cdc25, regulate the activity of cyclin/CDK complexes through inhibitory phosphorylation and dephosphorylation events.ref.9.4 ref.8.30 ref.2.4 Positive feedback regulation and mitotic phosphatases contribute to the control of the duration of mitosis. These regulatory mechanisms ensure proper cell cycle progression, the maintenance of genomic integrity, and the fidelity of cell division. Understanding these regulatory processes is important for elucidating the mechanisms underlying cell cycle control and their dysregulation in various diseases, including cancer.ref.7.37 ref.7.22 ref.7.2 Further research in this field will provide insights into potential therapeutic targets for the treatment of cell cycle-related disorders.ref.7.2 ref.8.30 ref.7.2

How are these regulators activated and inactivated during the cell cycle?

Activation of regulators

The regulators of cell cycle progression, such as cyclins and cyclin-dependent kinases (CDKs), are essential for the proper ordering of cell cycle phases and the completion of one phase before the onset of the next phase. These regulators are activated through various mechanisms.ref.9.4 ref.2.4 ref.7.2

1.1 Cyclin-dependent kinases (CDKs) activationref.15.1 ref.9.4 ref.15.1

CDKs are activated when they form a complex with their corresponding cyclin proteins. Different cyclin-CDK complexes promote progression through different phases of the cell cycle. For example, the Cyclin D/CDK4-6 complex promotes progression in the G1 phase, the Cyclin E/CDK2 complex promotes the transition from G1 to S phase, the Cyclin A/CDK2 complex promotes progression in S and G2 phases, and the Cyclin B/CDK1 complex promotes the G2/M phase transition.ref.9.4 ref.2.5 ref.8.30

1.2 Transcriptional regulation

Another mechanism of activation involves transcriptional regulation. E2F transcription factor 1 (E2F) regulates the sequential activation of cyclins. E2F activity is tightly controlled by binding to the retinoblastoma protein (Rb).ref.9.5 ref.2.5 ref.9.9 Cyclins phosphorylate Rb, leading to the activation of E2F and subsequent transcription of cyclins.ref.9.5 ref.9.9 ref.9.5

Inactivation of regulators

The inactivation of regulators is equally important for proper cell cycle progression. Several mechanisms contribute to the inactivation of CDKs and the binding of cyclin-dependent kinase inhibitors (CKIs).ref.9.4 ref.2.4 ref.7.2

2.1 Phosphorylation/dephosphorylation

Inactivation of CDKs can occur through inhibitory phosphorylation by proteins such as Wee1 and activation through dephosphorylation by phosphatases such as Cdc25. Wee1 phosphorylates CDKs, rendering them inactive, while Cdc25 dephosphorylates CDKs, leading to their activation.ref.4.2 ref.11.11 ref.17.2

2.2 Association with protein inhibitors

CDKs can be inhibited by binding to specific regulatory subunits called cyclin-dependent kinase inhibitors (CKIs). Examples of CKIs include p21, p27, and p57. These CKIs bind to CDKs and prevent their activity by forming complexes with their CDK partners.ref.2.4 ref.9.4 ref.14.5 This binding prevents CDKs from phosphorylating target proteins involved in DNA replication and mitosis, thereby inhibiting cell cycle progression.ref.14.5 ref.14.5 ref.15.1

2.3 Targeted proteolysis

Cyclins and CDK inhibitors can be degraded through the ubiquitin-dependent proteolysis system. The degradation of cyclins is mediated by complexes formed with anaphase-promoting complex (APC) and Cdc20 homolog 1 (Cdh1) or Cell division cycle 20 (Cdc20) (APCCdh1, APCCdc20). CKIs can also be degraded by SCF (Skp1/cullin/F-box protein related complexes).ref.35.6 ref.2.45 ref.30.6

Feedback regulation and cell cycle progression

The binding of cyclins to CDKs leads to their activation and subsequent progression through different phases of the cell cycle. This activation is tightly regulated at multiple levels to ensure the proper ordering of cell cycle phases and the completion of one phase before the onset of the next.ref.9.5 ref.9.4 ref.2.4

One example of feedback regulation is the control of Cyclin B activity, which drives mitosis. Cyclin B activity is controlled by a double negative feedback loop from the inhibitory kinase Wee1 and a positive feedback loop from the activating phosphatase Cdc25. Wee1 phosphorylates and inhibits CDKs, including CDK1 bound to Cyclin B, while Cdc25 dephosphorylates and activates CDKs.ref.9.10 ref.4.6 ref.7.40 The balance between Wee1 and Cdc25 activities determines the activation state of Cyclin B/CDK1 and ultimately controls the progression into mitosis.ref.9.10 ref.9.10 ref.4.6

The duration of mitosis is short, constant, and uncoupled from variability in early cell cycle phases. This is achieved through the coordination of CDK activity, such as Cdc28 in S. cerevisiae, which is regulated by the availability of cyclin partners, inhibitory phosphorylation, and binding to CDK inhibitors.ref.7.36 ref.7.36 ref.7.3

Phosphorylation of Rb and sequential activation of cyclins

The phosphorylation of Rb by cyclins contributes to the activation of E2F and the transcription of cyclins through a positive feedback loop mechanism. Cyclin D and Cyclin E sequentially phosphorylate Rb, leading to the activation of E2F. Once E2F is activated, it transactivates the transcription of Cyclin A and Cyclin B.ref.9.5 ref.9.31 ref.9.31 This sequential activation of cyclins is essential for the progression of the cell cycle, allowing entry into S phase, progression in S and G2 phase, and finally the G2/M phase transition into mitosis.ref.9.34 ref.9.5 ref.9.4

Additional proteins and factors involved in regulation

Besides Wee1 and phosphatases like Cdc25, there are other proteins or factors involved in the inhibitory phosphorylation or dephosphorylation of CDKs.ref.4.2 ref.4.1 ref.11.26

5.1 Polo kinase and 14-3-3 proteins

Polo kinase and 14-3-3 proteins also play a role in regulating the activity of CDKs and the progression of the cell cycle. Polo kinase phosphorylates CDKs, promoting their activation, while 14-3-3 proteins bind to phosphorylated CDKs, sequestering them in an inactive state.ref.9.4 ref.9.4 ref.3.7

5.2 Protein phosphatases (PPs)

Protein phosphatases (PPs), such as PP2A, are also involved in the dephosphorylation and inactivation of CDKs. PP2A dephosphorylates CDKs, leading to their inactivation and the subsequent inhibition of cell cycle progression.ref.1.4 ref.1.6 ref.1.17

Ubiquitin-dependent proteolysis system

The ubiquitin-dependent proteolysis system plays a significant role in regulating the degradation of cyclins and CDK inhibitors in the cell cycle. Several factors and enzymes are involved in this process.ref.2.4 ref.2.32 ref.67.9

6.1 Anaphase-promoting complex/cyclosome (APC/C)ref.57.1 ref.59.2 ref.57.1

The APC/C is responsible for tagging substrates like securin, Aurora kinase, and cyclin B for degradation. The degradation of cyclin B by the APC/C is crucial for the proper progression from G2 phase to mitosis.ref.30.6 ref.57.1 ref.66.3

6.2 Mitotic checkpoint complex (MCC)ref.59.0 ref.59.2 ref.59.2

The mitotic checkpoint complex (MCC) inhibits the APC/C until all chromosomes are properly attached to the spindle. The MCC is composed of proteins like Mad2, Mad3, and Bub3, and it ensures the fidelity of chromosome segregation during mitosis.ref.59.0 ref.59.2 ref.59.2

6.3 E3 ubiquitin ligasesref.2.10 ref.2.10 ref.2.10

E3 ubiquitin ligases, such as the SKP/CUL/RBX/F-box (SCF) complex, are also involved in the degradation of cyclins and CDK inhibitors. The SCF complex recognizes specific substrates and adds ubiquitin molecules to them, marking them for degradation.ref.2.10 ref.2.10 ref.2.10

In conclusion, the regulators of cell cycle progression, including cyclins and CDKs, are activated and inactivated through various mechanisms to ensure the proper ordering of cell cycle phases and the completion of one phase before the onset of the next phase. These mechanisms involve the formation of cyclin-CDK complexes, transcriptional regulation, phosphorylation/dephosphorylation, association with protein inhibitors, and targeted proteolysis. Feedback regulation and sequential activation of cyclins further contribute to the progression of the cell cycle.ref.9.4 ref.2.4 ref.7.2 Additionally, other proteins and factors, such as polo kinase, 14-3-3 proteins, and protein phosphatases, play a role in regulating CDK activity. The ubiquitin-dependent proteolysis system, involving the APC/C, MCC, and E3 ubiquitin ligases, ensures the degradation of cyclins and CDK inhibitors. Together, these regulatory mechanisms ensure the proper progression of the cell cycle and the fidelity of chromosome segregation during mitosis.ref.2.4 ref.9.4 ref.9.6

What are the checkpoints in the cell cycle and how do they ensure proper progression into mitosis?

Introduction

The cell cycle is a highly regulated process that ensures the accurate duplication and division of cells. It is governed by checkpoints that verify the completion of each phase before allowing progression into the next phase. The checkpoints in the cell cycle include the G1-to-S transition, the G2-to-M transition, and the M-to-G1 transition.ref.2.4 ref.7.2 ref.6.4 These checkpoints coordinate the timing of cell cycle phases, prevent premature entry into mitosis, and ensure accurate DNA replication and chromosome segregation. This essay will explore the regulation and significance of these checkpoints in maintaining the fidelity of the cell cycle and preventing diseases such as cancer.ref.7.2 ref.2.4 ref.7.1

G1-to-S Transition

The G1-to-S transition is a critical checkpoint in the cell cycle that ensures the accurate progression from the G1 phase to the S phase, where DNA replication occurs. This transition is regulated by cyclin-dependent kinases (CDKs) and cyclins. CDKs are enzymes that control the progression of the cell cycle, while cyclins are regulatory proteins that activate CDKs.ref.6.4 ref.2.4 ref.8.30 At the G1-to-S transition, the levels of cyclins increase, leading to the activation of CDKs. The activated CDKs phosphorylate target proteins involved in DNA replication, promoting the initiation of the S phase.ref.9.4 ref.6.4 ref.8.30

G2-to-M Transition

The G2-to-M transition is another crucial checkpoint in the cell cycle, regulating the progression from the G2 phase to mitosis. Like the G1-to-S transition, the G2-to-M transition is regulated by CDKs and cyclins. The activation of CDKs at this checkpoint promotes the entry into mitosis.ref.8.30 ref.6.4 ref.2.4 Specifically, CDK1, along with its regulatory protein Cyclin B1, plays a central role in this transition. The activity of CDK1-Cyclin B1 complex is controlled by a complex interplay of kinases and phosphatases, such as cdc25 and Plk, which regulate the phosphorylation status of CDK1. The dephosphorylation of CDK1 and the increased expression of cyclin B1 stimulate the activation of CDK1-Cyclin B1 complex, leading to the initiation of mitosis.ref.8.30 ref.2.4 ref.9.4

M-to-G1 Transition (Finish Transition)

The M-to-G1 transition, also known as the "Finish" transition, marks the completion of mitosis and the initiation of a new cell cycle. This transition is regulated by various mechanisms. One important regulator is the CDK complex, specifically CDK1 and Cyclin B1.ref.6.4 ref.8.30 ref.2.4 The activity and expression of CDK1 and Cyclin B1 are tightly controlled in a cell cycle-dependent manner. At the M-to-G1 transition, the activity of CDK1-Cyclin B1 complex is downregulated, allowing the cells to exit from mitosis and enter the G1 phase of the next cell cycle. Transcription factors like the forkhead family of transcription factors and NFκB also play a role in the transcriptional regulation of genes important for mitotic entry and exit.ref.8.30 ref.2.4 ref.8.30 NFκB, in particular, controls the transcription of cyclin B1, cyclin B2, Plk-1, and cdc25, thereby influencing the progression through the M-to-G1 transition.ref.8.30 ref.8.24 ref.8.30

Importance of Checkpoints in the Cell Cycle

The checkpoints in the cell cycle play a vital role in maintaining the fidelity of chromosome duplication and cell division. They prevent errors that can lead to diseases such as cancer. The accurate completion of each phase before the onset of the next phase ensures that DNA replication and chromosome segregation occur correctly.ref.2.4 ref.7.2 ref.79.0 Failure to regulate cell cycle progression at the checkpoints can have severe consequences. Genomic instability, characterized by an increased frequency of DNA mutations and chromosomal abnormalities, can result from errors in the checkpoints. The accumulation of mutations and genomic instability is a hallmark of cancer development.ref.79.0 ref.79.1 ref.79.0 Additionally, defects in the regulation of cell cycle progression can lead to uncontrolled cell growth and the formation of tumors.ref.24.2 ref.79.1 ref.79.0

Consequences of Checkpoint Malfunction in Cancer Development

Errors in the regulation of cell cycle progression at the checkpoints can contribute to the development of cancer. When the checkpoints fail to detect and repair DNA damage or other abnormalities, cells with these defects can continue dividing and proliferating. This can lead to the accumulation of mutations and genomic instability, which are key drivers of cancer development.ref.79.1 ref.73.9 ref.14.5 The specific consequences and mechanisms by which errors in the checkpoints contribute to cancer development may vary depending on the type of DNA damage and the phase of the cell cycle in which it occurs. Nevertheless, the malfunction of the checkpoints is a critical factor in the initiation and progression of cancer.ref.79.1 ref.73.9 ref.79.1

Conclusion

The checkpoints in the cell cycle are essential for maintaining the fidelity of chromosome duplication and cell division. They ensure proper progression into mitosis by coordinating the timing of cell cycle phases and verifying the accurate completion of each phase. CDKs, cyclins, and other regulatory proteins play a central role in regulating these checkpoints.ref.2.4 ref.7.2 ref.7.2 Failure to regulate cell cycle progression at the checkpoints can lead to genomic instability and the development of cancer. Understanding the molecular mechanisms and signaling pathways involved in the regulation of these checkpoints is crucial for developing targeted therapies to treat diseases such as cancer.ref.7.2 ref.14.5 ref.2.4

What are the molecular signals that trigger entry into mitosis?

Molecular Signals and Regulatory Mechanisms for Entry into Mitosis

The progression from the G2 phase to mitosis is regulated by a complex network of molecular signals and regulatory mechanisms. One of the key molecular signals involved in triggering entry into mitosis is the activation of the cyclin B and cyclin-dependent kinase 1 (CDK1) complex. This complex is responsible for driving the cell cycle from G2 phase to mitosis.ref.8.30 ref.9.4 ref.9.34 The activation of the cyclin B-CDK1 complex is regulated by several G2-M kinases and phosphatases, such as cdc25 and Plk (Nilsson and Hoffmann, 2000; Barr et al., 2004).ref.8.30 ref.2.4 ref.9.6

The expression of cyclin B, cyclin B2, Plk-1, and cdc25 is transcriptionally regulated. Studies have shown that blocking NFκB signaling at G2-M inhibits the transcription of these genes, indicating the involvement of NFκB in their transcriptional regulation. This finding provides new insights into the transcriptional mechanisms governing G2-M progression of the cell cycle.ref.8.30 ref.8.28 ref.8.2 The transcriptional regulation of these genes is not well defined, but it has been reported that the forkhead family of transcription factors, including FOXM1 and FoxO, play a critical role in the expression of genes important for mitotic entry and exit.ref.8.30 ref.8.24 ref.8.26

In addition to transcriptional regulation, the activity and expression of cdc25 and Plk are also regulated in a cell cycle-dependent manner. The rise of G1-S cyclins is accompanied by the appearance of Cyclin A during S phase. In the late G2 phase, a transient activation of Cyclin B is observed, enabling swift G2-M transition.ref.8.30 ref.8.30 ref.9.34 This transient activation is enabled by regulatory feedbacks imposed on Cyclin B by Cdc25 and Wee1. The activation and degradation of cyclins and their regulators are tightly regulated during the cell cycle progression.ref.9.6 ref.9.9 ref.9.10

Regulation of Cyclin/Cdk Complexes and Feedback Loops

The key molecular signals that trigger entry into mitosis involve the activation of various cyclin/Cdk complexes at different stages of the cell cycle. The progression from G1 phase to S phase is promoted by the activation of the cyclin D/Cdk4-6 complex. The transition from G1 to S phase is facilitated by the cyclin E/Cdk2 complex, while the progression in S and G2 phase is facilitated by the cyclin A/Cdk2 complex.ref.9.4 ref.9.34 ref.8.30 Finally, the G2/M phase transition allowing entry into mitosis is triggered by the cyclin B/Cdk1 complex.ref.8.30 ref.9.4 ref.9.34

These cyclin/Cdk complexes are regulated through various mechanisms. Transcriptional regulation plays a role in controlling the expression of cyclins and Cdks. Association with protein inhibitors, such as p21 and p27, also regulates the activity of cyclin/Cdk complexes.ref.9.4 ref.2.4 ref.15.1 Phosphorylation/dephosphorylation events and cyclin degradation further modulate the activity of these complexes.ref.2.4 ref.9.8 ref.15.1

The activation of Cyclin B and CDK1 complex, which is crucial for the G2/M phase transition, is controlled by a double negative feedback loop from the inhibitory kinase Wee1 and a positive feedback loop from the activating phosphatase Cdc25. Wee1 phosphorylates CDK1, inhibiting its activity, while Cdc25 dephosphorylates and activates CDK1. This regulatory mechanism ensures a precise and coordinated entry into mitosis.ref.9.10 ref.9.9 ref.8.30

Regulation of G2-M Kinases and Phosphatases

G2-M kinases and phosphatases play a crucial role in regulating the activation of the cyclin B-CDK1 complex during the G2-M phase transition. The activation of the cyclin B-CDK1 complex is stimulated by dephosphorylation of CDK1 and increased expression of cyclin B. This process is regulated by several G2-M kinases and phosphatases, including cdc25 and Plk.ref.8.30 ref.2.4 ref.9.6

The activity and expression of cdc25 and Plk are regulated in a cell cycle-dependent manner. They are involved in the dephosphorylation of CDK1 and the stimulation of cyclin B expression, ultimately enabling the transition into mitosis. The regulation of cdc25 and Plk is complex and involves multiple factors and mechanisms.ref.14.8 ref.3.7 ref.14.8 For example, Cdc25 is regulated by 14-3-3 proteins and protein phosphatases. Phosphorylation of Cdc25C at serine 287 triggers binding of 14-3-3 proteins and inhibition of phosphatase activity.ref.4.6 ref.4.2 ref.4.6

Role of NFκB Signaling in G2-M Progression

NFκB signaling has been found to play a role in regulating the transcription of cyclin B1, cyclin B2, Plk-1, and cdc25 genes during the G2-M phase transition of the cell cycle. Blocking NFκB signaling at G2-M inhibits the transcription of these genes, suggesting that NFκB is involved in their transcriptional regulation. This finding provides new insights into the transcriptional mechanisms governing G2-M progression of the cell cycle.ref.8.30 ref.8.28 ref.8.24

ERK5 activation has been shown to stimulate NFκB signaling, which in turn regulates the expression of these G2-M specific genes. The ERK5-NFκB signaling pathway is required for G2-M progression in cultured primary human cells. Therefore, NFκB signaling, activated by ERK5, plays a critical role in regulating the transcription of cyclin B, cyclin B2, Plk-1, and cdc25 genes during the G2-M phase transition of the cell cycle.ref.8.26 ref.8.26 ref.8.2

In conclusion, the progression from the G2 phase to mitosis is regulated by a complex network of molecular signals and regulatory mechanisms. The activation of cyclin B-CDK1 complex, along with the activation of other cyclin/Cdk complexes at different stages of the cell cycle, triggers entry into mitosis. G2-M kinases and phosphatases, such as cdc25 and Plk, play a crucial role in regulating the activation of the cyclin B-CDK1 complex.ref.8.30 ref.2.4 ref.9.34 NFκB signaling has been found to be involved in the transcriptional regulation of genes important for G2-M progression. The precise regulation of these molecular signals and regulatory mechanisms ensures the proper progression of the cell cycle and the accurate division of cells. Further research is needed to fully understand the intricate details of these regulatory processes and their implications in cell cycle control.ref.8.30 ref.8.2 ref.8.2

How are these signals transmitted and received by the cell?

Introduction

The regulation of the cell cycle is a complex process that ensures the proper ordering of cell cycle phases and the completion of each phase before the onset of the next. In order for cells to maintain their integrity and faithfully transmit hereditary information, the progression of the cell cycle must be tightly regulated. Failure to regulate cell cycle progression properly can lead to various disease states, such as cancer.ref.7.2 ref.7.2 ref.2.4 This essay will discuss the key mechanisms involved in the regulation of the cell cycle, including the interplay of cyclin-dependent kinases (CDKs) and cyclins, the role of checkpoints and regulatory mechanisms, and the importance of targeted proteolysis and transcriptional regulation.ref.9.4 ref.2.4 ref.7.2

Regulation of CDKs and Cyclins

One important mechanism in the regulation of the cell cycle is the interplay between CDKs and cyclins. CDKs are activated by cyclins, and the activity of CDK-cyclin complexes drives the progression of the cell cycle. The levels of cyclins and CDK inhibitors, which inhibit CDK activity, are regulated through various mechanisms.ref.9.4 ref.2.4 ref.15.1

CDK activity is regulated by association with specific regulatory subunits called cyclins. Additionally, CDK activity is regulated through phosphorylation and dephosphorylation events, as well as targeted proteolysis. The phosphorylation and dephosphorylation of CDKs can affect their activity and stability.ref.2.4 ref.15.1 ref.9.4 Furthermore, targeted proteolysis plays a crucial role in regulating the levels of cyclins and CDK inhibitors during cell cycle progression. Proteins such as Cdh1 and SCF are involved in the degradation of cyclins and CDK inhibitors, helping to regulate the timing and progression of the cell cycle.ref.9.7 ref.9.6 ref.9.4

Checkpoints and Regulatory Mechanisms

The proper ordering of cell cycle phases and the completion of each phase before the onset of the next are ensured through checkpoints and regulatory mechanisms. These checkpoints help maintain the fidelity of chromosome duplication and cell division.ref.2.4 ref.7.2 ref.7.2

One of the key checkpoints in the cell cycle is the G1-to-S checkpoint, which determines whether the cell is ready to enter DNA synthesis (S) phase. The G1-to-S transition requires the activity of CDKs, specifically CDKA and CDKB in plants, which directly drive the cell cycle transitions. Another important checkpoint is the G2-to-M checkpoint, which ensures that DNA replication is complete and that the cell is ready to enter mitosis (M) phase.ref.6.4 ref.14.6 ref.9.4 The G2-to-M transition involves the activation of cyclin B-CDK1 complexes and the regulation of various G2-to-M transition kinases and phosphatases.ref.8.30 ref.2.4 ref.6.4

The regulation of the cell cycle also involves the activation and deactivation cycles of cyclin-dependent kinases (Cdks) and cyclin counteracting phosphatases, the synthesis and degradation of regulatory cyclins, and the activation and deactivation cycles of checkpoints at specific cell cycle stages. The sequential activation of cyclins and proteolytic degradators is crucial for the progression of the cell cycle. Damage-induced pathways activate cytokine- and growth factor-induced pathways, which lead to the activation of cyclins D, E, A, and B in a sequential manner, allowing for the progression through different phases of the cell cycle.ref.2.4 ref.9.4 ref.7.2

Transcriptional Regulation

The transcriptional regulation of cyclins and CDK inhibitors during cell cycle progression involves multiple mechanisms. Transcriptional regulation plays a role in controlling the expression of cyclins and CDK inhibitors. For example, the E2F transcription factor 1 (E2F) is involved in regulating the sequential activation of cyclins.ref.9.4 ref.8.30 ref.2.4 Additionally, the forkhead family of transcription factors, including FOXM1 and FoxO, have been implicated in the expression of genes important for mitotic entry and exit.ref.8.30 ref.2.5 ref.8.30

Conclusion

In conclusion, the regulation of the cell cycle is a complex process that involves the interplay of various proteins and signaling pathways. The progression of the cell cycle is tightly regulated to ensure the proper ordering of cell cycle phases and the completion of each phase before the onset of the next. CDKs and cyclins play a crucial role in driving the progression of the cell cycle, and their activity and expression are tightly regulated in a cell cycle-dependent manner.ref.9.4 ref.7.2 ref.15.1 Additionally, checkpoints and regulatory mechanisms help maintain the fidelity of chromosome duplication and cell division. Targeted proteolysis and transcriptional regulation also play important roles in regulating the levels of cyclins and CDK inhibitors during cell cycle progression. The regulation of the cell cycle is crucial for maintaining the integrity of the genome and ensuring the faithful transmission of hereditary information.ref.7.2 ref.2.4 ref.9.4

Spindle Assembly

How are the microtubules organized to form the mitotic spindle?

Interphase Microtubules and Cell Polarity

During interphase, microtubule bundles play a critical role in maintaining cell polarity. They deposit factors required for the nucleation of actin filaments at the cell tips, ensuring proper cell shape and organization. Additionally, plus end microtubule polymerization exerts transient forces that help position the nucleus in the center of the cell.ref.65.1 ref.65.2 ref.91.16

Actin Redistribution and Cytokinetic Actomyosin Ring

As cells enter mitosis, actin is redistributed from the cell tips to the medial cell cortex to form the cytokinetic actomyosin ring. This coincides with the disappearance of interphase microtubules and the formation of a mitotic spindle composed of interpolar microtubules. The actin redistribution process involves several steps, such as the deposition of factors by interphase microtubule bundles for actin filament nucleation at cell tips.ref.65.2 ref.65.1 ref.29.6

Mitotic Spindle Structure and Function

The mitotic spindle is a complex structure composed of different types of microtubules and protein complexes. It consists of interpolar microtubules that overlap in a central zone and originate from two spindle pole bodies (SPBs). The SPBs are embedded in opposite sides of a persistent nuclear envelope.ref.86.4 ref.28.5 ref.65.2 Additional microtubules originate from each SPB and terminate at the kinetochores, which are protein structures on the chromosomes.ref.65.2 ref.87.47 ref.86.4

Phases of Mitotic Spindle

The mitotic spindle goes through three distinct phases: prophase, metaphase, and anaphase. During prophase, a short spindle is formed, preparing the cell for chromosome segregation. In metaphase, the spindle maintains its length, and the centromeres of the chromosomes make frequent and rapid movements between the poles.ref.65.2 ref.65.3 ref.87.2 Finally, in anaphase, the spindle elongates along the longitudinal axis of the cell, facilitating chromosome segregation.ref.65.3 ref.65.2 ref.86.7

Kinetochore Microtubules and Non-Kinetochore Microtubules

The spindle microtubules can be separated into kinetochore microtubules (K-MTs) and non-kinetochore microtubules (nK-MTs), each with distinct properties and functions. K-MTs are parallel bundles of microtubules that engage with kinetochores and are involved in the attachment and movement of chromosomes during mitosis. They remain relatively stable and do not undergo depolymerization at the microtubule minus-end.ref.86.4 ref.91.16 ref.86.6 On the other hand, nK-MTs are highly dynamic and undergo depolymerization at the microtubule minus-end. They contribute to poleward microtubule flux within the spindle and separation of centrosomes. Both K-MTs and nK-MTs play crucial roles in spindle assembly and chromosome segregation.ref.86.4 ref.91.16 ref.87.48

Mitotic Kinesins and Spindle Self-Organization

Mitotic kinesins, such as KIF11 and KIF15, are key members of the force-generating teams in the spindle. They interact with dynamic microtubules, walking directionally along them, and play essential roles in spindle self-organization and chromosome segregation. These kinesins anchor, crosslink, align, and sort microtubules into polarized bundles.ref.91.1 ref.91.16 ref.91.3 They also influence microtubule dynamics by interacting with microtubule tips. The mechanisms of these kinesins are specialized to allow each type to make a specific contribution to spindle self-organization and chromosome segregation.ref.91.1 ref.91.3 ref.91.1

Medial Microtubule-Organizing Centers (MTOCs)

The medial microtubule-organizing centers (MTOCs) are crucial components of the mitotic spindle. They consist of the nuclear envelope (NE) and the spindle pole bodies (SPBs). The NE maintains the integrity of the nucleus during mitosis, while the SPBs duplicate on the outer surface of the NE and function as MTOCs.ref.86.12 ref.65.2 ref.86.4 The SPBs organize the mitotic spindle by originating and terminating microtubules. They ensure the proper attachment of chromosomes to the spindle and facilitate chromosome segregation. The specific mechanisms by which MTOCs function as nuclear attachment sites and the forces powering chromosome segregation and spindle elongation are areas of ongoing research.ref.65.2 ref.86.4 ref.86.12

Role of Spindle Pole Bodies (SPBs)

The spindle pole bodies (SPBs) play a crucial role in the formation and organization of the mitotic spindle. During anaphase A, sister chromatids separate and move back to the SPBs. In anaphase B, the spindle elongates along the longitudinal axis of the cell, and the cytoplasmic face of the SPBs is associated with astral microtubules.ref.65.3 ref.65.2 ref.28.5 These astral microtubules maintain a fixed angle to the spindle axis and aid in spindle alignment by pushing the SPBs away from the cell cortex. Additionally, the SPBs are involved in the capture of microtubules by kinetochores, leading to proper chromosome biorientation during metaphase. The fidelity of chromosome biorientation is ensured by the spindle assembly checkpoint (SAC), which includes proteins such as Mad1, Mad2, Mad3, Bub1, Bub3, and Mps1.ref.65.3 ref.65.3 ref.65.3 The SPBs contribute to the organization and dynamics of the mitotic spindle, facilitating chromosome movement and segregation.ref.65.3 ref.86.13 ref.65.2

In conclusion, the mitotic spindle is a complex structure composed of different types of microtubules and protein complexes. Interphase microtubules play a critical role in maintaining cell polarity and nucleating actin filaments at cell tips. Actin redistribution from cell tips to the medial cell cortex leads to the formation of the cytokinetic actomyosin ring during mitosis.ref.65.2 ref.86.4 ref.65.1 The mitotic spindle, organized by medial microtubule-organizing centers (MTOCs), consists of interpolar microtubules and microtubules originating from spindle pole bodies (SPBs) that terminate at kinetochores. Kinetochore microtubules (K-MTs) and non-kinetochore microtubules (nK-MTs) have different properties and functions in spindle assembly and chromosome segregation. Mitotic kinesins, such as KIF11 and KIF15, play essential roles in spindle self-organization and chromosome segregation by interacting with microtubules.ref.86.4 ref.65.2 ref.91.1 The SPBs contribute to spindle formation, organization, and dynamics, including chromosome movement and segregation. Ongoing research aims to uncover the specific mechanisms by which MTOCs function as nuclear attachment sites and the forces powering chromosome segregation and spindle elongation.ref.86.12 ref.86.13 ref.86.4

What are the proteins involved in spindle assembly and how do they contribute to spindle formation and stability?

Introduction

Spindle assembly is a complex process that is critical for proper cell division and chromosome segregation. It involves the coordinated action of various proteins and factors to ensure the formation of a functional spindle apparatus. The proteins involved in spindle assembly and their contributions to spindle formation and stability are still being studied and understood.ref.27.1 ref.20.22 ref.28.41 Some of the proteins that have been identified include kinesins, microtubule-associated proteins (MAPs), the condensin complex, and chromosomal passenger proteins. These proteins interact with each other and with microtubules to ensure proper spindle formation and function. In this essay, we will discuss the specific roles of MAPs, kinesins, and the condensin complex in spindle assembly and stability.ref.87.48 ref.26.2 ref.28.41

Microtubule-Associated Proteins (MAPs) and their Role in Stabilizing Microtubules

MAPs play crucial roles in stabilizing microtubules and ensuring proper spindle assembly during cell division. Several specific MAPs have been identified, each with its own unique role in microtubule stability and organization. One such MAP is γ-tubulin, which is essential for microtubule nucleation in plants and required for the organization of microtubule structures in interphase and cell division.ref.51.4 ref.55.5 ref.87.47

Kinesins, another class of MAPs, are motor proteins that participate in various biological processes, including the organization of spindle microtubules and chromosome segregation. Several kinesins have been identified to be required for the structure, assembly, and positioning of mitotic and meiotic spindles in animals and fungi. Kinesins contribute to spindle organization and dynamics in several ways.ref.87.48 ref.91.16 ref.87.48 They walk directionally along dynamic microtubules, anchor, crosslink, align, and sort microtubules into polarized bundles, and interact with microtubule tips to influence microtubule dynamics. Kinesins are key members of the force-generating teams in mitotic spindles, working collectively with dynein to drive reliable chromosome segregation and contribute to spindle self-organization.ref.91.1 ref.91.3 ref.87.48

Op18/Stathmin is another MAP that plays a role in spindle assembly by negatively regulating microtubule stability. Op18 is a microtubule-destabilizing protein that is negatively regulated by phosphorylation. In Xenopus egg extracts, Op18 is basally phosphorylated in interphase and hyperphosphorylated in mitosis in the presence of mitotic chromatin, indicating that factors on chromosomes may promote microtubule polymerization and spindle assembly by inactivating Op18.ref.26.2 ref.26.21 ref.26.2

MPS1 is a protein that may guide microtubule minus-end migration in meiosis and could be involved in spindle pole organization and attracting the minus-end of spindle microtubules before spindle assembly. PSS1, a Kinesin-1-like protein in rice, has microtubule-stimulated ATPase activity and is required for proper chromosome alignment and segregation in meiosis. While spindle morphology is only slightly affected in pss1 mutants, PSS1 may be involved in regulating chromosome movements along the spindles.ref.87.51 ref.87.50 ref.87.51 MAB1, a MATH-BTB domain protein in maize, is required for organizing microtubule spindles and nuclei positioning in meiosis II and the first mitotic division. It may act through the control of a spindle apparatus regulator(s).ref.87.52 ref.87.51 ref.87.51

The Role of the Condensin Complex in Spindle Assembly

The condensin complex plays a specific function in establishing chromosomal architecture necessary for proper spindle assembly and chromosome segregation. In Xenopus laevis egg extracts, immunodepletion of condensin inhibited microtubule growth and organization around chromosomes, reducing the percentage of sperm nuclei capable of forming spindles and causing defects in anaphase chromosome segregation. The condensin complex is required throughout anaphase for chromosome resolution and segregation.ref.27.1 ref.27.22 ref.27.1 It actively maintains chromosome organization, preventing decondensation and sister entanglements. Additionally, the condensin complex contributes to kinetochore organization and attachment to microtubules during anaphase. Inhibition of condensin function during anaphase inhibits chromosome segregation.ref.27.1 ref.27.26 ref.27.25 The disruption of condensin function does not dramatically inhibit compaction or longitudinal shortening of chromosomes, suggesting the involvement of other activities.ref.27.26 ref.27.22 ref.27.27

The condensin complex is involved in chromosome compaction and resolution, contributing to mitotic chromosome structure. It forms ring structures that generate chromosome supercoiling or cross-linking. The disruption of condensin function does not dramatically inhibit compaction or longitudinal shortening of chromosomes, suggesting the involvement of other activities.ref.33.18 ref.33.18 ref.27.27 The contribution of condensin to spindle assembly is indirect, promoting the localization and/or activity of other factors. The condensin complex is required for proper spindle assembly and chromosome segregation in Xenopus egg extracts. It establishes chromosomal architecture necessary for microtubule organization during spindle assembly.ref.27.27 ref.27.1 ref.27.19

Conclusion

In conclusion, spindle assembly is a complex process that involves the coordinated action of various proteins and factors. MAPs, including γ-tubulin, kinesins, Op18/Stathmin, MPS1, PSS1, and MAB1, play crucial roles in stabilizing microtubules and ensuring proper spindle assembly during cell division. Kinesins contribute to spindle organization and dynamics by walking along microtubules, anchoring and crosslinking them, and influencing microtubule dynamics.ref.91.16 ref.87.48 ref.28.37 The condensin complex is involved in establishing chromosomal architecture necessary for proper spindle assembly and chromosome segregation. It actively maintains chromosome organization and contributes to kinetochore organization and attachment to microtubules during anaphase. Further research is needed to fully understand the mechanisms of these proteins and their interactions with microtubules in spindle assembly and stability.ref.27.1 ref.27.5 ref.27.1

How is the positioning and orientation of the spindle regulated?

The Regulation of Spindle Positioning and Orientation

The establishment of cell polarity is essential for the regulation of spindle positioning and orientation. This process involves several events that need to occur. First, cell polarity must be established, specifying cortical regions that can capture the spindle.ref.75.9 ref.75.24 ref.75.9 Second, the spindle apparatus needs to be able to interact with the cortex, typically through astral microtubules nucleated by centrosomes at the spindle poles. This process relies on a conserved molecular machinery that includes cortical and microtubule binding proteins, as well as molecular motors that exert torque on the spindle. The core set of molecules involved in spindle positioning includes G alphai, LGN, Numa, and Dynein.ref.75.9 ref.75.13 ref.75.9 G alphai binds to the cortex, LGN and Numa link G alphai to microtubules and Dynein, and Dynein provides the forces required to orient the spindle.ref.75.9 ref.75.10 ref.75.10

In addition to intrinsic cues, extrinsic cues from the environment can also regulate spindle orientation. The Wnt/planar cell polarity (PCP) pathway has been shown to play a role in this regulation. Wnt signaling can bias centrosome segregation, and the transmembrane receptor Frizzled and its effector Dishevelled are involved in this process.ref.75.24 ref.75.23 ref.75.24 These proteins can interact with Mud/Numa, linking Wnt signaling to the spindle orientation machinery.ref.75.24 ref.75.10 ref.75.23

Furthermore, the orientation of cell division can be regulated by signaling events between cells. The Wnt/PCP pathway has been implicated in regulating spindle orientation, and the Wnt signaling pathway can polarize the activity of the anthrax toxin receptor 2a, which, in cooperation with RhoA, activates the formin zDia2. This activation leads to the generation of actin filaments that help orient the spindle.ref.75.24 ref.75.23 ref.75.24

Overall, the establishment of cell polarity, the involvement of intrinsic and extrinsic cues, and the regulation of centrosome segregation by signaling pathways all contribute to the regulation of spindle positioning and orientation.ref.75.9 ref.75.24 ref.75.9

The Role of Actin Cables and Signaling Pathways in Spindle Orientation

The Wnt/planar cell polarity (PCP) pathway has been identified as a key regulator of spindle orientation. The Wnt signaling pathway polarizes the activity of the anthrax toxin receptor 2a, which, in cooperation with RhoA, activates the formin zDia2. This activation leads to the generation of actin filaments that help orient the spindle.ref.75.24 ref.75.24 ref.75.24 The precise role of actin cables in spindle positioning is still being determined.ref.75.25 ref.75.24 ref.75.10

In Drosophila S2 cells, the localization of Dishevelled (Dsh) to restricted cortical regions recruits the actin-binding protein Canoe/Afadin. This recruitment activates Rho signaling and contributes to spindle orientation.ref.75.25 ref.75.24 ref.75.25

The core set of molecules involved in spindle positioning includes G alphai, LGN, Numa, and Dynein. G alphai can be myristoylated and binds to the cortex. This binding regulates the activity of Pins by increasing its affinity for Mud.ref.75.10 ref.75.9 ref.75.10 Pins/LGN binds Mud/Numa, which can interact with cytoplasmic Dynein. Dynein exerts forces to orient the spindle. These molecules function in anchoring and positioning the spindle.ref.75.10 ref.75.9 ref.75.13 They also play important roles in directing spindle orientation in different contexts, such as progenitor cells in the mouse neocortex and symmetric divisions in developing epithelia. The proteins involved function similarly in different contexts. However, the specific role of spindle orientation in the outcome of progenitor/stem cell division varies and is not fully understood in many progenitor cells.ref.75.10 ref.75.9 ref.75.24

The Notch signaling pathway may also play a role in regulating centrosome and spindle behavior. However, the molecular details of how exposure to Wnt regulates the orientation of mitotic spindles are not well understood. The precise role of actin cables and the signaling pathways involved in spindle orientation are still being investigated.ref.75.24 ref.75.24 ref.75.26

The Significance of Regulating Spindle Orientation for Tissue Morphogenesis and Stem Cell Division

Regulating spindle orientation is crucial for tissue morphogenesis and stem cell division as it plays a crucial role in determining cell fate decisions and maintaining tissue homeostasis. The orientation of the mitotic spindle during cell division can result in either symmetric or asymmetric cell divisions, which can have different outcomes in terms of cell fate and tissue development.ref.75.9 ref.71.4 ref.75.24

In asymmetric cell divisions, the orientation of the spindle determines which daughter cell inherits specific cellular components or signaling molecules, leading to different cell fates. This process is important for tissue morphogenesis and the generation of diverse cell types during development.ref.75.9 ref.71.4 ref.75.15

Additionally, the regulation of spindle orientation is essential for stem cell division as it can influence the balance between self-renewal and differentiation. The correct orientation of the spindle is crucial for maintaining the stem cell pool and generating the appropriate number and types of daughter cells.ref.75.9 ref.71.4 ref.75.24

Various intrinsic and extrinsic cues, as well as molecular mechanisms, are involved in regulating spindle orientation and cell fate decisions. These include the involvement of centrosomes, microtubules, cortical and microtubule binding proteins, and molecular motors. However, the exact signaling pathways and mechanisms involved in regulating spindle orientation and cell fate decisions are still being investigated.ref.75.9 ref.75.24 ref.75.10

The Interaction Between the Spindle Apparatus and the Cortex Through Astral Microtubules

The interaction between the spindle apparatus and the cortex through astral microtubules is a complex process that involves various mechanisms. In some systems, cortical pulling forces exerted on astral microtubules represent the major mechanism for outward-directed forces on the spindle poles.ref.28.42 ref.28.62 ref.28.52

This mechanism is observed in C. elegans embryos, where cortical pulling forces generated by a combination of astral microtubule depolymerization and dynein motors walking towards the minus ends of astral microtubules drive spindle elongation and spindle positioning.ref.28.51 ref.28.62 ref.28.52

In other systems, such as fission yeast, the role of astral microtubules in spindle elongation is less clear. While cortical pulling is the dominant mechanism driving spindle elongation in fission yeast, experiments involving centrosome ablation have revealed the existence of a normally cryptic, redundant mechanism. This mechanism involves interpolar microtubule plus end polymerization at the spindle midzone generating an outward force that drives spindle elongation and chromosome segregation during anaphase.ref.28.49 ref.28.52 ref.28.62

Furthermore, recent studies have shown that spindle orientation is primarily determined by interphase microtubules, rather than astral microtubules, in some systems. The precise timing and mechanisms of spindle elongation and chromosome segregation can vary between different organisms and even within different cells of the same organism.ref.65.6 ref.28.54 ref.18.15

Overall, the interaction between the spindle apparatus and the cortex through astral microtubules is a dynamic and complex process that involves multiple mechanisms and can vary depending on the specific system being studied.ref.28.42 ref.91.28 ref.91.3

What are the mechanisms that ensure proper attachment of chromosomes to the spindle microtubules?

Role of Active Error Correction Mechanisms in Chromosome Attachment

Active error correction mechanisms are essential for ensuring proper attachment of chromosomes to the spindle microtubules and destabilizing inappropriate attachments. These mechanisms serve to destabilize incorrect attachments, such as both sister kinetochores bound to a single pole, and promote the formation of bi-oriented attachments where sister kinetochores attach to microtubules emanating from opposite spindle poles. The process of attachment correction involves the movement of chromosomes to the spindle equator during prometaphase, a phase known as congression.ref.86.7 ref.86.7 ref.58.1 Additionally, selective stabilization of bi-oriented attachments is crucial for accurate chromosome segregation during anaphase.ref.86.7 ref.92.21 ref.65.41

During prometaphase, chromosomes undergo congression, which involves the movement of chromosomes towards the spindle equator. This movement is facilitated by the interaction between kinetochores, which are protein structures located on the centromeres of chromosomes, and microtubules. The kinetochores capture microtubules from both spindle poles, resulting in bi-oriented chromosomes.ref.86.7 ref.65.3 ref.65.2 These bi-oriented chromosomes then move towards the metaphase plate, which is the equatorial plane where they align. The movement of chromosomes to the spindle equator is significant for spindle assembly because it ensures proper alignment of chromosomes and accurate chromosome segregation during anaphase.ref.65.3 ref.86.7 ref.65.2

In addition to the movement of chromosomes, kinetochores also play a role in mediating the spindle assembly checkpoint (SAC). The SAC is a surveillance mechanism that senses incorrect attachments and generates a soluble signal that inhibits anaphase onset until all sister kinetochores are correctly attached. The exact nature of the signal that the kinetochore senses and how it is satisfied remain unknown.ref.86.7 ref.86.8 ref.63.2 However, it is believed that the signal reflects the occupancy of microtubule attachment sites and/or the imposition of tension or force balance across sister kinetochores.ref.65.41 ref.86.7 ref.25.1

Forces and Mechanisms Involved in Chromosome Congression

The movement of chromosomes to the spindle equator during prometaphase, known as congression, is influenced by various forces and mechanisms. These forces act to balance poleward and anti-poleward forces on chromosomes, resulting in their positioning at the equator.ref.86.7 ref.18.14 ref.18.15

One important force involved in chromosome congression is the interaction between kinetochores and microtubules. The kinetochores capture microtubules from both spindle poles, creating a tension that pulls the chromosomes towards the equator. This tension is crucial for the alignment of chromosomes at the metaphase plate.ref.86.7 ref.92.4 ref.21.2

Another force involved in chromosome congression is the polar ejection force. This force is generated by motor proteins and acts to push chromosomes away from the spindle poles towards the equator. The polar ejection force helps to counteract the poleward forces and facilitates the movement of chromosomes towards the equator.ref.18.14 ref.18.15 ref.91.24

Motor activities on chromosomes also contribute to chromosome congression. Motor proteins, such as dynein and kinesin, exert forces on chromosomes that influence their movement and positioning at the equator. Dynein, for example, acts in an anti-poleward direction and helps to maintain the balance of forces during congression.ref.18.15 ref.91.33 ref.91.24

The organization of spindle microtubules also plays a role in the spatial distribution of forces involved in chromosome congression. The spatial arrangement of microtubules and their interactions with kinetochores and motor proteins determine the direction and magnitude of forces acting on chromosomes.ref.86.7 ref.91.28 ref.18.15

Despite the progress made in understanding the forces and mechanisms involved in chromosome congression, the process is not fully understood. Further research is needed to elucidate the molecular mechanisms underlying chromosome congression and to uncover additional forces and factors that contribute to this process.

Theories on the Nature of the Signal and its Satisfaction in Chromosome Attachment

The nature of the signal that the kinetochore senses and how it is satisfied to ensure proper attachment of chromosomes to the spindle microtubules is the subject of current theories and hypotheses. Two prominent models, the attachment model and the tension model, propose different mechanisms for the initiation of anaphase and the satisfaction of the spindle assembly checkpoint.ref.65.5 ref.86.7 ref.86.8

The attachment model suggests that anaphase is initiated when all potential attachment sites on kinetochores are occupied by spindle microtubules. According to this model, attachment is necessary for the application of tension, and tension is required for the checkpoint to be satisfied. In other words, the presence of microtubules at all potential attachment sites generates tension across sister kinetochore pairs, which signals that the chromosomes are properly attached and aligned.ref.65.5 ref.65.41 ref.86.7

On the other hand, the tension model proposes that the checkpoint is only satisfied when tension is applied across sister kinetochore pairs. This model suggests that the checkpoint signal may be amplified by the recruitment of checkpoint proteins to kinetochores. Tension is generated when microtubules exert forces on kinetochores, and this tension is required to satisfy the checkpoint.ref.65.5 ref.65.41 ref.86.7 When microtubules become detached from kinetochores, the tension is lost, and the checkpoint signal is blocked.ref.65.41 ref.65.5 ref.86.8

It is important to note that these models are based on computational modeling and experimental data. While they provide valuable insights into the nature of the signal and its satisfaction in spindle assembly, further research is needed to fully understand the molecular mechanisms involved. Future studies may involve the use of advanced imaging techniques, genetic manipulations, and biochemical assays to investigate the dynamics of kinetochore-microtubule interactions and the regulation of the spindle assembly checkpoint.ref.25.0 ref.91.28 ref.91.28

In conclusion, active error correction mechanisms are crucial for destabilizing inappropriate attachments and ensuring the proper attachment of chromosomes to the spindle microtubules. The movement of chromosomes to the spindle equator during prometaphase, known as congression, and the selective stabilization of bi-oriented attachments are central to this process. Various forces, including kinetochore-microtubule interactions, polar ejection forces, and motor activities on chromosomes, contribute to chromosome congression.ref.86.7 ref.86.7 ref.64.3 The nature of the signal that the kinetochore senses and how it is satisfied in spindle assembly are still the subject of ongoing research and are the focus of the attachment and tension models. Further studies are needed to fully understand the molecular mechanisms involved in chromosome attachment and to uncover additional factors that contribute to this process.ref.86.7 ref.86.8 ref.86.7

How is the tension across the spindle regulated?

Mechanisms of Mitotic Motors in Generating Force within the Spindle

The document excerpts discuss the mechanisms of mitotic motors in generating force within the spindle. The interactions of mitotic motors with dynamic microtubules play a crucial role in spindle self-organization and faithful chromosome segregation. Each mitotic motor has a mechanism for generating force, and this mechanism responds to external forces in a predictable, measurable way.ref.91.28 ref.91.1 ref.91.1

The force-velocity curve of a mitotic motor defines how it generates force. The document suggests that understanding the force-velocity curves of individual motors is important for comprehending spindle self-organization and faithful chromosome segregation. The force-velocity curve describes how the motor's force generation is affected by the velocity at which it is moving.ref.91.28 ref.91.28 ref.91.28 As the velocity of the motor increases, the force it can generate decreases, and vice versa.ref.91.28 ref.91.28 ref.91.28

In vitro biophysical approaches are needed to measure local forces in simplified subsystems and describe the intrinsic mechanisms of individual force generators. These approaches contribute to our understanding of spindle self-organization and chromosome segregation by providing insights into the mechanical design principles of the spindle and the forces involved in spindle elongation. For example, theoretical models combined with computer simulations and structural analysis have proposed that the compressive strength of elongating spindles is optimized to support the drag forces that resist spindle elongation.ref.91.28 ref.91.28 ref.91.2 The optimization is achieved through the crosslinking of interpolar microtubules (ipMTs) into rigid, paracrystalline arrays with square and hexagonal symmetry within and outside the central midzone, respectively.ref.28.51 ref.28.26 ref.28.52

Live cell microscopy approaches are also required to dissect the positions and motions of individual kinesin molecules and teams of kinesin molecules in the spindle. These experiments help in understanding the interactions of various mechanisms with dynamic microtubules, thus producing robust structure and function at the level of the entire spindle.ref.91.28 ref.91.3 ref.28.51

The document mentions the use of in vivo force sensors to measure local forces in the intact spindle. These sensors provide valuable information about the forces involved in spindle self-organization and chromosome segregation.ref.91.28 ref.91.28 ref.91.28

Explicit models can be created to predict collective behavior by determining the characteristic mechanical behavior of each mitotic kinesin. These models can be tested and refined by mutating the motors to alter their performance and measuring their altered performance curves in vitro and in living cells.ref.91.28 ref.91.2 ref.91.2

Importance of Force-Velocity Curves in Spindle Assembly

The force-velocity curve plays a vital role in spindle assembly and understanding how spindle self-organization and faithful chromosome segregation occur.ref.91.28 ref.91.28 ref.91.28

The force-velocity curve determines how the motor responds to external forces. If the external force is greater than the force the motor can generate at a given velocity, the motor will slow down or stall. Conversely, if the external force is smaller than the force the motor can generate at a given velocity, the motor will continue to move at that velocity.

Understanding the force-velocity curves of individual motors is crucial for understanding spindle self-organization and chromosome segregation. The generation of force by mitotic motors and their response to external forces are key factors in the proper functioning of the spindle during cell division.ref.91.28 ref.91.28 ref.91.1

Contributions of In Vitro Biophysical Approaches to Understanding Spindle Self-Organization and Chromosome Segregation

In vitro biophysical approaches provide valuable insights into the mechanical design principles of the spindle and the forces involved in spindle elongation, thus contributing to our understanding of spindle self-organization and chromosome segregation.ref.91.28 ref.28.57 ref.91.28

Theoretical models combined with computer simulations and structural analysis have proposed that the compressive strength of elongating spindles is optimized to support the drag forces that resist spindle elongation. This optimization is achieved through the crosslinking of interpolar microtubules (ipMTs) into rigid, paracrystalline arrays with square and hexagonal symmetry within and outside the central midzone, respectively.ref.28.18 ref.28.52 ref.28.26

In vitro studies using extracts from Xenopus laevis eggs have shown that chromatin-associated kinases, such as Plx1, play a role in regulating microtubule stabilization and spindle assembly. These studies provide insights into the biophysical mechanisms underlying spindle self-organization and chromosome segregation.ref.26.21 ref.26.1 ref.26.22

In conclusion, the mechanisms of mitotic motors in generating force within the spindle are essential for spindle self-organization and faithful chromosome segregation. Each mitotic motor has a mechanism for generating force, and this mechanism responds to external forces in a predictable, measurable way defined by its force-velocity curve. In vitro biophysical approaches and live cell microscopy approaches are both required to understand how the interactions of these various mechanisms with dynamic microtubules produce robust structure and function at the level of the entire spindle.ref.91.28 ref.91.28 ref.91.1 In vitro reconstitution approaches are needed to measure local forces in simplified subsystems and describe the intrinsic mechanisms of individual force generators, while live cell experiments are needed to dissect the positions and motions of individual kinesin molecules and teams of kinesin molecules in the spindle. The use of in vivo force sensors can measure local forces in the intact spindle, providing further insights. By determining the characteristic mechanical behavior of each mitotic kinesin, explicit models can be created to predict collective behavior.ref.91.28 ref.91.28 ref.91.28 These models can be tested and refined by mutating the motors to alter their performance and measuring their altered performance curves in vitro and in living cells. Overall, a comprehensive understanding of how the molecular mechanisms of individual spindle motors interact with each other and with regulated microtubule dynamics is necessary to understand spindle self-organization and chromosome segregation.ref.91.28 ref.91.28 ref.91.28

Chromosome Condensation

What are the molecular events that lead to chromosome condensation in mitosis?

The Role of Chromatin Condensation in Chromosome Segregation during Mitosis

Chromatin condensation is a critical process in cell division that facilitates the proper segregation of chromosomes during mitosis. The alternation between a condensed and decondensed state of chromatin is necessary for this process. During cell division, chromatin transitions from a condensed state to facilitate chromosome segregation, and then to a decondensed form when DNA replicates.ref.32.2 ref.31.27 ref.32.3 In most tissues, the S phase (DNA replication) and mitosis are separated by defined G1 and G2 gap phases. However, in early embryogenesis, there are rapid oscillations between replication and mitosis.ref.32.1 ref.32.3 ref.32.2

Replicated DNA is required for chromosome condensation and the concentration of condensin II on chromosomal axes. Condensin II is a key component of the condensin complex, which consists of SMC (structural maintenance of chromosomes) proteins and non-SMC proteins. The condensin complex plays a crucial role in establishing and maintaining the condensed state of chromosomes.ref.32.3 ref.37.1 ref.27.2 During late telophase, replication initiates on condensed chromosomes, promoting the rapid decondensation of the chromatin.ref.32.3 ref.32.33 ref.32.2

The CDC-45-MCM-GINS (CMG) DNA helicase is responsible for driving the release of condensin I complexes from chromatin. It also activates or displaces inactive MCM-2-7 complexes. MEL-28/ELYS, a nucleoporin, tethers condensed chromatin to the nuclear envelope, contributing to chromatin decondensation.ref.32.36 ref.32.36 ref.32.35 This highlights the functional link between the chromosome-condensation cycle and DNA replication in early embryos.ref.32.2 ref.32.37 ref.32.17

The Functions and Activities of SMC Subunits within the Condensin Complexes

The condensin complexes, composed of SMC proteins and non-SMC proteins, have diverse functions and activities related to chromosome condensation. SMC proteins play multiple roles in chromosome organization and function, including sister cohesion, dosage compensation, and recombination-mediated repair.ref.27.2 ref.37.2 ref.36.28

The condensin complex can reconfigure DNA structure in an ATP hydrolysis-dependent manner. In vertebrates, condensin I and II localize at centromeres and along the axis of metaphase sister chromatid arms. This localization supports the idea that condensin acts as a central scaffold network that is important for the architecture of mitotic chromosomes.ref.37.2 ref.37.32 ref.37.2

The condensin complex is essential for chromosome segregation and architecture in all eukaryotes. Depletion of condensin can result in defects in chromosome compaction and anaphase segregation. It is required throughout mitosis to promote spindle formation and ensure proper chromosome segregation.ref.33.18 ref.27.13 ref.27.1

Furthermore, condensin complexes are non-randomly distributed across chromosomes and often found at the boundaries of topologically associating domains (TADs) within chromosome territories. This suggests a role in transcriptional regulation and global chromosomal organization. Additionally, the condensin complex is involved in anaphase chromosome resolution, further highlighting its importance in maintaining chromosomal integrity.ref.36.28 ref.33.18 ref.37.2

The Role of Chromosome Associated Proteins (CAPs) in Chromosome Condensation

Chromosome associated proteins (CAPs) contribute to chromosome condensation and are essential for the establishment and maintenance of the condensed state of chromosomes. In vertebrates, there are two condensin complexes: condensin I and condensin II. These complexes have distinct roles in mitotic chromosome structure.ref.37.1 ref.27.2 ref.37.30

Condensin I consists of CAP-D2, CAP-G, and CAP-H, while condensin II consists of CAP-D3, CAP-G2, and CAP-H2. CAP-D3, a component of condensin II, is primarily responsible for providing rigidity to the chromosomes by establishing an initial chromosome axis. On the other hand, condensin I arranges loops of chromatin around this axis.ref.37.30 ref.37.30 ref.37.7

Depletion of condensin II results in chromosomes that are more stretched and lack axial rigidity, impairing proper chromosome segregation. In contrast, depletion of condensin I leads to wider and shorter chromosomes with a diffuse scaffold. These observations highlight the complementary roles of the condensin complexes in compacting the chromatin and ensuring accurate chromosome segregation during cell division.ref.37.31 ref.37.33 ref.37.32

In conclusion, chromatin condensation is a dynamic process essential for proper chromosome segregation during mitosis. The alternation between a condensed and decondensed state of chromatin is facilitated by the condensin complexes, which play crucial roles in establishing and maintaining the condensed state of chromosomes. The SMC subunits within the condensin complexes have diverse functions and activities related to chromosome organization and function.ref.37.1 ref.31.27 ref.32.3 Additionally, chromosome associated proteins (CAPs) contribute to chromosome condensation and play distinct roles in mitotic chromosome structure. The condensin complexes and CAPs work together to compact the chromatin and ensure accurate chromosome segregation during cell division. Further research into the mechanisms underlying chromatin condensation will deepen our understanding of the fundamental processes that govern chromosome dynamics and integrity.ref.37.1 ref.32.3 ref.37.30

How are the condensin complexes involved in chromosome compaction?

Introduction

The condensin complexes play a crucial role in chromosome condensation and segregation during mitosis. These multisubunit ATPase complexes establish and maintain an ordered loop structure in mitotic chromosomes, driving self-compaction in a cell cycle-dependent manner. The exact mechanism of condensin function is still actively debated, but it is clear that condensin complexes bring together distant segments of the same chromosome, generating mitotic loops.ref.33.18 ref.33.18 ref.21.2 This process of loop formation is supported by evidence of condensin localization to the chromosome axis, where loops often appear to be anchored. Additionally, condensin action leads to DNA supercoiling of the mitotic chromosome fibers, which drives the resolution of sister chromatid intertwines. The condensin complexes play distinct roles in mitotic chromosome structure, with condensin I mediating more frequent short-range lateral interactions among chromatin loops, and condensin II mediating axial stacking of the laterally assembled configurations.ref.33.18 ref.37.31 ref.33.18

Role of Condensin in Chromosome Condensation

Establishment and Maintenance of Loop Structures

The condensin complexes are responsible for establishing and maintaining an ordered loop structure in mitotic chromosomes. This loop structure is essential for chromosome compaction. One model proposes that condensin complexes bring together two distant segments of the same chromosome, generating a mitotic loop.ref.33.18 ref.33.18 ref.37.1 This structural model is consistent with the preferred localization of condensin to the chromosome axis, where loops often appear to be anchored. Supporting this model, in HeLa cells, condensin II accumulates on chromatin during S phase and promotes the separation of replicated DNA, suggesting its role in bringing together distant segments of the same chromosome during chromosome condensation.ref.33.18 ref.37.1 ref.37.31

DNA Supercoiling and Resolution of Sister Chromatid Intertwines

Another model suggests that condensin action leads to DNA supercoiling of the mitotic chromosome fibers, which exposes inter-chromatid linkages to the outside of chromatid masses, facilitating their resolution by topoisomerase II. DNA supercoiling induced by condensin complexes drives the resolution of sister chromatid intertwines, ensuring proper chromosome segregation during anaphase. The absence of condensin results in defective chromosome segregation and compromised chromosome compaction.ref.33.18 ref.33.18 ref.37.32 In Xenopus egg extracts, the absence of condensin II results in more robust and obvious anaphase bridges, while the absence of condensin I leads to highly attenuated bridges that are only seen after careful examination. These defects in chromosome compaction and segregation observed in the absence of condensin can lead to excessive chromatin bridge formation and a cut phenotype during cytokinesis.ref.37.29 ref.37.1 ref.27.27

Cooperating with Other Factors in Chromosome Condensation

The condensin complexes cooperate with other factors to promote chromosome condensation. They work together with the chromokinesin KIF4 to promote lateral compaction of chromatid arms. Additionally, other factors such as topoisomerase II and histone modifications also contribute to chromosome compaction and resolution.ref.21.2 ref.27.26 ref.37.29 The combination of intra-chromosomal compaction provided by chromatin organization and condensin action is sufficient for the resolution of most sister chromatid intertwines in cells.ref.33.19 ref.37.29 ref.33.19

Role of Condensin in Chromosome Segregation

Spindle Formation and Chromosome Segregation

The condensin complexes are required throughout mitosis to promote spindle formation and ensure proper chromosome segregation. Depletion of condensin leads to defects in chromosome compaction and anaphase segregation. The absence of condensin prevents chromosomes from being disentangled during anaphase, leading to a failure to fully segregate mitotic chromosomes.ref.27.1 ref.27.11 ref.27.23 Condensin-depleted chromosomes are more compromised in terms of their three-dimensional architecture compared to chromosomes depleted of condensin II. The defects in chromosome compaction and segregation observed in the absence of condensin significantly impact the ability of the mitotic spindle to form and inhibit chromosome segregation during anaphase.ref.27.11 ref.27.23 ref.37.29

Variations in Condensin Function

The contribution of condensin to mitotic chromosome condensation varies between different organisms. More severe defects in chromosome compaction and segregation are observed in Xenopus and Drosophila compared to C. elegans.ref.27.28 ref.27.2 ref.33.18 This suggests that the exact role and requirement of condensin complexes may vary across different species. Further studies are needed to fully understand the role of condensin complexes in chromosome compaction and segregation.ref.27.28 ref.27.27 ref.33.18

Conclusion

In conclusion, the condensin complexes play a crucial role in chromosome compaction and segregation during mitosis. They establish and maintain an ordered loop structure in mitotic chromosomes, driving self-compaction in a cell cycle-dependent manner. The condensin complexes bring together distant segments of the same chromosome, generating mitotic loops.ref.33.18 ref.33.18 ref.37.32 Additionally, condensin action leads to DNA supercoiling of the mitotic chromosome fibers, which drives the resolution of sister chromatid intertwines. The condensin complexes are required throughout mitosis to promote spindle formation and ensure proper chromosome segregation. Defects in condensin function result in defective chromosome compaction and anaphase segregation, leading to compromised chromosome architecture and excessive chromatin bridge formation.ref.33.18 ref.27.1 ref.33.18 Further studies are needed to fully understand the exact mechanism of condensin function and its role in chromosome compaction and segregation.ref.27.27 ref.33.18 ref.37.32

What are the roles of histones and other chromatin-associated proteins in chromosome condensation?

The Roles of Histone H1 and CENP-A in Chromosome Condensation

Histones and other chromatin-associated proteins play crucial roles in the condensation of chromosomes during mitosis. One such protein, histone H1, has been found to contribute to the architecture of chromosome arms and is necessary for proper mitotic chromosome condensation (1). Additionally, the histone H3 variant CENP-A is enriched at centromeres and may confer structural rigidity to centromeric chromatin, defining and maintaining functional centromeres (2).ref.21.17 ref.21.4 ref.21.2 The presence of histone H1 and CENP-A at specific locations along the chromatin template helps define the structurally and functionally distinct domains of arm and centromeric chromatin (2). These findings suggest that histone H1 and CENP-A play important roles in establishing the structural environments of chromosome arms and centromeres during chromosome condensation.ref.21.24 ref.21.24 ref.21.17

Histone H1 is involved in defining chromosome arm architecture and contributes to chromosome compaction (2). Depletion of histone H1 in Xenopus laevis egg extracts resulted in the assembly of aberrant elongated chromosomes that could not be properly segregated in anaphase (3). While histone H1 depletion did not significantly affect the recruitment of known structural or functional chromosomal components, such as condensins or chromokinesins, it did affect chromosome alignment and segregation (3).ref.21.23 ref.21.1 ref.21.4 Histone H1 is required for proper mitotic chromosome compaction, alignment, and segregation (3). The deposition of condensation factors, such as condensin, on the chromatin template is likely defined at the level of DNA kilobases rather than the physical length of the template (1). Further quantitative comparison of interphase chromatin lengths and condensation factor levels in control and H1-depleted chromosomes can help distinguish between different models of chromosome condensation (3).ref.21.23 ref.21.11 ref.21.20

CENP-A, on the other hand, is enriched at centromeres and may confer structural rigidity to centromeric chromatin, defining and maintaining functional centromeres (2). The distinct domains of arm and centromeric chromatin are likely defined by the local deposition of histone H1 and CENP-A (2). These findings suggest that H1 and CENP-A play important roles in establishing the structural environments of chromosome arms and centromeres during chromosome condensation.ref.21.24 ref.21.24 ref.21.17

The Roles of Condensin I and II in Chromosome Architecture

Condensin I and condensin II are ATPase complexes that shape the architecture of mitotic chromosomes (1). Condensin I mediates short-range lateral interactions among chromatin loops, promoting compaction of chromosome rosettes, while condensin II mediates axial stacking of the laterally assembled configurations (1). In the absence of condensin II, chromosomes fail to compact axially, becoming bent and twisted, while in the absence of condensin I, chromatids show fine chromatin fibers associated with failure of cytokinesis and cell death (4).ref.37.31 ref.37.31 ref.37.30

Super-resolution microscopy studies have revealed that condensin-I-depleted mitotic chromosomes are wider and shorter, with a diffuse chromosome scaffold, whereas condensin-II-depleted chromosomes retain a more defined scaffold, with chromosomes more stretched and seemingly lacking in axial rigidity (4). CAP-D3, a subunit of condensin II, knockout results in masses of chromatin-containing anaphase bridges, while CAP-H, a subunit of condensin I, knockout anaphases have a more subtle defect (4). These observations indicate that the two condensin complexes play distinct roles in mitotic chromosome structure, with condensin I mediating more frequent short-range lateral interactions among chromatin loops, and condensin II mediating axial stacking of the laterally assembled configurations (4).ref.37.1 ref.37.30 ref.37.18

The exact mechanisms of chromosome condensation and the interplay among histones, condensin I, and condensin II are still being studied. However, it is clear that histone H1, CENP-A, condensin I, and condensin II all contribute to the proper architecture and compaction of mitotic chromosomes during condensation.ref.37.29 ref.37.30 ref.21.2

The Deposition of Condensation Factors on the Chromatin Template

The deposition of condensation factors, such as condensin, on the chromatin template is determined by the presence of replicated DNA and the spacing of DNA-defined intervals (5). The condensation machinery can be loaded onto interphase chromatin either at intervals defined by the physical length of the template or at specific DNA-defined intervals (5). However, factors such as condensin are deposited at DNA-defined intervals rather than the physical length of the template (5).ref.21.23 ref.21.23 ref.21.23

The presence of condensin is required throughout anaphase for proper chromosome segregation (5). The condensin complex actively maintains chromosome organization and prevents decondensation and sister entanglements (5). It is also required for spindle formation and proper kinetochore attachment and movement within the spindle (5).ref.27.1 ref.27.13 ref.27.12

The deposition of chromatin condensation factors at defined DNA intervals is supported by biophysical studies and chromosome immunoprecipitation experiments (5). Understanding the mechanisms governing the deposition and function of condensation factors on the chromatin template is an ongoing area of research.ref.21.23 ref.21.23 ref.32.1

In conclusion, histones, such as histone H1 and the histone H3 variant CENP-A, and condensin complexes, such as condensin I and condensin II, play crucial roles in chromosome condensation. Histone H1 contributes to the architecture of mitotic chromosomes by playing a role in chromosome compaction and arm architecture. CENP-A is involved in defining and maintaining functional centromeres.ref.21.24 ref.37.29 ref.21.2 Condensin I mediates short-range lateral interactions among chromatin loops, while condensin II mediates axial stacking of the laterally assembled configurations. The deposition of condensation factors on the chromatin template is determined by the presence of replicated DNA and the spacing of DNA-defined intervals. Further research is needed to fully understand the exact mechanisms and interplay among these factors in chromosome condensation.ref.33.18 ref.37.30 ref.37.30

How is the timing and coordination of chromosome condensation regulated with other mitotic events?

Role of CDC-45-MCM-GINS (CMG) DNA helicase in promoting chromatin decondensation during late telophase

During late telophase, the CDC-45-MCM-GINS (CMG) DNA helicase plays a crucial role in promoting chromatin decondensation. It achieves this by driving the release of condensin I complexes from chromatin and the activation or displacement of inactive MCM-2-7 complexes. The nucleoporin MEL-28/ELYS also contributes to chromatin decondensation by interacting with MCM-2–7 complexes.ref.32.36 ref.32.36 ref.32.35 The initiation of DNA replication, which is dependent on both nuclear assembly and CDC-45, further promotes the dissociation of MCMs from MEL-28, allowing for the rapid decondensation of chromatin once replication has initiated.ref.32.31 ref.32.36 ref.32.32

The CMG helicase also plays a role in focusing replication initiation on highly condensed chromatin, potentially aiding rapid and synchronous replication. Additionally, the active replicative helicase may induce the release of chromatin-bound complexes that maintain condensation, thereby promoting decondensation. It is important to note that the specific mechanism by which CMG promotes chromatin decondensation during the late telophase stage is not provided in the document.ref.32.35 ref.32.36 ref.32.23 However, it is suggested that DNA unwinding by the CMG helicase could displace condensins from DNA and allow decondensation to occur.ref.32.36 ref.32.35 ref.32.36

Mechanism of nucleoporin MEL-28/ELYS in tethering condensed chromatin to the nuclear envelope

The nucleoporin MEL-28/ELYS plays a critical role in tethering condensed chromatin to the nuclear envelope. This is achieved through its interaction with MCM-2–7 complexes. During DNA replication, MCM-2–7 complexes dissociate from MEL-28, leading to the rapid decondensation of chromatin.ref.32.36 ref.32.31 ref.32.32 The activation or displacement of inactive MCM-2–7 complexes by the CDC-45-MCM-GINS (CMG) DNA helicase drives this process. The release of condensin I complexes from chromatin also contributes to chromatin decondensation. It is important to note that the document does not provide specific information on the precise mechanism by which MEL-28/ELYS tethers condensed chromatin to the nuclear envelope.ref.32.36 ref.32.36 ref.32.35

Regulation of cell-cycle progression and coordination of mitotic events

The regulation of cell-cycle progression and coordination of mitotic events involve several checkpoint mechanisms. These mechanisms ensure that critical events, such as DNA replication and chromosome alignment, are completed before proceeding to the next stage of the cell cycle. Changes in cyclin-dependent kinase (CDK) kinase activity and cyclin association drive major transitions in the cell cycle, such as the initiation of S phase, mitosis, and the segregation of chromatids.ref.7.2 ref.9.4 ref.2.4

The progression of chromosome condensation and the concentration of condensin II on chromosomal axes require replicated DNA. During late telophase, replication initiates on condensed chromosomes and promotes the rapid decondensation of chromatin. The CDC-45-MCM-GINS (CMG) DNA helicase plays a crucial role in this process by driving the release of condensin I complexes from chromatin and the activation or displacement of inactive MCM-2-7 complexes.ref.32.17 ref.32.2 ref.32.35 The nucleoporin MEL-28/ELYS also contributes to chromatin decondensation by tethering condensed chromatin to the nuclear envelope. The regulation of chromosome condensation is functionally linked with DNA replication in an early embryo.ref.32.36 ref.32.32 ref.32.33

Role of condensin in chromosome condensation during mitosis

Condensin is a protein complex that is essential for proper chromosome segregation and compaction during mitosis. Its function involves generating positively supercoiled chromatin loops and preventing chromosome stretching and distortion in response to spindle forces. Changes in cyclin-dependent kinase (CDK) kinase activity and cyclin association regulate the timing and coordination of chromosome condensation.ref.33.18 ref.27.1 ref.37.1 CDKs and Dbf4-dependent kinases activate the Mcm2-7 helicase, which promotes the interaction between the helicase and other proteins involved in chromosome condensation.ref.32.4 ref.33.18 ref.32.3

The association of condensin with chromosomes is crucial for spindle formation and proper chromosome segregation. Depletion of condensin subunits results in chromosome segregation defects. The condensin complex is required throughout mitosis to generate a chromosome structure that allows sister chromatid resolution, kinetochore orientation, and prevents chromosome stretching and distortion.ref.27.1 ref.27.23 ref.27.27

Relationship between DNA replication and chromosome condensation

The relationship between DNA replication and chromosome condensation has been studied by depleting the licensing factor MCM-7, which inhibits DNA replication. The results showed that chromosome condensation occurs abnormally when DNA replication is inhibited. This suggests that DNA replication plays a role in the regulation of chromosome condensation.ref.32.8 ref.32.1 ref.32.35

Fate of transcripts during mitosis

The fate of transcripts during mitosis is stable, and RNA molecules remain present throughout all stages of mitosis. This indicates that the presence of RNA is maintained, even as the cell undergoes chromosome condensation and other mitotic events. Further research is needed to understand the specific mechanisms that contribute to the stability and regulation of transcripts during mitosis.ref.40.2 ref.40.2

In conclusion, the CDC-45-MCM-GINS (CMG) DNA helicase plays a crucial role in promoting chromatin decondensation during late telophase by driving the release of condensin I complexes from chromatin and the activation or displacement of inactive MCM-2-7 complexes. The nucleoporin MEL-28/ELYS also contributes to chromatin decondensation by tethering condensed chromatin to the nuclear envelope. The regulation of cell-cycle progression and coordination of mitotic events involve multiple checkpoint mechanisms and changes in cyclin-dependent kinase (CDK) kinase activity and cyclin association.ref.32.36 ref.32.35 ref.32.36 Condensin is essential for proper chromosome segregation and compaction during mitosis, preventing chromosome stretching and distortion. The relationship between DNA replication and chromosome condensation is evident, as inhibition of DNA replication leads to abnormal chromosome condensation. The fate of transcripts during mitosis remains stable, with RNA molecules present throughout all stages of mitosis.ref.32.33 ref.32.3 ref.32.2 Further research is needed to fully understand the mechanisms underlying these processes.ref.32.35 ref.32.36 ref.32.33

What are the mechanisms that ensure accurate segregation of condensed chromosomes during mitosis?

Mechanisms of Chromosome Segregation during Mitosis

During mitosis, accurate segregation of condensed chromosomes is crucial for proper cell division. Several processes are involved in ensuring this accurate segregation. One important factor is the condensin complex, which is responsible for chromosome condensation and resolution.ref.33.18 ref.37.1 ref.21.2 The condensin complex consists of two subtypes, condensin I and condensin II, both of which contribute to chromatid condensation and resolution. Disruption of condensin function can have significant effects on chromosome compaction and segregation.ref.33.18 ref.37.1 ref.33.18

Additionally, topoisomerase II plays a role in higher order chromosome architecture during mitosis. It promotes the shortening of the chromatid axes and acts in opposition to the actions of condensins and the chromokinesin KIF4. Topoisomerase II also contributes to chromatid condensation and resolution.ref.34.2 ref.21.2 ref.34.19 The absence of condensin II, for example, results in chromosomes that fail to compact axially and become bent and twisted. On the other hand, the absence of condensin I leads to wider laterally and shorter axially chromosomes. These observations highlight the importance of condensins and topoisomerase II in regulating chromosome architecture and ensuring accurate chromosome segregation during mitosis.ref.37.31 ref.37.30 ref.37.32

Process of Chromosome Condensation and Segregation during Mitosis

The process of chromosome condensation and segregation during mitosis is a dynamic and autonomous multi-step process. It involves several stages, including aggregation, condensation, and segregation. Initially, during interphase, the chromatin aggregates into distinct chromosomal filaments through a transversal condensation process.ref.31.27 ref.31.27 ref.31.26 This aggregation of chromatin sets the stage for subsequent chromosome condensation and segregation.ref.31.26 ref.31.27 ref.31.23

Condensation progression to metaphase is a uniform process along the chromosome arms, resulting in a reduction in length and an increase in diameter. The condensin complex, along with topoisomerase II, plays a crucial role in establishing and maintaining the condensed state of the chromosomes. The condensin complex is responsible for establishing and maintaining an ordered loop structure in mitotic chromosomes.ref.37.1 ref.31.27 ref.37.29 This loop structure drives self-compaction in a cell cycle-dependent manner. One proposed model suggests that condensin complexes bring together distant segments of the same chromosome, generating a mitotic loop. This structural model aligns with the observed localization of condensin to the chromosome axis.ref.33.18 ref.33.18 ref.37.1

In addition to the structural role of condensins, there is evidence that their action leads to DNA supercoiling of the mitotic chromosome fibers. This supercoiling is essential for resolving sister chromatid intertwinings and ensuring proper chromosome segregation during anaphase. Disruption of condensin function can result in a failure to fully segregate mitotic chromosomes and a defect in intra-chromosomal compaction.ref.33.18 ref.33.18 ref.37.32 However, the exact mechanism by which condensins function and contribute to chromosome compaction and segregation is still a subject of debate and requires further investigation.ref.33.18 ref.37.32 ref.27.26

Chromosome Condensation and Segregation during Meiosis

In meiosis, the process of chromosome condensation is similar to mitosis but with some differences. Meiotic chromosomes undergo linear condensation, and the process follows a sigmoidal pattern. This sigmoidal pattern involves retardation in early prophase and acceleration from prophase to metaphase I.ref.31.21 ref.31.24 ref.31.3 The extreme condensation of chromosomes in metaphase is important for proper segregation of meiosis I and II.ref.31.27 ref.31.24 ref.31.26

Sister chromatid cohesion is also crucial for accurate segregation during meiosis. The cohesion of sister chromatids forms a single "structural" chromosome in rye during meiosis I. This cohesion ensures that the homologous chromosomes segregate correctly during meiosis.ref.31.26 ref.31.25 ref.87.2 The condensin complex, along with other factors involved in chromosome condensation and resolution, plays a vital role in establishing the necessary chromosome architecture for proper segregation during meiosis.ref.21.2 ref.31.26 ref.33.19

Regulation of Chromosome Segregation during Mitosis

The mechanisms involved in the accurate segregation of condensed chromosomes during mitosis are regulated by cell-specific and chromosome region-specific factors. The condensin complex, topoisomerase II, and the processes of aggregation, condensation, and segregation are all regulated by these factors. Furthermore, posttranslational modifications of histones and other chromosomal proteins likely contribute to the regulation of mitotic chromatin compaction.ref.21.2 ref.21.2 ref.31.27

The condensin complex, in particular, plays multiple crucial roles in mitotic chromosome structure and segregation. It is involved in anaphase chromosome resolution and is required for proper chromosome segregation. The condensin complex contributes to the lateral compaction of chromatid arms and is mutually dependent on the chromokinesin KIF4 for dynamic localization on the chromatid axes.ref.33.18 ref.27.27 ref.21.2 It also plays a role in spindle assembly and function, promoting microtubule growth and organization around chromosomes. The condensin complex is essential throughout mitosis to ensure proper chromosome segregation and spindle formation.ref.27.27 ref.27.1 ref.27.5

In addition to the condensin complex, topoisomerase II is involved in regulating higher order chromosome architecture during mitosis. By promoting the shortening of chromatid axes and acting in opposition to the actions of condensins and KIF4, topoisomerase II contributes to chromatid condensation and resolution. The coordination of these various factors and processes is necessary to achieve accurate segregation of condensed chromosomes during mitosis.ref.34.1 ref.34.2 ref.21.2

In conclusion, the accurate segregation of condensed chromosomes during mitosis is a complex process that involves several mechanisms and factors. The condensin complex, topoisomerase II, and the processes of aggregation, condensation, and segregation all contribute to ensuring the proper compaction and segregation of chromosomes. These processes are regulated by cell-specific and chromosome region-specific factors.ref.21.2 ref.31.27 ref.31.26 Further research is needed to fully understand the exact mechanisms by which these factors and processes function and interact to achieve accurate chromosome segregation during mitosis.ref.21.2 ref.31.27 ref.31.26

Cytokinesis

What are the key molecular events that drive cytokinesis?

Introduction to Cytokinesis

Cytokinesis is a fundamental process in cell division that involves the division and separation of cytoskeletal microtubules and the plasma membrane. This process is essential for the formation of two daughter cells with equal genetic material. In order to achieve successful cytokinesis, a series of tightly regulated molecular events take place, including the ingression of a cleavage furrow, the division and separation of microtubules and the plasma membrane, and the remodeling of crosslinks between microtubules and the plasma membrane.ref.44.18 ref.44.3 ref.55.4 These events are coordinated by various molecular regulators and proteins, which are still being investigated to fully understand the precise mechanisms and signal transduction pathways involved in cytokinesis.ref.55.4 ref.7.2 ref.7.2

Microtubule Severing and Increase in Microtubule Number

A. Role of Katanin, Spastin, and Fidgetin Katanin, spastin, and fidgetin are members of the AAA ATPase family and play important roles in cytokinesis by severing microtubules and increasing the overall number of microtubules. Katanin is a heterodimer consisting of a regulatory p80 subunit and a catalytic p60 subunit.ref.44.4 ref.44.3 ref.44.6 It is involved in substrate recognition and targeting of the catalytic subunit. Spastin localizes to centrosomes and spindle poles and is involved in endosomal trafficking. Fidgetin, on the other hand, did not yield any discernible phenotype when depleted.ref.44.6 ref.44.4 ref.44.7 These proteins are crucial for the cleavage and remodeling of the subpellicular microtubule cytoskeleton during cytokinesis in trypanosomes.ref.44.20 ref.44.1 ref.44.21

Trypanosomes have a vermiform shape, with parallel microtubules running longitudinally. During cytokinesis in trypanosomes, the cleavage furrow ingresses along the longitudinal axis of the cell, causing the plasma membrane to invaginate and crosslinks between the microtubules to be broken. To maintain the shape of the daughter cells, microtubules need to be severed and shortened or re-formed on either side of the cleavage plane.ref.44.18 ref.44.18 ref.44.3 In post-mitotic procyclic cells, an increase in new, short, tyrosinated microtubules is observed, suggesting that microtubule severing may aid the synthesis of new microtubules by increasing the pool of free tubulin subunits. Crosslinks between the microtubules and the plasma membrane are also likely to be remodeled during furrow ingression. The identification of proteins such as KAT80, KAT60a, KAT60b, KAT60c, and SPA as essential for cytokinesis in bloodstream trypanosomes provides important insights into how the division of the cytoskeleton is accomplished.ref.44.18 ref.44.20 ref.44.24

Chromatin Decondensation and Nuclear Envelope Dynamics

A. Role of CDC-45-MCM-GINS DNA Helicase The CDC-45-MCM-GINS (CMG) DNA helicase is involved in replication initiation, which promotes the rapid decondensation of chromatin during cytokinesis. The dissociation of MCMs from the nucleoporin MEL-28/ELYS allows for the rapid decondensation of chromatin once replication has initiated.ref.32.36 ref.32.5 ref.32.35 Loss of MEL-28 rescues the chromosome-condensation defect seen upon CDC-45 depletion, indicating that MEL-28 chromatin relocalization contributes to chromosome decondensation. The coupling of DNA replication, chromosome decondensation, and condensation ensures the correct order of cell-cycle events and the faithful propagation of chromosomal DNA.ref.32.31 ref.32.32 ref.32.37

The interplay between the nuclear envelope and the mitotic spindle is crucial in cytokinesis. The spindle, driven by cytoplasmic dynein, causes the deformation and rupture of the nuclear membranes, leading to the release of condensed chromosomes into the cytoplasm. This process serves to delay nuclear envelope breakdown until functional spindle microtubules have been assembled.ref.64.6 ref.64.3 ref.86.9 Nup358, a nucleoporin, has been suggested as a possible dynein-binding partner on the nuclear envelope, but its involvement in dynein binding and nuclear envelope breakdown is still under investigation. Furthermore, Nup358 also plays a role in kinetochore function and chromatid segregation. The kinetochores mediate the spindle assembly checkpoint, which ensures that sister kinetochores are correctly attached before anaphase onset.ref.64.6 ref.64.21 ref.64.3 The relationship between membranes, nuclear pores, and kinetochores is crucial for generating the forces necessary for chromosome segregation during mitosis. Further investigation of these processes in non-model organisms can provide valuable insights into conserved mechanisms and alternative evolutionary approaches.ref.86.2 ref.86.19 ref.86.19

Conclusion

Cytokinesis is a complex process that involves the division and separation of cytoskeletal microtubules and the plasma membrane. The key molecular events that drive cytokinesis are tightly regulated and involve various molecular regulators and proteins. Katanin, spastin, and fidgetin, members of the AAA ATPase family, play important roles in severing microtubules and increasing the overall number of microtubules.ref.44.3 ref.55.4 ref.44.18 The CDC-45-MCM-GINS DNA helicase is involved in replication initiation and chromatin decondensation. The nucleoporin MEL-28/ELYS tethers condensed chromatin to the nuclear envelope, facilitating chromatin decondensation. The interplay between the nuclear envelope and the mitotic spindle, as well as the involvement of nucleoporins in dynein binding and kinetochore function, are important aspects of cytokinesis.ref.86.8 ref.17.1 ref.17.1 Further research is needed to fully elucidate the precise mechanisms and signal transduction pathways involved in cytokinesis and to gain a deeper understanding of the mechanics and effectors of cell division in different organisms.ref.55.4 ref.44.18 ref.44.3

How is the contractile ring formed and regulated?

The Formation and Regulation of the Contractile Ring during Cytokinesis

The formation and regulation of the contractile ring during cytokinesis involve several molecular mechanisms. In the context of an epithelium, the assembly of a mechanically stable metaphase cortex depends on the RhoGEF Pbl/Ect2. Pbl/Ect2 induces a lateral shift in the distribution of polarity regulators Cdc42, aPKC, and Par6, which leads to the assembly of a relatively isotropic Diaphanous-dependent actomyosin cytoskeleton.ref.29.4 ref.29.1 ref.29.0 This actomyosin cytoskeleton is critical for mitosis and cell division in a crowded tissue environment. The exact nucleators involved in mitotic actin filament assembly are still unclear, but Dia homologs are likely to play a key role.ref.29.5 ref.29.25 ref.29.1

One well-understood actin nucleator is the Arp2/3 complex. The Arp2/3 complex, downstream of the RhoGEF Pbl/Ect2, is involved in the remodeling of the actomyosin cortex at mitotic entry. Another key actin nucleator is Dia, which is required for the assembly of the mitotic actin cortex.ref.29.5 ref.29.1 ref.29.8 Dia, like the Arp2/3 complex, is downstream of the RhoGEF Pbl/Ect2. The regulation of cytokinesis also involves other proteins such as katanin, spastin, and fidgetin, which influence microtubule dynamics. However, the specific mechanisms of contractile ring formation and regulation are still being studied, and further research is needed to fully understand the process.ref.29.26 ref.29.8 ref.29.1

The Role of the Diaphanous-dependent Actomyosin Cytoskeleton in the Assembly of a Mechanically Stable Metaphase Cortex

The Diaphanous-dependent actomyosin cytoskeleton plays a crucial role in the assembly of a mechanically stable metaphase cortex during cytokinesis. The assembly of a stable actomyosin-rich lateral cortex is observed in cells during metaphase. This assembly is dependent on the broad-specificity RhoGEF Pbl/Ect2.ref.29.1 ref.29.4 ref.29.8 Pbl/Ect2 induces a lateral shift in the distribution of polarity regulators Cdc42, aPKC, and Par6, which leads to the assembly of a relatively isotropic Diaphanous-dependent actomyosin cytoskeleton.ref.29.0 ref.29.1 ref.29.24

The Diaphanous-dependent actomyosin cytoskeleton is the critical actin nucleator required for the generation of the metaphase cortex. Pbl/Ect2 activates Rho, which in turn induces a switch from Arp2/3 to Diaphanous-mediated cortical actin nucleation. This switch is essential for the assembly of the mitotic actin cortex during cytokinesis.ref.29.1 ref.29.8 ref.29.0 The Diaphanous-dependent actomyosin cytoskeleton drives the assembly of the mitotic cortex, which is necessary for successful cell division in a crowded tissue environment.ref.29.8 ref.29.1 ref.29.4

The Roles of the Arp2/3 Complex and Dia in the Remodeling of the Actomyosin Cortex at Mitotic Entry

Both the Arp2/3 complex and Dia contribute to the remodeling of the actomyosin cortex at mitotic entry. The Arp2/3 complex is involved in the assembly of the mitotic actin cortex, while Dia is required for the formation of the mitotic actin cortex. The Arp2/3 complex is responsible for nucleating actin filaments, while Dia is a key actin nucleator that drives the assembly of the mitotic cortex.ref.29.5 ref.29.25 ref.29.24

The Arp2/3 complex and Dia have distinct roles in the remodeling of the actomyosin cortex during mitotic entry. The Arp2/3 complex, downstream of the RhoGEF Pbl/Ect2, is involved in the remodeling process. Dia, also downstream of Pbl/Ect2, is required for the assembly of the mitotic actin cortex.ref.29.5 ref.29.1 ref.29.8 Together, these two actin nucleators contribute to the dynamic remodeling of the actomyosin cortex during cytokinesis.ref.29.1 ref.29.25 ref.29.5

The Role of Katanin, Spastin, and Fidgetin in Regulating Microtubule Dynamics during Cytokinesis

Katanin, spastin, and fidgetin play significant roles in regulating microtubule dynamics during cytokinesis. Katanin is involved in redistributing c-tubulin during mitosis and is required for severing the flagellum from the basal bodies before mitosis in certain organisms. Spastin localizes to centrosomes and spindle poles during cell division and is also involved in regulating axon branching.ref.44.5 ref.44.4 ref.44.3 Mutations in the spastin gene are associated with Hereditary Spastic Paraplegia, a condition characterized by the degeneration of corticospinal tracts. Fidgetin's exact role in cytokinesis is still unclear, but it localizes to the nucleus and may be involved in spindle formation during mitosis.ref.44.6 ref.44.4 ref.44.7

These proteins, katanin, spastin, and fidgetin, belong to the AAA ATPase family and sever microtubules along their length, shortening them and increasing the overall number of microtubules. Katanin is a heterodimer consisting of a regulatory p80 subunit and a catalytic p60 subunit. Spastin and fidgetin have high homology to katanin p60 but do not interact with a regulatory subunit.ref.44.4 ref.44.4 ref.44.6 These proteins can oligomerize into hexameric rings in the presence of ATP, stimulating their ATPase activity and creating a central pore into which the C-terminal tail of tubulin is pulled, breaking the microtubule.ref.44.4 ref.91.16 ref.87.48

In trypanosomes, katanins, spastin, and fidgetin are involved in cytokinesis. Depletion of these proteins in bloodstream trypanosomes resulted in distinct phenotypes, indicating their non-redundant and essential functions in cytokinesis. KAT60a may be important for converting the cleavage cleft to a furrow, while KAT60b is involved in cleaving the cytoskeleton at the anterior of the cell.ref.44.1 ref.44.0 ref.44.20 KAT60c likely plays multiple roles in cytokinesis. The exact mechanisms and targets of these proteins in trypanosome cytokinesis are still not fully understood.ref.44.20 ref.44.25 ref.44.29

Conclusion

In conclusion, the formation and regulation of the contractile ring during cytokinesis involve several molecular mechanisms. The assembly of a mechanically stable metaphase cortex depends on the RhoGEF Pbl/Ect2, which induces a lateral shift in the distribution of polarity regulators. This shift leads to the assembly of a Diaphanous-dependent actomyosin cytoskeleton, critical for successful cell division in a crowded tissue environment.ref.29.25 ref.29.1 ref.29.4 The Arp2/3 complex and Dia contribute to the remodeling of the actomyosin cortex at mitotic entry, with distinct roles in the assembly of the mitotic actin cortex. Katanin, spastin, and fidgetin play crucial roles in regulating microtubule dynamics during cytokinesis, with each protein having distinct functions at different stages of the process. Further research is needed to fully understand the specific mechanisms and targets of these proteins in contractile ring formation and regulation during cytokinesis.ref.29.1 ref.29.5 ref.29.25

What are the proteins involved in membrane ingression and vesicle fusion during cytokinesis?

Introduction

Cytokinesis is the process by which cells physically divide into two daughter cells. It is a complex and highly regulated process that involves the coordination of various cellular and molecular events. In trypanosomes, a group of single-celled parasites, cytokinesis is particularly intricate and involves the cleavage and remodeling of the subpellicular microtubule cytoskeleton.ref.44.18 ref.55.2 ref.55.4 This process is facilitated by several proteins, including KAT80, KAT60a, KAT60b, KAT60c, and SPA. These proteins play essential roles in membrane ingression and vesicle fusion during cytokinesis. In this essay, we will explore the functions of these proteins and their contributions to the cleavage and remodeling of the subpellicular microtubule cytoskeleton during cytokinesis in trypanosomes.ref.44.18 ref.44.0 ref.44.20

Cleavage and Remodeling of the Subpellicular Microtubule Cytoskeleton during Cytokinesis

The process of cleavage and remodeling of the subpellicular microtubule cytoskeleton during cytokinesis in trypanosomes is crucial for the division of the cell. This process involves the action of several proteins, including KAT80, KAT60a, KAT60b, KAT60c, and SPA. These proteins contribute to the severing and shortening of microtubules on either side of the cleavage plane, allowing for the maintenance of the vermiform shape in the daughter cells.ref.44.18 ref.44.20 ref.44.0

Microtubule severing proteins, such as katanin and spastin, play distinct roles at different stages of cytokinesis. They may have different microtubule cleavage specificities based on differences in microtubule post-translational modifications and interactions with microtubule-binding proteins. The process of microtubule severing is important for increasing the pool of free tubulin subunits and aiding in the synthesis of new microtubules.ref.44.21 ref.44.18 ref.44.20 Additionally, crosslinks between the microtubules and the plasma membrane are likely to be remodeled during cytokinesis. However, the exact mechanisms by which these proteins contribute to the cleavage and remodeling of the subpellicular microtubule cytoskeleton during cytokinesis are still not fully understood.ref.44.18 ref.44.18 ref.44.3

Functions of the Proteins during Cytokinesis

A. Katanins Katanins are microtubule severing enzymes that regulate microtubule dynamics. They sever microtubules along their length, shortening them and increasing the overall number of microtubules.ref.44.4 ref.44.21 ref.44.4 This helps in remodeling the cytoskeleton during cytokinesis. The katanin p60 subunits, including KAT60a, KAT60b, and KAT60c, were found to be enzymatically active and functional in the absence of p80 katanin (KAT80). The role of KAT80 in promoting furrow ingression suggests that it may interact with the KAT60 proteins, although direct interactions have not been detected so far.ref.44.24 ref.44.24 ref.44.25

Spastin is another microtubule severing enzyme that influences microtubule dynamics. It also severs microtubules along their length, leading to their shortening and an increase in the number of microtubules. Spastin plays an essential and non-redundant role in cytokinesis in Trypanosoma brucei.ref.44.6 ref.44.0 ref.44.5

Fidgetin is a member of the AAA ATPases family and is involved in microtubule dynamics. However, its specific function during cytokinesis is not mentioned in the provided document excerpts.

Cyclins are proteins that regulate the activity of cyclin-dependent kinases (CDKs) during the cell cycle. In the context of cytokinesis, A-type cyclins, such as CYCA1;2/TAM, have been shown to have a meiotic function in Arabidopsis. Knockout plants of CYCA1;2/TAM exhibit defects in homolog chromosome pairing and crossover formation during prophase I of meiosis.ref.2.37 ref.15.11 ref.15.11 B-type cyclins, such as CYCB3;1, have prominent roles during mitosis and contribute to the spatial and temporal regulation of cell wall formation in pollen mother cells.ref.2.37 ref.2.37 ref.2.5

MAPs are important for cytokinesis, and their depletion or overexpression can result in abnormal cell cycle progression. For example, the depletion of WCB and CAP5.5/CAP5.5V MAPs or the overexpression of CAP15 and CAP17 MAPs can lead to aberrant cell types and furrowing during cytokinesis. However, the specific functions of these MAPs during cytokinesis are not mentioned in the provided document excerpts.ref.55.5 ref.44.24 ref.51.5

APC/C is a ubiquitin ligase complex that regulates mitotic progression and cytokinesis. During meiosis I, APC/C destroys the separase inhibitor SECURIN to allow the proper progression from metaphase I to anaphase I. The specific functions of APC/C subunits in cytokinesis are not mentioned in the provided document excerpts.ref.57.1 ref.58.2 ref.59.2

Lysosomes and autophagic vesicles are present and active during cell division. Lysosomes decrease in number and increase in size from prophase to telophase, and they are involved in the degradation of cellular components during cytokinesis. Autophagy and lysosome-dependent degradation play a role in mitotic progression and can prevent chromosome mis-segregation.ref.80.8 ref.80.7 ref.80.5

Mutations and Abnormalities in Proteins and their Effects on Cytokinesis

There are known mutations or abnormalities in proteins that can lead to disruptions in cytokinesis. Depletion or overexpression of certain MAPs, such as WCB, CAP5.5/CAP5.5V, CAP15, and CAP17, can result in abnormal cell cycle progression and aberrant cell types during cytokinesis. Additionally, a family of microtubule severing proteins is required for cytokinesis.ref.55.5 ref.44.24 ref.52.18 Depletion of TbAIR9, a protein involved in cytokinesis, can lead to defects in kinetoplast positioning and cytokinesis in trypanosomes. Mutations in CENP-A and hMis12, kinetochore proteins, can also result in aberrant cytokinesis. Furthermore, loss of CITK (Citron Kinase) can lead to disruptions in cytokinesis and DNA damage accumulation.ref.92.13 ref.52.18 ref.48.3 Mutations in CitK can cause cytokinesis failure and an increase in DNA double-strand breaks (DSBs).ref.48.3 ref.48.3 ref.48.5

Conclusion

In conclusion, the proteins KAT80, KAT60a, KAT60b, KAT60c, and SPA play crucial roles in membrane ingression and vesicle fusion during cytokinesis in trypanosomes. These proteins are involved in the cleavage and remodeling of the subpellicular microtubule cytoskeleton, contributing to the severing and shortening of microtubules, as well as the remodeling of crosslinks between microtubules and the plasma membrane. These proteins are non-redundant and have distinct roles at different stages of cytokinesis.ref.44.18 ref.44.1 ref.44.20 The exact mechanisms by which these proteins contribute to cytokinesis are still not fully understood, and further research is needed to unravel the intricacies of this process.ref.44.21 ref.44.20 ref.44.20

How is the completion of cytokinesis coordinated with other events in mitosis?

Coordination of Cytokinesis with Other Events in Mitosis

The completion of cytokinesis, the final stage of cell division, is a tightly regulated process that is coordinated with other events in mitosis. This coordination is essential to ensure the proper ordering of cell-cycle phases and the fidelity of cell division. Several regulatory mechanisms are involved in this coordination.ref.7.2 ref.2.4 ref.7.2

One key regulatory mechanism is the activation and deactivation cycles of cyclin-dependent kinases (Cdks) and cyclin counteracting phosphatases. Cdks form functional heterodimers with regulatory cyclin subunits and control cell cycle progression at the G1-to-S and G2-to-M phase transitions. Through the phosphorylation of target proteins involved in DNA replication and mitosis, Cdks regulate the transitions within the cell cycle.ref.2.4 ref.9.4 ref.15.1 The activity of Cdks is regulated at multiple levels, including the interaction with inhibitors or scaffolding proteins, phosphoregulation of the Cdk subunit, and the association with specific regulatory subunits called cyclins. The activation and deactivation cycles of Cdks are also regulated by phosphorylation and dephosphorylation, interaction with inhibitory proteins, and targeted proteolysis. Additionally, the activity of Cdks is controlled by cyclin degradation through the ubiquitin-dependent proteolysis system.ref.2.4 ref.15.1 ref.14.5 These regulatory mechanisms ensure the proper ordering of cell-cycle phases and the completion of each phase before the onset of the next.ref.7.2 ref.7.2 ref.2.4

The activation and deactivation cycles of checkpoints also play a role in coordinating the different phases of the cell cycle, including cytokinesis. Checkpoints are regulatory mechanisms that monitor the integrity of the genome and ensure that each phase of the cell cycle is completed correctly before the cell progresses to the next phase. The activation and deactivation cycles of these checkpoints are crucial for the coordination of cytokinesis with other events in mitosis.ref.7.2 ref.2.4 ref.7.2 They help ensure the fidelity of cell division and prevent the occurrence of genomic instability.ref.2.4 ref.7.2 ref.14.12

Positive feedback is another regulatory mechanism that is involved in coordinating cytokinesis with other events in mitosis. Positive feedback functions as a molecular mechanism to ensure the temporal insulation of mitosis from variability in earlier cell-cycle phases. It helps keep the duration of mitosis short, constant, and insulated.ref.7.22 ref.7.37 ref.7.35 This is achieved through the activation of positive feedback loops, which amplify and maintain the activation of Cdk1-cyclin B1 complexes. The activation of Cdk1-cyclin B1 complexes triggers the onset of mitosis and the subsequent events of mitotic progression. The positive feedback loops involving Cdc25, Wee1, and Cdk1-cyclin B1 contribute to the bistability of Cdk1 activation, ensuring a switch-like behavior.ref.7.19 ref.7.40 ref.7.22 This short and constant duration of mitosis is crucial for the fidelity of cell division and cell survival.ref.7.37 ref.7.0 ref.7.0

Positive feedback regulation also contributes to the coordination of cytokinesis by ensuring proper Cdk1 activation thresholds for the progression and exit from mitosis. The temporal insulation of mitosis is not dependent on the spindle assembly checkpoint (SAC) but is regulated by mitotic phosphatases, such as PP1 and PP2A, which reverse Cdk1 substrate phosphorylation. Inhibition of positive feedback compromises the short and constant duration of mitosis, leading to cell death during or shortly after mitosis.ref.7.22 ref.7.37 ref.7.37 Computational modeling supports the role of positive feedback in temporal insulation and modularity in mitosis. Overall, positive feedback is a crucial mechanism that ensures the precise timing and coordination of mitotic events and cytokinesis.ref.7.10 ref.7.35 ref.7.22

Specific Examples of Regulatory Cyclins

In addition to the regulatory mechanisms discussed above, the coordination of cytokinesis with other events in mitosis also involves specific regulatory cyclins. These cyclins form complexes with Cdks and promote progression through different phases of the cell cycle.ref.9.4 ref.7.2 ref.7.2

Cyclin D forms complexes with Cdks 4, 5, and 6 and is involved in the regulation of the G1 phase of the cell cycle. Cyclin D/Cdk4-6 complexes promote the phosphorylation of the retinoblastoma protein (Rb), leading to the release of E2F transcription factors and the activation of genes required for DNA replication. The activity of Cyclin D/Cdk4-6 complexes is regulated through various mechanisms, including transcriptional regulation, association with protein inhibitors, phosphorylation/dephosphorylation, and cyclin degradation.ref.9.4 ref.9.4 ref.2.5

Cyclin E forms complexes with Cdk2 and is involved in the transition from G1 to S phase. Cyclin E/Cdk2 complexes promote the initiation of DNA replication by phosphorylating proteins involved in DNA replication initiation, such as the Mcm complex. The activity of Cyclin E/Cdk2 complexes is regulated through similar mechanisms as Cyclin D/Cdk4-6 complexes.ref.9.4 ref.9.4 ref.9.34

Cyclin A forms complexes with Cdk2 and is involved in the progression in S and G2 phase of the cell cycle. Cyclin A/Cdk2 complexes promote the completion of DNA replication and the preparation for mitosis. The activity of Cyclin A/Cdk2 complexes is regulated through similar mechanisms as Cyclin D/Cdk4-6 and Cyclin E/Cdk2 complexes.ref.9.4 ref.9.4 ref.9.34

Cyclin B forms complexes with Cdk1 and is involved in the G2/M phase transition, allowing entry into mitosis. Cyclin B/Cdk1 complexes promote the phosphorylation of proteins involved in the disassembly of the nuclear envelope, the condensation of chromosomes, and the formation of the mitotic spindle. The activity of Cyclin B/Cdk1 complexes is regulated through similar mechanisms as the other cyclin/Cdk complexes.ref.8.30 ref.9.4 ref.9.9

The regulation of these regulatory cyclin/Cdk complexes is crucial for the proper coordination of cytokinesis with other events in mitosis. Through their interaction with Cdks, these cyclins ensure the orderly progression through the cell cycle and the completion of each phase before the onset of the next. The synthesis and degradation of these regulatory cyclins, as well as the activation and deactivation cycles of Cdks and cyclin counteracting phosphatases, form a complex network that controls the progression of the cell cycle during cytokinesis.ref.9.4 ref.2.4 ref.7.2

Conclusion

In conclusion, the completion of cytokinesis is coordinated with other events in mitosis through various regulatory mechanisms. The activation and deactivation cycles of Cdks and cyclin counteracting phosphatases, as well as the synthesis and degradation of regulatory cyclins, play a crucial role in regulating the transitions within the cell cycle and ensuring the proper ordering of cell-cycle phases. Positive feedback mechanisms also contribute to the coordination of cytokinesis by ensuring the short and constant duration of mitosis and the proper Cdk1 activation thresholds.ref.7.2 ref.7.22 ref.7.2 Together, these regulatory mechanisms ensure the precise timing and coordination of mitotic events and cytokinesis, which are essential for the fidelity of cell division and cell survival.ref.7.2 ref.7.2 ref.7.3

What are the mechanisms that ensure equal division of cellular contents during cytokinesis?

Regulation of Cell Cycle Progression during Cytokinesis

During cytokinesis, the equal division of cellular contents is achieved through the coordination of various mechanisms that ensure the faithful transmission of genetic information and the proper distribution of cellular components. One of the key mechanisms involved in this process is the regulation of cell cycle progression. Transitions within the cell cycle are regulated by the activation and deactivation cycles of cyclin-dependent kinases (Cdks) and the synthesis and degradation of regulatory cyclins.ref.7.2 ref.7.2 ref.9.4

Cdks are a family of protein kinases that play a crucial role in controlling the cell cycle. They form complexes with cyclin subunits, and these complexes control the transitions between different phases of the cell cycle, such as the G1-to-S transition and the G2-to-M transition. In plants, two major classes of Cdks, CDKA and CDKB, directly drive cell cycle transitions.ref.2.4 ref.15.1 ref.2.4 CDKA is most closely related to mammalian CDK1 and CDK2 and regulates both the G1-to-S and G2-to-M transitions.ref.2.4 ref.2.5 ref.15.16

The regulation of Cdks occurs at multiple levels. First, they associate with cyclins, which are regulatory subunits that confer substrate specificity and regulate the activity of Cdks. Second, Cdks are regulated through phosphorylation and dephosphorylation events.ref.9.4 ref.2.4 ref.15.1 Phosphorylation of specific residues can either activate or inactivate Cdks, depending on the context. Third, Cdks interact with inhibitory proteins, such as CKIs (Cdk inhibitors), which bind to Cdks and prevent their activation. Lastly, Cdks are subject to targeted proteolysis, which is mediated by the ubiquitin-proteasome system and ensures the timely degradation of cyclins and Cdks at specific points in the cell cycle.ref.2.4 ref.4.2 ref.15.1

These regulatory mechanisms ensure the proper ordering of cell cycle phases and the completion of one phase before the onset of the next. Checkpoints also play a role in verifying the accurate completion of each phase before progression into the next phase. Together, these mechanisms ensure the fidelity of cell cycle progression and contribute to the equal division of cellular contents during cytokinesis.ref.7.2 ref.2.4 ref.7.2

Furrow Formation and Guidance in Animal Cells during Cytokinesis

In animal cells, cytokinesis involves the formation of a furrow that divides and separates the cytoskeletal microtubules and plasma membrane. The mechanisms responsible for furrow formation and guidance are not fully understood, but current research suggests that the furrow is formed through a search-and-capture mechanism that stabilizes microtubules attached to the cell cortex or organelles.ref.44.18 ref.49.15 ref.49.3

One of the key proteins involved in this process is EB1 (end-binding protein 1), which associates with the plus-ends of microtubules and regulates their dynamics. EB1 plays a role in stabilizing microtubules and promoting their attachment to the cell cortex or organelles. Another protein involved in furrow formation and guidance is dynein, which diffuses in the cytoplasm, attaches to microtubules, and undergoes 1-D diffusion along the microtubules to reach the plus end.ref.28.38 ref.28.43 ref.64.6 Dynein can then anchor at the cell cortex and generate pulling forces on the spindle poles, contributing to various aspects of mitosis, including anaphase B spindle elongation.ref.28.43 ref.64.6 ref.28.53

In addition to EB1 and dynein, other proteins and factors are likely involved in furrow formation and guidance. Katanins and spastin, for example, are microtubule-severing enzymes that play distinct roles at different stages of cytokinesis. These proteins may have different microtubule cleavage specificities, which could be influenced by post-translational modifications of microtubules and interactions with microtubule-binding proteins.ref.44.20 ref.44.18 ref.44.21

While the search-and-capture mechanism provides insights into the process of furrow formation and guidance, the exact details are still not fully understood. Ongoing research aims to uncover the precise molecular interactions and signaling pathways involved in these processes.

Cell Plate Formation and Guidance in Plant Cells during Cytokinesis

In plant cells, cytokinesis involves the formation of a cell plate that expands toward the cell periphery. The establishment of a cortical division zone, marked by a preprophase band (PPB) of microtubules and an actin-depleted zone, guides the cell plate. The cortical division zone is maintained throughout mitosis and is thought to be established by the presence of specific proteins and the rearrangement of the plasma membrane.ref.49.4 ref.49.3 ref.49.3

The preprophase band (PPB) of microtubules is thought to predict the future division plane and is removed before cytokinesis. It predefines the orientation and position of the division plane. The phragmoplast, which consists of microtubules, guides the expansion of the cell plate toward the cell periphery.ref.49.2 ref.49.1 ref.49.2 The communication between the cortical division zone and the leading edge of the expanding cell plate is achieved through short-range interactions mediated by microtubules and possibly actin. The contact between the growing cell plate and the cortical division zone is intense, as observed by the movement of microtubule plus-end marker EB1a-GFP between the two sites.ref.49.15 ref.49.15 ref.49.10

The establishment of the cortical division zone is regulated by various proteins. KCA1, for example, is excluded from the division zone and is responsible for establishing the division zone. TPLATE and AIR9 are targeted to the division zone and play a role in membrane trafficking.ref.49.12 ref.49.10 ref.49.18 The exact mechanisms and timeframes of division plane determination, phragmoplast guidance, and cell plate insertion site establishment are still areas of ongoing research.ref.49.18 ref.49.15 ref.49.2

The plasma membrane (PM) and its connection to the cell wall are vital for the maintenance of the cortical positional information during cytokinesis. The integrity of the PM and/or the PM-to-cell wall connection is important for the proper functioning of the division zone. Alterations in membrane composition or anchor proteins may play a role in the establishment and maintenance of the cortical division zone.ref.49.12 ref.49.7 ref.49.6 However, the exact nature of the molecular signals that regulate the expansion of the cell plate toward the cell periphery is still unknown.ref.49.6 ref.49.15 ref.49.3

Conclusion

In conclusion, the equal division of cellular contents during cytokinesis is achieved through the coordination of cell cycle progression, the formation of a furrow or cell plate, and the guidance of the phragmoplast or cell plate. The regulation of cell cycle progression ensures the proper ordering of cell cycle phases and the completion of one phase before the onset of the next. Furrow formation and guidance in animal cells involve a search-and-capture mechanism that stabilizes microtubules attached to the cell cortex or organelles.ref.49.3 ref.55.4 ref.49.1 Cell plate formation and guidance in plant cells involve the establishment of a cortical division zone marked by a preprophase band of microtubules and an actin-depleted zone.ref.49.3 ref.49.1 ref.49.15

While significant progress has been made in understanding these mechanisms, there are still many unanswered questions. The exact details of furrow formation and guidance in animal cells, as well as cell plate formation and guidance in plant cells, remain areas of ongoing research. Further investigation into the molecular interactions and signaling pathways involved in these processes will provide valuable insights into the fundamental principles of cytokinesis and the regulation of cell division.ref.49.15 ref.49.15 ref.49.15

Mitotic Checkpoint and Error Correction

How does the mitotic checkpoint system ensure accurate chromosome segregation?

Introduction

The mitotic checkpoint system, also known as the Spindle Assembly Checkpoint (SAC), plays a critical role in ensuring accurate chromosome segregation during the transition from metaphase to anaphase in the two meiotic cell divisions. This checkpoint monitors the attachment of kinetochores to microtubules and ensures correct kinetochore-microtubule attachments and faithful chromosome segregation. It functions through unattached kinetochores, generating a soluble mitotic checkpoint complex (MCC) consisting of Mad2, Cdc20, BubR1, and Bub3.ref.63.2 ref.59.2 ref.22.0 The MCC directly inhibits the anaphase-promoting complex/cyclosome (APC/C), which is an E3 ubiquitin ligase complex responsible for regulating kinetochore function and sister chromatid cohesion. In this essay, we will explore the intricate mechanisms and key components of the mitotic checkpoint system, as well as its relationship with the APC/C, cyclin B, securin, and the activation of separase.ref.58.1 ref.59.2 ref.30.2

The Mitotic Checkpoint System

The mitotic checkpoint system acts through unattached kinetochores to ensure accurate chromosome segregation. When microtubules are not properly attached to kinetochores, the checkpoint is engaged, leading to the production of the MCC. The MCC is a soluble complex that inhibits the APC/C, preventing the degradation of cyclin B and securin, and hence delaying anaphase onset.ref.59.0 ref.58.1 ref.59.2

The formation of the MCC is a crucial step in the activation of the mitotic checkpoint. The core components of the MCC are Mad2 and Cdc20. Mad2 is an essential checkpoint protein that can adopt two different conformations: "open" (O-Mad2) or "closed" (C-Mad2).ref.59.2 ref.59.3 ref.59.0 In O-Mad2, the C-terminal seatbelt occupies the MIM binding site, preventing complex formation. At unattached kinetochores, O-Mad2 is recruited to Mad1:C-Mad2 complexes and converted to C-Mad2. C-Mad2 then assembles with Cdc20 to form the core of the MCC.ref.59.3 ref.59.2 ref.25.3 This assembly of the Cdc20:C-Mad2 complex is crucial for the inhibition of the APC/C and the delay of mitotic exit.ref.25.3 ref.59.2 ref.59.3

In addition to Mad2 and Cdc20, the MCC also consists of BubR1 and Bub3. BubR1 plays a role in MCC assembly and is recruited along with Bub3 to complete the assembly of the MCC. The fully assembled MCC, comprising Mad2, Cdc20, BubR1, and Bub3, then binds to the APC/CCdc20 complex as a substrate analog and directly inhibits its activity.ref.25.3 ref.25.3 ref.66.3 This inhibition of the APC/C by the MCC delays mitotic exit, allowing for accurate chromosome segregation.ref.59.3 ref.30.1 ref.30.21

Relationship with the APC/C, Cyclin B, Securin, and Separase

The APC/C, together with its co-activator Cdc20, is responsible for the degradation of cyclin B and securin, which allows for sister chromatid separation and mitotic exit. The APC/C remains inhibited until all chromosomes are properly attached to microtubules via their kinetochores. Unattached kinetochores catalyze the production of the MCC, which directly binds to the APC/C and inhibits its activity.ref.57.1 ref.58.2 ref.66.3

The degradation of cyclin B and securin by the APC/C triggers the activation of separase and the inactivation of cyclin-dependent kinase 1 (Cdk1), leading to sister chromatid disjunction and mitotic exit. The simultaneous destruction of securin and cyclin B by the APC/C results in the dissolution of attachment monitoring mechanisms during mitotic exit. If Cdk1 is not inactivated concomitantly with separase-induced sister chromatid separation, the mitotic checkpoint can be engaged, leading to the inhibition of the APC/C and the reaccumulation of securin and cyclin B.ref.58.2 ref.58.24 ref.58.23 This reaccumulation could interfere with chromatid separation and mitotic exit. However, the reaccumulation of securin and cyclin B after sister chromatid separation is slow and not significant within a short timeframe due to the requirement for resynthesis and partial inhibition of the APC/C. The degradation of cyclin B and the reversal of Cdk1-dependent phosphorylation are crucial for the generation of stable kinetochore-microtubule attachments that underlie the correct partitioning of sister chromatids in anaphase.ref.58.24 ref.58.2 ref.58.23

Regulation and Coordination of the Mitotic Checkpoint System

The delicate balance between MCC assembly and disassembly/degradation pathways, as well as their coordination through space and time, is crucial to consider in the study of cancer. Two separate catalytic pathways, TRIP13/p31comet-mediated MCC disassembly through C-Mad2 to O-Mad2 conformational conversion and APC15-dependent ubiquitination of Cdc20 in MCC, are essential for inactivating MCC made either in interphase or in mitosis. MCC complexes are extraordinarily stable once assembled, with a negligible rate of spontaneous inactivation.ref.59.31 ref.59.4 ref.59.32 Chronic mitotic arrest occurs in cells lacking both MCC disassembly and degradation pathways, even in the absence of actively signaling kinetochores. Low levels of TRIP13 are unable to quickly reactivate APC/CCdc20 that has been pre-incubated with MCC components but are sufficient to disassemble free MCC and prevent APC/CCdc20 inhibition. This indicates that free MCC is the preferred substrate of TRIP13, but low-level activity on APC/C-MCC complexes is not eliminated.ref.59.31 ref.59.32 ref.59.26

Detection of Unattached Kinetochores and Dependence on Tension

The mitotic checkpoint system detects unattached kinetochores through the recruitment of checkpoint proteins such as Mad1 and Mad2 to the kinetochores. This signal is amplified when microtubules become detached from kinetochores, leading to the engagement of the mitotic checkpoint and the inhibition of anaphase onset. The engagement of the mitotic checkpoint is dependent on the presence of tension across sister kinetochores and the correct attachment of chromosomes.ref.65.41 ref.58.8 ref.58.7 If tension is lost or kinetochores become detached, the mitotic checkpoint can be activated. The inactivation of Cdk1 at anaphase onset disables the mitotic checkpoint surveillance in human cells. Preventing cyclin B1 proteolysis at the time of sister chromatid disjunction destabilizes kinetochore-microtubule attachments and triggers the engagement of the mitotic checkpoint.ref.58.1 ref.58.23 ref.58.1 Conversely, acute pharmacological inhibition of Cdk1 abolishes the engagement and maintenance of the mitotic checkpoint upon microtubule depolymerization. Therefore, the mitotic checkpoint system detects unattached kinetochores through the recruitment of checkpoint proteins and is dependent on tension and correct attachment of chromosomes.ref.58.1 ref.58.23 ref.58.2

Involvement in Response to DNA Damage

The mitotic checkpoint system also plays a role in responding to chromosomal breakage during M-phase. In the presence of DNA double-strand breaks (DSBs), the checkpoint prevents the onset of mitosis and delays spindle assembly. This checkpoint affects centrosome-driven spindle assembly and involves the phosphorylation of XCEP63 by ATM and ATR, leading to its delocalization from the centrosome and a delay in spindle assembly.ref.24.2 ref.24.0 ref.24.0

Conclusion

The mitotic checkpoint system is a complex and highly regulated mechanism that ensures accurate chromosome segregation during cell division. Through the generation of the MCC, the system inhibits the APC/C and delays mitotic exit until all chromosomes have successfully attached to spindle microtubules. Key components such as Mad2, Cdc20, BubR1, and Bub3 play essential roles in the formation of the MCC and its interaction with the APC/C.ref.59.0 ref.59.2 ref.59.2 The delicate balance between MCC assembly and disassembly/degradation pathways, as well as the detection of unattached kinetochores and the dependence on tension, further highlight the complexity and precision of the mitotic checkpoint system. Understanding the mechanisms and regulation of this system is crucial for unraveling the underlying causes of chromosomal instability and the development of targeted cancer therapies.ref.59.31 ref.59.0 ref.58.1

What are the molecular mechanisms that detect errors and delay cell cycle progression until the errors are corrected?

Introduction

The cell cycle is a highly regulated process that ensures the accurate replication and segregation of genetic material during cell division. To maintain genomic stability, cells have evolved mechanisms to detect errors and delay cell cycle progression until these errors are corrected. One of the key mechanisms involved in error detection is the mitotic checkpoint, also known as the spindle-assembly checkpoint.ref.30.2 ref.59.2 ref.68.3 This checkpoint monitors the attachment of chromosomes to spindle microtubules and delays the onset of anaphase until all chromosomes have successfully attached. The mitotic checkpoint functions through the assembly of the mitotic checkpoint complex (MCC), which inhibits the activity of the anaphase-promoting complex/cyclosome (APC/C), a key regulator of cell cycle progression. In addition to the mitotic checkpoint, DNA damage response proteins, such as ATM and ATR, also contribute to cell cycle delay during mitosis.ref.59.2 ref.63.2 ref.58.1 In this essay, we will discuss the molecular mechanisms involved in error detection and cell cycle delay during mitosis.ref.72.2 ref.24.2 ref.68.3

The Mitotic Checkpoint

A. Spindle Checkpoint and Chromosome Attachment The mitotic checkpoint recognizes and detects errors in chromosome attachment to spindle microtubules through a molecular safety mechanism called the spindle checkpoint. This checkpoint monitors the bi-orientation of chromosomes, ensuring that sister chromatids are attached to microtubules from opposite spindle poles.ref.62.0 ref.63.2 ref.59.2 When sister chromatids fail to bi-orient, spindle checkpoint components, including Bub1, Bub3, Mad1, and Mad2, are hierarchically recruited to kinetochores. The key component controlling mitotic checkpoint assembly is Mad2, which can adopt two different conformations: "open" (O-Mad2) or "closed" (C-Mad2). C-Mad2 binds partner proteins, including Cdc20 and Mad1, through a distinctive "seat belt" interaction.ref.62.0 ref.65.40 ref.58.7 In O-Mad2, the C-terminal seat belt occupies the MIM binding site to prevent complex formation. The engagement of the mitotic checkpoint leads to the accumulation of mitotic checkpoint proteins at anaphase kinetochores, inhibition of the APC/C, and reaccumulation of securin. The inhibition of Cdk1 at the time of sister chromatid disjunction destabilizes kinetochore-microtubule attachments and triggers the engagement of the mitotic checkpoint.ref.58.1 ref.58.7 ref.58.1

The mitotic checkpoint complex (MCC) consists of several components, including Mad2, Cdc20, BubR1, and Bub3. The key component controlling MCC assembly is Mad2, which can adopt two different conformations: "open" (O-Mad2) or "closed" (C-Mad2). C-Mad2 binds partner proteins, including Cdc20 and Mad1, through a distinctive "seat belt" interaction.ref.66.3 ref.25.3 ref.59.2 In O-Mad2, the C-terminal seat belt occupies the binding site to prevent complex formation. O-Mad2 is recruited to Mad1:C-Mad2 complexes at unattached kinetochores, where it is converted to C-Mad2 and assembled with Cdc20 to form the core of the MCC. BubR1 and Bub3 are then recruited to complete the assembly of the MCC.ref.59.3 ref.59.2 ref.25.3 The MCC binds to the APC/C and inhibits its recognition of cyclin B and securin, thereby delaying mitotic exit. Once all kinetochores have become attached to microtubules, the mitotic checkpoint is silenced, and APC/C is reactivated for cyclin B and securin ubiquitination, initiating anaphase onset.ref.59.3 ref.87.42 ref.30.21

The mitotic checkpoint is tightly regulated to ensure its proper function. One of the key regulators of the mitotic checkpoint is Cdk1, a key cell cycle regulator. Inactivation of Cdk1 disables the mitotic checkpoint surveillance at anaphase onset.ref.58.1 ref.59.2 ref.58.1 Acute pharmacological inhibition of Cdk1 abolishes the engagement and maintenance of the mitotic checkpoint upon microtubule depolymerization. Additionally, the assembly and disassembly of the mitotic checkpoint components are regulated by various post-translational modifications, including phosphorylation, ubiquitination, and sumoylation. These modifications regulate the stability, localization, and activity of the mitotic checkpoint proteins, ensuring the precise control of cell cycle progression.ref.58.12 ref.58.1 ref.58.2

DNA Damage Response and Mitotic Arrest

In addition to the mitotic checkpoint, DNA damage response proteins, such as ATM and ATR, contribute to mitotic arrest following DNA damage. ATM and ATR are activated in response to DNA damage and phosphorylate various proteins involved in cell cycle regulation, such as cyclin B1 and cdc25C. This leads to the inhibition of the mitotic kinase cdc2 and prevents cells from progressing into mitosis.ref.67.4 ref.73.9 ref.14.4 ATM and ATR also play a role in the repair of DNA damage during mitosis. They are involved in the recognition and processing of DNA double-strand breaks (DSBs) and the recruitment of repair factors to the damaged sites. The exact mechanisms by which ATM and ATR impact mitosis progression in the presence of DNA damage are still not fully understood.ref.24.3 ref.73.10 ref.71.26 Some studies suggest that ATM and ATR-dependent pathways may affect spindle assembly and mitotic progression, while others indicate that their activation is primarily linked to kinetochore damage and the spindle assembly checkpoint (SAC). Further research is needed to elucidate the precise roles of ATM and ATR in mitotic arrest following DNA damage.ref.24.3 ref.24.5 ref.24.2

Other Mechanisms of Error Detection and Cell Cycle Delay

Besides the mitotic checkpoint and the DNA damage response, there are other mechanisms that play a role in error detection and cell cycle delay during mitosis. One such mechanism is the DNA damage response (DDR) pathway. DNA double-strand breaks (DSBs) are one of the most deleterious types of DNA damage, and during mitosis, the repair of DSBs primarily occurs through error-prone nonhomologous end joining (NHEJ), alternative end joining (Alt-EJ), and single-strand annealing (SSA) pathways.ref.72.2 ref.69.1 ref.69.1 Additionally, there is evidence for the existence of a mid-anaphase DNA damage checkpoint that is dependent on the yeast homologs of securin and separase. This checkpoint can delay mitotic progression in response to DNA damage. Furthermore, studies have shown that cells can adapt to the presence of unrepaired chromosomal breakages and enter mitosis, overcoming the G2/M checkpoint.ref.72.2 ref.72.1 ref.72.17 It has also been demonstrated that DNA repair can occur in mitosis, as evidenced by the repair of DNA double-strand breaks in mitotic cells. However, the exact mechanisms by which cells react to DNA damage in mitosis and the extent of DNA repair in mitosis are still not fully understood.ref.72.6 ref.69.34 ref.72.1

Chromosomal Instability and Therapeutic Potential

Bypassing or disabling the mitotic checkpoint surveillance at anaphase onset can lead to increased chromosome missegregation during mitosis, resulting in aneuploidy or an abnormal number of chromosomes. This defect is seen more frequently in tumor cell divisions compared to normal cell divisions, and a large fraction of human tumors display a chromosome instable phenotype. Inhibition of the mitotic checkpoint has been shown to increase cell death in tumor cells, suggesting that it could be a useful anti-cancer strategy.ref.79.0 ref.79.0 ref.79.1 However, complete inactivation of the mitotic checkpoint is not compatible with cell viability. The exact cause of chromosomal instability (CIN) in tumors is still largely unknown, but defects in mitotic checkpoint function have been implicated as a causal event for CIN. Mutations or altered expression of mitotic checkpoint genes have been associated with CIN and aneuploidy in various human cancers.ref.79.1 ref.79.1 ref.79.0 However, it is important to note that checkpoint malfunctions as a direct cause of CIN in tumor cells have not been found, and partial checkpoint defects may be well tolerated by the tumor cell population. Therefore, the therapeutic potential of inhibiting the mitotic checkpoint in cancer treatment is still being explored.ref.79.1 ref.79.1 ref.79.1

Conclusion

The molecular mechanisms involved in error detection and cell cycle delay during mitosis are complex and involve the mitotic checkpoint, the assembly of the mitotic checkpoint complex, the regulation of Cdk1 activity, and the activation of DNA damage response proteins. These mechanisms ensure the proper attachment of chromosomes to the spindle and prevent the propagation of DNA damage during mitosis. Further research is needed to fully understand these mechanisms and their implications for human health and disease, including cancer.ref.68.3 ref.59.2 ref.72.1 By unraveling the intricacies of these processes, we can potentially develop new strategies for cancer therapy and improve our understanding of the fundamental processes that govern cell division.ref.72.1 ref.72.1 ref.68.3

How do the spindle assembly checkpoint and the DNA damage checkpoint function to maintain genomic stability during mitosis?

Introduction

The spindle assembly checkpoint (SAC) and the DNA damage checkpoint are two critical mechanisms that function to maintain genomic stability during mitosis. The SAC ensures accurate chromosome segregation by monitoring the attachment of chromosomes to the mitotic spindle, while the DNA damage checkpoint responds to DNA double-strand breaks (DSBs) and delays mitotic progression to allow for repair. Both checkpoints involve the activation of specific proteins and the inhibition of the anaphase-promoting complex/cyclosome (APC/C).ref.72.9 ref.63.2 ref.59.2 However, there are also differences between the two checkpoints, such as the requirement of a functional kinetochore for the standard SAC but not for the DNA-damage-induced mitotic arrest. In this essay, we will explore the regulatory mechanisms and protein components of these checkpoints, their similarities and differences, and the potential consequences of their failure.ref.72.3 ref.72.9 ref.63.2

Mechanisms of the Spindle Assembly Checkpoint

The spindle assembly checkpoint ensures proper chromosome alignment and attachment to the mitotic spindle during mitosis. Several key mechanisms contribute to its ability to monitor chromosome attachment:ref.62.0 ref.87.3 ref.63.2

1. Continuous transcription of mitotic regulators: Sustained activation of the spindle assembly checkpoint requires de novo transcription and translation of cyclin B1, as shown in a study by Mena et al. (2010). This suggests that continuous transcription of mitotic regulators is necessary to sustain the activation of the checkpoint.ref.63.1 ref.63.0 ref.63.0

2. Formation of the mitotic checkpoint complex (MCC): The MCC is a diffusible signal that inhibits the APC/C, which promotes the degradation of cyclin B1 and securin, leading to anaphase onset. The MCC consists of Mad2, Cdc20, BubR1, and Bub3.ref.58.9 ref.66.3 ref.59.2 Unattached kinetochores generate the MCC, which directly inhibits the APC/C activity.ref.59.2 ref.66.3 ref.59.0

3. Role of Mad2: Mad2 is an essential checkpoint protein that plays a key role in MCC assembly. It can adopt two different conformations: "open" (O-Mad2) and "closed" (C-Mad2).ref.59.3 ref.59.1 ref.59.3 C-Mad2 binds partner proteins, including Cdc20 and Mad1, through a distinctive "seat belt" interaction. In O-Mad2, the C-terminal seatbelt occupies the MIM binding site to prevent complex formation. The conformational switch of Mad2 is crucial for the assembly of the MCC.ref.59.3 ref.25.3 ref.59.4

4. Kinetochore-microtubule attachments: The spindle assembly checkpoint monitors the microtubule attachment to kinetochores. When sister chromatids fail to bi-orient, spindle checkpoint components, including Bub1, Bub3, Mad1, and Mad2, are hierarchically recruited to kinetochores.ref.62.0 ref.65.41 ref.63.2 The correct attachment of all kinetochores to the mitotic spindle satisfies the checkpoint and allows anaphase onset.ref.86.7 ref.65.5 ref.65.41

5. Nuclear envelope breakdown (NEBD): NEBD is the process of disassembling the nuclear envelope at the onset of mitosis. This allows the chromosomes to be accessible to the mitotic spindle.ref.64.2 ref.86.8 ref.86.9 NEBD involves the dispersal of nucleoporins, physical deformation and tearing of the nuclear envelope, and disassembly of the nuclear lamina. NEBD is coordinated with the disassembly of the interphase microtubule cytoskeleton and the formation of asters around the centrosomes.ref.64.2 ref.86.8 ref.86.9

6. Checkpoint adaptation: Cells can adapt to the presence of unrepaired chromosomal breakages and overcome the G2/M checkpoint, entering mitosis. The sensitivity of the G2/M checkpoint to DNA damage varies, and cells with defects in DNA repair can enter mitosis with a high number of unrepaired DNA double-strand breaks.ref.24.2 ref.24.2 ref.14.13 The engagement of the mitotic checkpoint depends on the kinetochore status and the activation of ATM and ATR pathways.ref.24.2 ref.24.2 ref.73.9

Mechanisms of the DNA Damage Checkpoint

The DNA damage checkpoint is activated in response to DNA damage, including double-strand breaks (DSBs), during mitosis. The activation of this checkpoint involves a complex series of signaling pathways, although the exact mechanisms are still being studied. Some key findings include:ref.72.4 ref.72.1 ref.72.2

1. Activation of ATM and ATR: The presence of DSBs activates the ATM and ATR pathways, which are key regulators of the DNA damage response (DDR). These pathways phosphorylate downstream checkpoint proteins and initiate the DNA damage checkpoint signaling cascade.ref.71.26 ref.67.4 ref.14.4

2. Role of yeast homologs of securin and separase: There is evidence of a mid-anaphase DNA damage checkpoint in yeast, which is dependent on the yeast homologs of securin and separase. This suggests that the DNA damage checkpoint can be activated during mitosis and not just during interphase.ref.72.2 ref.72.3 ref.72.17

3. Crosstalk between the DNA damage response (DDR) and the spindle assembly checkpoint (SAC): The DDR senses mitotic DNA damage and activates the SAC, potentially through specific damage to the kinetochores. This crosstalk ensures that mitotic progression is delayed until DNA repair is completed.ref.72.2 ref.72.17 ref.72.4

4. Minimal DNA repair in mitosis: Studies have shown that mitotic cells are capable of repairing DNA double-strand breaks, although the efficiency of repair is reduced compared to interphase cells. Mitotic repair is mediated by DNA-PKcs and ATM, and the repair process is attenuated when both proteins are compromised.ref.72.6 ref.69.34 ref.72.1

Similarities and Differences between the Spindle Assembly Checkpoint and the DNA Damage Checkpoint

The spindle assembly checkpoint and the DNA damage checkpoint share some similarities in their regulatory mechanisms and protein components. Both checkpoints involve the activation of specific proteins, such as Mad2, and the inhibition of the APC/C. However, there are also differences between the two checkpoints.ref.59.2 ref.65.38 ref.63.2 For example, a functional kinetochore is not required for DNA-damage-induced mitotic arrest, unlike the standard SAC. Further research is needed to fully understand the similarities and differences between these two checkpoints.ref.72.3 ref.72.2 ref.87.42

Consequences of Checkpoint Failure

A failure in either the spindle assembly checkpoint or the DNA damage checkpoint during mitosis can have potential consequences and contribute to genomic instability. Failure in the spindle assembly checkpoint can lead to chromosome missegregation and aneuploidy, which is an abnormal number of chromosomes. This can also result in chromosomal rearrangements, contributing to genomic instability and the development of cancer.ref.79.1 ref.79.0 ref.79.0

Similarly, failure in the DNA damage checkpoint can lead to the generation of chromosomal rearrangements and aneuploidy. Cells may progress into mitosis with unrepaired DNA damage, resulting in the accumulation of DNA double-strand breaks. This can lead to chromosomal abnormalities, such as translocations and telomere fusions, which are associated with genomic instability and the development of cancer.ref.72.2 ref.79.0 ref.24.2

In summary, the spindle assembly checkpoint and the DNA damage checkpoint are crucial for maintaining genomic stability during mitosis. They ensure accurate chromosome segregation and prevent the progression of cells with damaged DNA. While there are similarities in their regulatory mechanisms and protein components, there are also differences between the two checkpoints.ref.87.3 ref.63.1 ref.72.2 Failure in either checkpoint can have significant consequences, contributing to genomic instability and the development of cancer. Further research is needed to fully understand the intricate mechanisms and regulation of these checkpoints.ref.72.2 ref.63.1 ref.72.2

What are the consequences of mitotic errors and how are they prevented or repaired?

Introduction

Mitosis is a fundamental process in cell division that ensures the proper segregation of chromosomes into daughter cells. However, errors can occur during mitosis, leading to chromosomal rearrangements, aneuploidy, and genomic instability. These errors can have severe consequences, including developmental disorders, cell death, and cancer.ref.79.0 ref.79.0 ref.79.0 To prevent and repair mitotic errors, cells have evolved various mechanisms, including the mitotic checkpoint and the DNA damage response. In recent years, there has been significant research focused on understanding the molecular mechanisms underlying mitotic error prevention and repair. This essay will explore the current areas of research in this field and discuss the potential implications of these findings for cancer therapy and our understanding of genomic instability-related diseases.ref.72.1 ref.72.6 ref.72.2

Mitotic Error Prevention

A. The Mitotic Checkpoint The mitotic checkpoint, also known as the spindle assembly checkpoint (SAC), is a crucial mechanism that ensures the fidelity of sister chromatid segregation during mitosis. It functions by delaying the onset of anaphase until all chromosomes have correctly attached to the spindle microtubules.ref.63.2 ref.63.1 ref.59.2 The SAC is activated by unattached kinetochores, which generate a soluble mitotic checkpoint complex (MCC) consisting of Mad2, Cdc20, BubR1, and Bub3. The MCC binds to the anaphase-promoting complex/cyclosome (APC/C) and inhibits its activity, thereby preventing the degradation of cyclin B and securin, which are essential for mitotic progression.ref.63.2 ref.22.0 ref.87.42

In addition to the mitotic checkpoint, DNA damage checkpoints can also be activated in mitosis in response to double-strand breaks (DSBs). The DNA damage response (DDR) in mitosis involves the activation of proteins such as ATM, ATR, H2AX, and MDC1. These proteins play a role in repairing DSBs through mechanisms such as homologous recombination (HR) and non-homologous end joining (NHEJ).ref.72.4 ref.69.1 ref.72.2 However, the repair of DSBs in mitosis is less efficient compared to interphase, and inaccurate repair can lead to chromatin bridges and DNA rearrangements.ref.24.2 ref.72.6 ref.69.1

Current Areas of Research

A. DNA Damage Checkpoint and Repair Proteins One area of ongoing research focuses on studying the proteins involved in the DNA damage checkpoint and repair processes during mitosis. Proteins such as ATM, ATR, XRCC4, and BRCA1 have been identified as key players in these pathways.ref.72.6 ref.73.9 ref.72.2 Understanding the precise roles of these proteins and their interactions will provide insights into the mechanisms of mitotic error prevention and repair.ref.72.6 ref.69.1 ref.73.9

Post-translational modifications of proteins are crucial for regulating their functions. Research is being conducted to investigate the role of post-translational modifications in the activation of the mitotic checkpoint and the repair of mitotic errors. For example, the phosphorylation of XRCC4 in mitosis has been shown to impact the efficiency of NHEJ and the formation of chromatin bridges.ref.72.6 ref.73.15 ref.72.5

Another area of research focuses on the mitotic DNA damage response (DDR), which involves the activation of various repair pathways in response to DNA damage during mitosis. The exact mechanisms and factors involved in this process are still being studied. It is becoming increasingly clear that the mitotic DDR is more complex than previously thought and may involve factors from multiple repair pathways.ref.69.1 ref.69.1 ref.69.1

Implications and Potential Applications

A. Targeted Therapies for Cancer The knowledge gained from studying mitotic error prevention and repair mechanisms has the potential to inform the development of targeted therapies for cancer. Inhibiting the mitotic checkpoint through the depletion of essential checkpoint components, such as Mps1, has been shown to increase cell death in tumor cells.ref.79.0 ref.88.2 ref.79.0 This approach could be explored further to selectively kill tumor cells while sparing normal cells.ref.88.2 ref.79.0 ref.79.0

A better understanding of the mechanisms underlying mitotic error prevention and repair can provide insights into the development of genomic instability-related diseases, such as cancer, premature aging, and developmental disorders. Chromosomal rearrangements and aneuploidy are common features of these diseases. By elucidating the causes and consequences of chromosomal instability, researchers can potentially develop strategies to prevent or mitigate these conditions.ref.79.0 ref.79.0 ref.79.1

Conclusion

Mitotic error prevention and repair mechanisms are crucial for maintaining genomic stability during cell division. The mitotic checkpoint and the DNA damage response play key roles in ensuring the fidelity of chromosome segregation and repairing DNA damage. Ongoing research is focused on understanding the proteins and molecules involved in these processes, as well as the mechanisms by which mitotic errors are prevented and repaired.ref.72.1 ref.72.6 ref.72.2 The findings from these studies have the potential to have a significant impact on cancer therapy and our understanding of genomic instability-related diseases. Further research is needed to fully unravel the intricacies of mitotic error prevention and repair and translate this knowledge into clinical applications.ref.69.1 ref.72.1 ref.72.6

How do mitotic errors contribute to genetic instability and disease?

Introduction

Mitotic errors and DNA damage can have significant consequences on genetic stability and contribute to the development of diseases, including cancer. In particular, chromosome missegregation during mitosis can lead to aneuploidy, which is an abnormal number of chromosomes, and genetic instability in tumor tissue. Aneuploidy is a common characteristic of cancer cells, and it is often caused by chromosome missegregation during mitosis.ref.79.0 ref.79.0 ref.79.0 Mitotic checkpoint function, which ensures the fidelity of sister chromatid segregation, has been implicated in chromosome instability (CIN) and aneuploidy in various human cancers. However, the exact cause of CIN in tumors is still not fully understood. It has been suggested that defects in the mitotic checkpoint could be a causal event for CIN in tumors.ref.79.1 ref.79.0 ref.79.1 Furthermore, mitotic errors can also lead to chromosomal rearrangements, telomere fusions, and altered cell fate, all of which can contribute to genetic instability and disease.ref.79.0 ref.79.0 ref.92.2

Chromosome Missegregation and Aneuploidy in Tumors

Chromosome Missegregation and Aneuploidy in Tumor Tissue

Chromosome missegregation during mitosis can lead to aneuploidy and genetic instability in tumor tissue. Aneuploidy, characterized by an abnormal number of chromosomes, is a common feature of cancer cells. This abnormal chromosome content is a result of chromosome missegregation during mitosis, which occurs more frequently in tumor cell divisions compared to normal cell divisions.ref.79.0 ref.79.0 ref.92.2 Tumor cells often display a chromosome instable phenotype, meaning they frequently missegregate chromosomes, leading to variegated aneuploidy within the tumor tissue. The exact cause of chromosome missegregation in tumors is not fully understood, but defects in mitotic checkpoint function have been implicated as a contributing factor. The mitotic checkpoint ensures the fidelity of sister chromatid segregation by delaying the segregation of chromosomes until all chromosome pairs are properly attached to the mitotic spindle.ref.79.0 ref.79.0 ref.79.1 Inhibition or partial inactivation of the mitotic checkpoint can lead to increased segregation errors and chromosome missegregation, which has been shown to result in increased cell death in tumor cells. It has been suggested that targeting the mitotic checkpoint could be a useful anti-cancer strategy. Additionally, chromosomes with larger kinetochores have been found to have a higher tendency to establish erroneous merotelic attachments and missegregate during anaphase, potentially leading to aneuploidy.ref.79.0 ref.79.0 ref.79.1 The size of the kinetochore, which is the critical chromosomal interface with spindle microtubules, is an important determinant of chromosome segregation fidelity. However, further research is needed to fully understand the mechanisms and potential therapeutic implications of chromosome missegregation in tumor tissue.ref.92.4 ref.92.3 ref.92.4

Mitotic Checkpoint Function and Chromosome Segregation

Role of the Mitotic Checkpoint in Sister Chromatid Segregation

The mitotic checkpoint plays a crucial role in ensuring the fidelity of sister chromatid segregation by monitoring the attachment of microtubules to kinetochores and delaying anaphase onset until all chromosomes are properly attached. This checkpoint acts during the transition between metaphase and anaphase of the two meiotic cell divisions to ensure correct kinetochore-microtubule attachments and faithful chromosome segregation. It is activated by the presence of unattached kinetochores and inhibits the anaphase-promoting complex/cyclosome (APC/C).ref.59.2 ref.63.2 ref.87.3 The activation of the mitotic checkpoint leads to the accumulation of mitotic checkpoint proteins at anaphase kinetochores, inhibition of the APC/C, and reaccumulation of securin. The mitotic checkpoint is blind to replication and recombination intermediates as well as rearranged chromosomes. The spindle assembly checkpoint (SAC) is also dependent on the yeast homologs of securin and separase (Pds1 and Esp1, respectively).ref.58.2 ref.63.2 ref.59.2 In plants, the checkpoints appear to be less stringent compared to yeast and animals, as completion of meiosis is achieved in several meiotic mutants creating imbalanced gametes. The mitotic checkpoint is regulated by the activity of CDK-cyclin complexes and the APC/C. Continuous transcription of mitotic regulators is required to sustain the activation of the spindle assembly checkpoint.ref.87.3 ref.59.2 ref.58.23

Mitotic Errors, DNA Damage, and Altered Cell Fate

Mechanisms of Altered Cell Fate

Mitotic errors and DNA damage can lead to altered cell fate, such as cell senescence, apoptosis, and necrosis. These outcomes are typically mediated by various signaling pathways and checkpoints.ref.72.1 ref.72.17 ref.72.17

cGAS-STING Axis and Cellular Responses

One mechanism of altered cell fate is the activation of the cGAS-STING axis, which is essential for both cell senescence and cell death via apoptosis and necrosis. This pathway is involved in the recognition of DNA damage and the initiation of cellular responses.ref.72.16 ref.72.17 ref.101.10

DNA Damage Response (DDR) and Spindle Assembly Checkpoint (SAC)

Another mechanism involves the activation of the DNA damage response (DDR) and the spindle assembly checkpoint (SAC). The DDR and SAC play a role in controlling mitotic progression and cell fate in the presence of DNA damage. The DDR proteins ATM and ATR have been shown to be required for mitotic arrest following DNA damage.ref.72.2 ref.24.3 ref.72.2 The SAC delays the segregation of chromosomes during mitosis until all chromosomes are properly attached to the mitotic spindle. Inhibition of the SAC can lead to increased segregation errors and cell death in tumor cells.ref.72.3 ref.72.9 ref.72.2

Minimal DNA Repair in Mitosis

Despite the inhibitions on DNA repair pathways, studies have shown that DNA double-strand breaks (DSBs) can be repaired in mitosis, and the NHEJ protein XRCC4 is phosphorylated specifically in mitosis, indicating its involvement in DNA repair. However, inaccurate repair in mitosis can lead to chromatin bridges, which can result in altered cell fate.ref.72.6 ref.69.3 ref.69.29

Mitotic DNA Damage and Repair Mechanisms

Repair Mechanisms for Mitotic DNA Damage

Mitotic DNA damage can result in chromosomal breaks and other types of DNA damage. When DNA double-strand breaks (DSBs) occur during mitosis, the most likely repair mechanisms are the error-prone nonhomologous end joining (NHEJ), alternative end joining (Alt-EJ), and single-strand annealing (SSA) pathways. However, repair in mitosis is limited and can lead to chromosomal translocations, telomere fusions, and aneuploidy.ref.72.2 ref.69.1 ref.69.3

Existence of a Mitotic DNA Damage Checkpoint

It has been proposed that cells with damaged DNA progress through mitosis with similar kinetics to unperturbed cells and then initiate the DNA damage response (DDR) and begin repair in G1 phase, rather than facilitating a response in mitosis. However, there is increasing evidence for the existence of a mitotic DNA damage checkpoint. Experiments in yeast have shown the existence of a mid-anaphase DNA damage checkpoint, which is dependent on the yeast homologs of securin and separase.ref.69.1 ref.72.2 ref.72.2 Additionally, recent studies have demonstrated that the DDR proteins ATM and ATR are required for mitotic arrest following DNA damage. The spindle assembly checkpoint (SAC) is also involved in the response to DNA damage in mitosis, and a functional kinetochore is not required for DNA-damage-induced mitotic arrest.ref.72.17 ref.72.3 ref.24.2

Chromatin Bridges and Repair in Mitosis

Mitotic DNA damage can lead to chromatin bridges, which occur when DNA from two chromosomes or chromatids is fused together. These chromatin bridges can be formed by incorrect DNA repair or telomere fusion. There is evidence that inaccurate repair in mitosis can lead to DNA bridges.ref.72.6 ref.72.7 ref.72.2 Furthermore, recent studies have shown that MDC1 foci, which form at DNA breaks in mitosis, recruit TopBP1, which then forms filamentous structures that tether DSBs and allow proper segregation of broken chromosomes until repair can be carried out in G1.ref.72.7 ref.72.8 ref.72.9

Conclusion

In summary, mitotic errors and DNA damage can have profound effects on genetic stability and disease development. Chromosome missegregation during mitosis can lead to aneuploidy and genetic instability in tumor tissue. The mitotic checkpoint plays a crucial role in ensuring the fidelity of sister chromatid segregation and defects in its function have been implicated in chromosome instability and aneuploidy in tumors.ref.79.1 ref.79.0 ref.79.0 Mitotic errors and DNA damage can also lead to altered cell fate, such as cell senescence, apoptosis, and necrosis. Additionally, mitotic DNA damage can result in chromosomal breaks and other types of DNA damage. While repair in mitosis is limited, there is evidence for the existence of a mitotic DNA damage checkpoint.ref.72.1 ref.72.17 ref.72.2 Further research is needed to fully understand the molecular mechanisms involved in these processes and their potential therapeutic implications.ref.79.1 ref.72.1 ref.79.0

Mitotic Signaling Pathways

What are the key signaling pathways involved in mitosis?

Introduction

Mitosis is a highly regulated process that ensures accurate chromosome segregation and proper cell division. This complex process involves the coordination of multiple signaling pathways, including the DNA damage response (DDR) pathway, the spindle assembly checkpoint (SAC), and the mitogen-activated protein kinase (MAPK) signaling pathway. These pathways work together to maintain genomic stability and prevent the formation of abnormal cells.ref.72.1 ref.72.2 ref.72.1 In this essay, we will explore the key proteins and mechanisms involved in each of these signaling pathways and their roles in mitotic progression.ref.72.1 ref.24.3 ref.72.4

The DNA Damage Response Pathway

The DDR pathway is activated in response to DNA damage, including double-strand breaks (DSBs). It plays a crucial role in repairing DNA damage during mitosis and maintaining genomic integrity. Several proteins are involved in the DDR pathway, including ATM, ATR, H2AX, and MDC1.ref.71.26 ref.69.1 ref.69.1

ATM and ATR are kinases that are activated by the MRN complex, which consists of Mre11, Rad50, and Nbs1. The MRN complex recognizes and processes DNA damage, leading to the activation of ATM and ATR. ATM phosphorylates the histone variant H2AX, generating g-H2AX, which promotes the recruitment of MDC1 to the site of damage.ref.14.4 ref.24.6 ref.73.13 MDC1 acts as an amplification factor for the DDR signal by recruiting the MRN-ATM complex to the damage site, leading to further phosphorylation of adjacent H2AX molecules. MDC1 also acts as an interaction platform for other DDR components, facilitating the recruitment of chromatin remodelers and ubiquitin ligase complexes.ref.14.4 ref.14.4 ref.71.26

The activation of ATM and ATR leads to the phosphorylation and activation of checkpoint kinases CHK1 and CHK2. CHK1 and CHK2 play important roles in the maintenance of the cell cycle checkpoint and inhibit cyclin-dependent kinases, leading to cell cycle arrest. These proteins and pathways are crucial for repairing DNA damage during mitosis and ensuring the maintenance of genomic integrity.ref.14.4 ref.14.4 ref.14.5

The Spindle Assembly Checkpoint

The SAC is a surveillance mechanism that ensures accurate chromosome segregation during mitosis. It is activated when kinetochores, protein structures on chromosomes, are not properly attached to spindle microtubules. The SAC involves the action of proteins such as Mad1, Mad2, Bub1, Bub3, and Cdc20.ref.87.44 ref.63.2 ref.87.42

Mad1 and Mad2 are involved in detecting unattached kinetochores and generating a diffusible signal that inhibits the Anaphase Promoting Complex/Cyclosome (APC/C). The APC/C is responsible for promoting chromosome segregation. Bub1 and Bub3 are also involved in the SAC and are bound to kinetochores early in mitosis.ref.87.42 ref.22.1 ref.66.3 BubR1, also known as Mad3, is a kinase that interacts with Mad2 and Bub3 to form the Mitotic Checkpoint Complex (MCC). The MCC physically inhibits APC/C by binding to its co-activator Cdc20. Cdc20 is normally a co-activator of APC/C, but it is inhibited by the MCC until all chromosomes are properly attached to kinetochores.ref.25.3 ref.66.3 ref.87.42

These proteins work together to ensure that chromosomes are accurately segregated during mitosis by delaying the onset of anaphase until all kinetochores are correctly attached to the mitotic spindle. This mechanism prevents chromosomal mis-segregation and the development of aneuploidy, which can lead to genetic disorders and cancer.ref.87.2 ref.30.1 ref.59.2

The MAPK Signaling Pathway

The MAPK signaling pathway is involved in various cellular processes, including mitotic spindle assembly. MAPKs, such as ERK1/2, p38, and JNK/SAPK, can regulate microtubule dynamics and ensure proper cell division. Phosphorylation of specific MAPK substrates, such as MAP65 proteins, can affect microtubule stability and bundling.ref.51.1 ref.51.5 ref.51.4

The MAPK signaling pathway acts downstream of various upstream signals, including growth factors and stress signals. Activation of MAPKs leads to the phosphorylation of downstream targets, including MAP65 proteins, which are involved in microtubule stabilization and organization. Proper microtubule dynamics are crucial for the assembly and alignment of the mitotic spindle, ensuring accurate chromosome segregation.ref.51.4 ref.51.1 ref.51.4

Integration of Signaling Pathways

The DDR pathway, SAC, and MAPK signaling pathway work together to ensure accurate chromosome segregation and proper cell division during mitosis. The DDR pathway plays a role in repairing DNA damage during mitosis and can impact mitotic progression and cell fate in the presence of DNA damage. The SAC ensures that sister chromatids are correctly attached to the mitotic spindle before allowing the onset of anaphase, preventing genetic instability and errors in chromosome segregation.ref.72.9 ref.72.4 ref.72.18 The MAPK signaling pathway regulates microtubule dynamics and plays a role in mitotic spindle assembly. Integration of these signaling pathways is crucial for maintaining genomic stability and preventing the formation of abnormal cells.ref.25.1 ref.72.1 ref.24.3

Consequences of SAC Failure

If the SAC fails to delay anaphase onset until all chromosomes are properly aligned and attached to the spindle, it can lead to genetic instability and errors in chromosome segregation. The SAC is responsible for ensuring the fidelity of chromosome segregation during mitosis. If a single kinetochore is unattached or weakly attached, it generates a signal that inhibits the progression of the cell cycle.ref.63.2 ref.86.8 ref.72.3 This signal involves proteins such as Mad1, Mad2, BubR1, Bub3, and Cdc20.ref.25.2 ref.63.2 ref.66.3

The inhibition of anaphase-promoting complexes (APCs) prevents the ubiquitination of proteins required for chromatid cohesion and the mitotic state. If the SAC fails, it can result in premature separation of sister chromatids and errors in chromosome segregation, leading to genetic instability and potential cell death. The consequences of SAC failure can vary depending on the specific context, such as the presence of DNA damage or exposure to spindle poisons.ref.25.2 ref.87.42 ref.66.3 However, it is generally understood that the SAC plays a crucial role in maintaining the fidelity of chromosome segregation during mitosis.ref.87.42 ref.30.2 ref.63.2

Conclusion

In conclusion, the key signaling pathways involved in mitosis, including the DDR pathway, SAC, and MAPK signaling pathway, play important roles in ensuring accurate chromosome segregation and proper cell division. The DDR pathway repairs DNA damage during mitosis and maintains genomic integrity. The SAC monitors the attachment of kinetochores to the mitotic spindle and delays anaphase onset until all chromosomes are properly aligned and attached.ref.72.17 ref.72.4 ref.72.18 The MAPK signaling pathway regulates microtubule dynamics and contributes to proper mitotic spindle assembly. These pathways work together to maintain genomic stability and prevent the formation of abnormal cells. Understanding the intricate mechanisms of these signaling pathways in mitosis is crucial for furthering our knowledge of cell division and its implications in various diseases, including cancer.ref.72.1 ref.25.1 ref.24.3

How do these pathways regulate cell cycle progression and mitotic events?

Introduction

The regulation of cell cycle progression and mitotic events is a complex process that ensures the accurate duplication and segregation of chromosomes during cell division. Positive feedback is a key mechanism that insulates mitosis from variability in other cell cycle phases, allowing for the short and constant duration of mitosis. The activation and deactivation cycles of cyclin-dependent kinases (Cdks) and the synthesis and degradation of regulatory cyclins play a crucial role in regulating cell cycle progression.ref.7.37 ref.7.22 ref.7.35 In addition, the spindle assembly checkpoint (SAC) and the DNA damage response (DDR) contribute to the control of mitotic progression and cell fate. Although the exact mechanisms and interactions between these pathways are still being studied, significant progress has been made in understanding their roles in mitotic regulation.ref.2.4 ref.7.2 ref.7.22

Regulation of Mitotic Events

A. Role of Regulatory Cyclins Regulatory cyclins, such as Cyclin B, play a crucial role in the regulation of mitotic events. The activity of Cdk1, in complex with Cyclin B1, is required for the initiation and progression of mitosis.ref.9.4 ref.9.34 ref.9.4 The switch-like increase in Cyclin B activity enables entry into mitosis, while subsequent proteolysis of Cyclin B promotes exit from mitosis. The activation of Cyclin B is regulated by a positive feedback loop involving the activating phosphatase Cdc25 and the inhibitory kinase Wee1. This positive feedback loop ensures the proper timing and duration of mitosis, allowing for the faithful segregation of chromosomes and completion of cell division.ref.9.10 ref.9.9 ref.9.6

The activation and deactivation cycles of Cdks during the cell cycle are regulated by various mechanisms. The activation of Cyclin D during late G1 and subsequent activation of Cyclin E indicate entry into S phase. This is followed by step-wise activation of E2F via Cyclin D and Cyclin E, leading to the sequential transcription of Cyclin A in the S phase and Cyclin B during mitosis.ref.9.34 ref.9.9 ref.9.6 CKI is activated in early G1 and is maximally elevated at late S phase. E2F activity peaks at S phase. APCCdh1 is active during late mitosis and early G1 phase, degrading Cyclin A and Cyclin B.ref.9.9 ref.9.34 ref.9.9 APCCdc20, which is a degradator of mitotic substrates, is activated during late mitosis, once Cyclin B is sufficiently activated. The degradation of G1-S cyclins is mediated by SCF, which becomes active at the end of S phase, leading to the degradation of Cyclin D and Cyclin E. These activation and deactivation cycles are regulated by various mechanisms, including transcriptional regulation, association with protein inhibitors, phosphorylation/dephosphorylation, and cyclin degradation.ref.9.9 ref.9.6 ref.9.9

Spindle Assembly Checkpoint (SAC)

A. Mechanism of SAC The spindle assembly checkpoint (SAC) ensures accurate chromosome segregation during mitosis by monitoring the attachment of kinetochores to the mitotic spindle. The SAC prevents the progression from metaphase to anaphase until all kinetochores are properly attached to microtubules.ref.63.2 ref.22.0 ref.86.8 If even a single kinetochore is unattached, it can signal to the rest of the cell and prevent cell cycle progression. The SAC generates a diffusible signal called the mitotic checkpoint complex (MCC), which inhibits the Anaphase Promoting Complex/Cyclosome (APC/C) and prevents the degradation of securin and cyclin B. This inhibition delays the onset of anaphase and ensures that all chromosomes are properly aligned before segregation occurs.ref.87.42 ref.66.3 ref.63.2

There are two proposed mechanisms for how the SAC signal is generated and propagated. One model suggests that the signal is generated through diffusive sequestration, where unattached kinetochores release the MCC into the cytoplasm, inhibiting the APC/C. This model requires a two-stage signal amplification cascade to generate sufficient inhibition.ref.25.4 ref.25.4 ref.66.3 The other model proposes that the signal is propagated through spatial gradients of an inhibitory signal that is transported along spindle microtubules. This active transport mechanism allows for rapid signal propagation and is supported by experiments. The specific mechanism of signal generation and propagation may involve diffusive sequestration or active transport along spindle microtubules.ref.25.18 ref.25.0 ref.25.5

DNA Damage Response (DDR)

A. Role of DDR in Mitotic Progression and Cell Fate The DNA damage response (DDR) plays a crucial role in the control of mitotic progression and cell fate. Mitosis is tightly regulated, and DNA damage sensing and repair are thought to be repressed during mitosis to prevent telomere fusion.ref.69.1 ref.69.1 ref.72.1 However, there is increasing evidence that mitotic progression is tightly controlled in the presence of DNA damage. DNA double-strand breaks (DSBs) are one of the most deleterious types of DNA damage, and the repair mechanisms during mitosis include nonhomologous end joining (NHEJ), alternative end joining (Alt-EJ), and single-strand annealing (SSA) pathways.ref.72.2 ref.69.1 ref.69.3

The DDR and the SAC have crosstalk and contribute to the control of mitotic progression and cell fate. The DDR proteins ATM and ATR are required for mitotic arrest following DNA damage, and there is evidence for the existence of a mitotic DNA damage checkpoint. Mitotic cells are capable of DNA repair, and repair is attenuated when DNA-PKcs and ATM are simultaneously compromised.ref.72.12 ref.24.3 ref.67.4 Mitotic DDR is more complex than previously thought and may involve factors from multiple repair pathways. The exact mechanisms by which the DDR communicates with the SAC to instigate cell cycle arrest are still unclear.ref.72.18 ref.72.12 ref.71.26

Conclusion

In conclusion, the regulation of cell cycle progression and mitotic events involves a complex series of signaling pathways. Positive feedback ensures the short and temporally insulated duration of mitosis. The activation and deactivation cycles of Cdks and the synthesis and degradation of regulatory cyclins play a crucial role in regulating cell cycle progression.ref.7.22 ref.7.37 ref.7.35 The SAC ensures accurate chromosome segregation by monitoring kinetochore attachment to the mitotic spindle, while the DDR responds to DNA damage during mitosis. The exact mechanisms and interactions between these pathways are still being studied and understood. Further research is needed to fully elucidate the intricate mechanisms involved in the regulation of mitotic events.ref.7.2 ref.2.4 ref.7.22

What are the upstream signals that initiate mitotic signaling?

The Role of MAPKs in Regulating Microtubule Dynamics during Cell Division

MAPKs, or Mitogen-Activated Protein Kinases, are involved in initiating mitotic signaling and play a crucial role in regulating microtubule dynamics during cell division. These upstream signals that activate MAPKs include the phosphorylation of specific SAFs, or Spindle Assembly Factors, by mitotic kinases. MAPKs have been found to be essential for the stability of the mitotic spindle and proper microtubule dynamics.ref.51.4 ref.51.1 ref.51.5

Studies have shown that MAPKs are involved in processes such as mitotic spindle formation and cytokinesis, both of which require precise microtubule organization and dynamics. In Arabidopsis, MAPKs have been found to localize to the phragmoplast during cytokinesis, suggesting their involvement in microtubule organization at this critical stage of cell division. The proper functioning of the cell division process relies on the stability and dynamics of microtubules, which are regulated by MAPKs.ref.51.4 ref.51.0 ref.51.5

Phosphorylation of SAFs, such as TPX2, by mitotic kinases can regulate their function in mitotic spindle assembly. TPX2 is a specific example of an SAF that is phosphorylated by mitotic kinases to modulate its role in mitotic spindle assembly. Phosphorylated SAFs, including TPX2, can influence the enzymatic activities, localization, and functions of mitotic kinases.ref.19.6 ref.19.5 ref.19.1 This phosphorylation event plays a critical role in controlling the activity of SAFs and their involvement in microtubule dynamics during cell division.ref.19.6 ref.19.26 ref.19.1

It is important to note that not all plant species may engage the MAPK signaling pathway to regulate microtubule dynamics during cell division. The role of MAPKs in regulating microtubule dynamics may vary among different plant species and may involve other factors such as CDKs (cyclin-dependent kinases). Further research is needed to investigate the specific processes controlled by the MAPK signaling pathway and the differences in its regulation among plant species.ref.51.6 ref.51.5 ref.51.24

Examples of MAPKs Involved in Mitotic Signaling Pathways

There are several specific examples of MAPKs that are known to be involved in mitotic signaling pathways. These include MPK4 and MPK6 in Arabidopsis, as well as MAPKs in Xenopus egg extracts. These MAPKs play a crucial role in regulating microtubule dynamics during cell division and ensuring an unperturbed cell division process.ref.51.4 ref.51.5 ref.51.0 Studies on mutants of the MAPK signaling pathway in plants have shown failures in mitotic spindle formation and cytokinesis, leading to the formation of bi- and multinucleate cells. However, it is important to note that the specific functions and roles of MAPKs in mitotic signaling pathways may vary among different plant species.ref.51.4 ref.51.5 ref.51.5

The Role of SAFs, Specifically TPX2, in Mitotic Spindle Assembly

SAFs, or Spindle Assembly Factors, are essential for the proper assembly of the mitotic spindle. TPX2 is a specific SAF that plays a crucial role in regulating mitotic spindle assembly. The phosphorylation of TPX2 by mitotic kinases can regulate its function in mitotic spindle assembly.ref.19.6 ref.19.5 ref.19.2

Phosphorylated TPX2, along with other phosphorylated SAFs, can influence the enzymatic activities, localization, and functions of mitotic kinases. This phosphorylation event plays a critical role in controlling the activity of TPX2 and its involvement in mitotic spindle assembly. The phosphorylation of TPX2 by mitotic kinases can regulate its function in mitotic spindle assembly by influencing its interactions with other proteins and its localization to the mitotic spindle.ref.19.6 ref.19.1 ref.19.5

The release of SAFs, including TPX2, from inhibitory complexes with importin-α1 is crucial for the proper formation of the mitotic apparatus. Importin-α1 is a key partner and regulator of SAFs containing a nuclear localization signal (NLS) motif. The phosphorylation of importin-α1 induces the release of NLS-containing SAFs, such as TPX2, to load onto the mitotic spindle, promoting their function in mitotic spindle assembly.ref.19.25 ref.19.24 ref.19.23

The spatial control of SAF interactions is critical for the proper formation of the mitotic apparatus. Microtubules (MTs) need to be specifically nucleated and stabilized only in the region of spindle formation, and proteins have to be recruited to the spindle for the regulation of chromosome segregation. The release of SAFs from inhibitory complexes with importin-α1 is directed by chromosomes through chromatin-generated Ran-GTP, which exists at a high concentration in the vicinity of chromosomes and a lower concentration farther away.ref.19.25 ref.19.2 ref.19.24 Disruption of the Ran-GTP gradient compromises the spatial organization of spindle assembly.ref.19.25 ref.19.2 ref.19.5

In conclusion, MAPKs play a crucial role in regulating microtubule dynamics during cell division. Their activation is initiated by upstream signals, including the phosphorylation of specific SAFs by mitotic kinases. MAPKs are involved in processes such as mitotic spindle formation and cytokinesis, and their proper functioning is essential for the stability of the mitotic spindle and proper microtubule dynamics.ref.51.4 ref.51.5 ref.51.21 Examples of MAPKs involved in mitotic signaling pathways include MPK4 and MPK6 in Arabidopsis, as well as MAPKs in Xenopus egg extracts. The role of SAFs, specifically TPX2, in mitotic spindle assembly is regulated by the phosphorylation of TPX2 by mitotic kinases. The phosphorylation of TPX2 and other SAFs influences their function in mitotic spindle assembly and their interactions with other proteins.ref.51.4 ref.19.5 ref.51.5 The release of SAFs from inhibitory complexes with importin-α1 is directed by chromosomes through chromatin-generated Ran-GTP, ensuring the proper spatial organization of spindle assembly. Further research is needed to investigate the specific processes controlled by the MAPK signaling pathway and the differences in its regulation among different plant species.ref.19.2 ref.19.2 ref.19.5

How do these signaling pathways interact and cross-regulate each other?

Mitotic Signaling Pathways and Their Functions

Mitotic signaling pathways play a crucial role in regulating the cell cycle and ensuring proper cell division. These pathways involve a complex network of signaling molecules and interactions that ultimately control mitosis. Here, we will discuss some examples of mitotic signaling pathways and their specific functions.ref.72.1 ref.7.2 ref.72.1

1. TGF-β pathway: The TGF-β pathway consists of two branches, Smad-dependent and Smad-independent signaling. When extracellular TGF-β ligands bind to membrane receptors, the Smad proteins are phosphorylated, leading to their translocation to the nucleus.ref.77.31 ref.77.31 ref.77.31 In the nucleus, Smad proteins induce the expression of specific genes that can either upregulate or downregulate mitosis. Additionally, TGF-β signaling can modify the composition of the extracellular matrix. Moreover, TGF-β can also modulate apoptotic processes through its interaction with the MAPK pathway.ref.77.31 ref.77.31 ref.77.31 The TGF-β pathway can exhibit either pro- or anti-oncogenic activity, contributing to resisting cell death and sustaining proliferative signaling. High levels of TGF activity are associated with tissue invasion, metastasis, and angiogenesis. Disruption to TGF signaling, often due to mutated membrane receptors, is frequently observed in colorectal cancer (CRC).ref.77.31 ref.77.30 ref.77.27

2. MAPK signaling pathway: The MAPK signaling pathway is involved in cell differentiation and growth. Extracellular Growth Factors (EGFs) bind to cell surface receptors, which then recruit a complex of signaling proteins.ref.77.31 ref.51.3 ref.8.2 This complex assists in the phosphorylation of Ras, initiating a cascade of kinase phosphorylations involving Raf, MEK, and ERK/MAPK. The activated ERK proteins then translocate to the nucleus, where they induce chromatin remodeling and transcription of target genes. Mutations in the MAPK signaling pathway can lead to failures in mitotic spindle formation and cytokinesis, resulting in the formation of bi- and multinucleate cells.ref.77.31 ref.77.32 ref.77.32

3. β-catenin/Wnt signaling pathway: The β-catenin/Wnt signaling pathway involves β-catenin, a mediator of Wnt signaling, which is associated with adherens junctions (AJs) at the cell surface. Contact inhibition, a process that regulates cell proliferation, has been reported for NF2/Merlin, a tumor suppressor gene. NF2/Merlin acts via the Hippo kinase pathway, controlling the nuclear localization of the transcriptional activator YAP.ref.78.2 ref.78.3 ref.77.27 The MAPK pathway is also implicated in contact inhibition and promotes cell cycle entry by regulating the expression of cyclin D1. Nectins, a family of cell adhesion molecules, are also involved in the regulation of cell motility and proliferation.ref.78.3 ref.78.3 ref.78.2

4. Mitotic spindle-associated MAPKs: MAPKs have been found to play a role in the formation and stability of the mitotic spindle. Activated MAPKs localize to the phragmoplast during cytokinesis, ensuring proper cell division.ref.51.4 ref.51.5 ref.51.0 Inactivation of MAPKs can lead to failures in mitotic spindle formation and cytokinesis, resulting in the formation of bi- and multinucleate cells.ref.51.4 ref.51.5 ref.51.5

Role of MAPK Signaling Pathways in Mitosis

The role of MAPK signaling pathways in mitosis has been extensively studied in various plant species. In plants such as Lupinus luteus, Pisum sativum, Vicia faba, and Lycopersicon esculentum, activated MAPKs have been observed to localize in the nucleus of both interphase and mitotic cells. Furthermore, in some species, activated MAPKs have been found to co-localize with mitotic microtubules, which are essential for proper cell division.ref.51.0 ref.51.0 ref.51.5 However, the extent of co-localization varies among different plant species. For example, L. esculentum shows the most evident co-localization of activated MAPKs with microtubules, while P.ref.51.13 ref.51.0 ref.51.12 sativum and V. faba display this co-localization in only about 50% of mitotic cells. On the other hand, L.ref.51.0 ref.51.12 ref.51.12 luteus does not show clear immunofluorescence signals at spindle microtubules and phragmoplast during mitosis. These findings suggest that not all plants engage the MAPK signaling pathway to regulate microtubule dynamics during cell division.ref.51.1 ref.51.0 ref.51.13

Consequences of Disruptions in Mitotic Signaling Pathways

When mitotic signaling pathways fail to interact or cross-regulate properly, it can have several consequences or potential disruptions. Firstly, chromosome mis-segregation during mitosis can occur, leading to the formation of aneuploid cells. Aneuploidy is known to be a hallmark of cancer and is associated with various serious diseases.ref.79.0 ref.72.1 ref.79.0 Secondly, disruptions in mitotic signaling pathways can affect the timing of exit from mitosis. The mitotic or spindle assembly checkpoint regulates cell cycle progression from metaphase to anaphase. Failure to properly regulate this checkpoint can result in the premature exit from mitosis, leading to abnormal cell division.ref.68.3 ref.72.1 ref.79.0 Thirdly, disruptions in mitotic signaling pathways can lead to mitotic catastrophe. Mitotic catastrophe is an intrinsic oncosuppressive mechanism that senses mitotic failure and responds by driving a cell to an irreversible antiproliferative fate of death or senescence. Fourthly, disruptions in mitotic signaling pathways can impact the induction of apoptosis during mitosis.ref.72.1 ref.68.3 ref.68.3 There is crosstalk between the spindle assembly checkpoint (SAC) and the apoptotic machinery, and disruptions in this crosstalk can affect the proper execution of apoptosis during mitosis. Lastly, disruptions in mitotic signaling pathways can affect the proper ordering of cell-cycle phases and the full completion of one phase before the onset of the next. This can lead to cell cycle defects and abnormal cell division.ref.68.3 ref.72.1 ref.72.13

Feedback Loops in Mitotic Signaling Pathways

There are known feedback loops within mitotic signaling pathways that play a crucial role in maintaining the proper regulation of mitosis. Positive feedback loops involving Cdc25, Wee1, and Cdk1-cyclin B1 have been identified. These feedback loops can give rise to bistability, a phenomenon in which cells can exist in either an active or inactive state.ref.7.40 ref.7.37 ref.7.22 Positive feedback regulation is important for keeping mitotic events synchronized and ensuring a short, constant duration of mitosis. It also generates temporal insulation and brings about modularity in mitosis. Positive feedback regulation is responsible for the short, constant duration of mitosis and its temporal insulation from earlier cell-cycle events.ref.7.35 ref.7.22 ref.7.0 It is crucial for the fidelity of cell division and cell survival. Moreover, positive feedback regulation may underlie temporal modularity in mitosis. It accounts for the experimentally observed short, constant duration of mitosis in somatic cells and for the temporal insulation of mitotic duration from variability in the duration of upstream cell-cycle events.ref.7.35 ref.7.10 ref.7.22 In summary, positive feedback regulation is a key mechanism to ensure that the duration of mitosis is kept short and constant, and it brings about temporal modularity in mitosis.ref.7.35 ref.7.0 ref.7.10

How are mitotic signaling pathways dysregulated in cancer and other diseases?

Dysregulation of Mitotic Signaling Pathways in Cancer and Other Diseases

Dysregulation of mitotic signaling pathways in cancer and other diseases can occur through various mechanisms. One key pathway involved is the Wnt pathway, which governs proliferation and cellular adhesion. Dysregulation of this pathway can disrupt mitosis, leading to sustained proliferative signaling and enabling replicative immortality.ref.78.3 ref.77.27 ref.77.27 The Wnt pathway has also been implicated in activating invasion and metastasis and resisting cell death.ref.77.27 ref.77.27 ref.77.28

Another pathway involved is the DNA damage response (DDR) pathway. DNA damage in mitosis can lead to activation of the spindle assembly checkpoint (SAC), which delays segregation of chromosomes until they are properly attached to the mitotic spindle. Dysregulation of the DDR pathway can result in increased segregation errors during mitosis, leading to mitotic catastrophe and altered cell fate.ref.72.2 ref.72.1 ref.72.4

Additionally, the mitotic checkpoint, which regulates the timing of exit from mitosis, can be targeted to kill tumor cells. Inhibition of the mitotic checkpoint can lead to increased segregation errors and cell death in tumor cells.ref.79.0 ref.79.1 ref.79.0

Overall, dysregulation of mitotic signaling pathways in cancer and other diseases can result in disrupted mitosis, increased segregation errors, and altered cell fate. Further research is needed to fully understand the molecular mechanisms and potential therapeutic targets associated with these dysregulations.ref.72.1 ref.79.0 ref.79.0

Dysregulation of the Wnt Pathway and Disrupted Mitosis

The dysregulation of the Wnt pathway disrupts mitosis and leads to sustained proliferative signaling. The Wnt pathway, specifically its canonical form, governs proliferation and cellular adhesion. When there are sufficiently high concentrations of extracellular Wnt, it inhibits the destruction complex of β-catenin, allowing cytoplasmic β-catenin to translocate to the nucleus and upregulate Wnt target genes involved in cell cycle progression, such as cyclin D.ref.10.4 ref.77.27 ref.78.3 This disruption of mitosis contributes to sustained proliferative signaling.ref.10.33 ref.10.41 ref.10.33

Neoplastic cells often display addiction to Wnt signaling, and Lgr5 expression, associated with Wnt pathway activity, is upregulated in colorectal tumor tissue. The Wnt pathway interacts with other networks, such as Notch and EGF, further influencing cell development and proliferation. Mathematical modeling has aided in understanding Wnt signaling, and the pathway has been implicated in CRC initiation.ref.77.28 ref.77.27 ref.77.27

The dysregulation of the Wnt pathway, along with mechanical strain, synergistically activates β-catenin-mediated transcription and cell cycle progression through mitosis. Mechanical strain induces Src-dependent phosphorylation of β-catenin, and Wnt3A activation increases β-catenin-mediated transcription. The combination of mechanical strain and Wnt/b-catenin activation triggers cells in S/G2 to divide.ref.10.1 ref.10.1 ref.10.6 This suggests that local Wnt signaling fine-tunes the effects of global mechanical strain to restrict cell divisions during tissue development and homeostasis.ref.10.1 ref.10.41 ref.10.5

Dysregulated Signaling Pathways in Cancer and Other Diseases

Besides the Wnt pathway, other signaling pathways that are dysregulated in cancer and other diseases include the Notch pathway, TGF-β pathway, Ras-MAPK pathway, and neural signaling pathways.ref.77.28 ref.77.27 ref.77.27

The Notch pathway regulates cell development and influences the distribution of cell types within the crypt epithelium. It interacts with the Wnt pathway and is believed to be a vital step in colorectal cancer (CRC) initiation. The protein β-catenin acts as a hub linking the Wnt and Notch pathways.ref.77.28 ref.77.29 ref.77.28

The TGF-β pathway has two branches, Smad-dependent and Smad-independent signaling. It modulates mitosis, modifies the composition of the extracellular matrix, and contributes to resisting cell death and sustaining proliferative signaling. Disruption to TGF signaling in CRC is often due to mutated membrane receptors.ref.77.31 ref.77.31 ref.77.31

The Ras-MAPK pathway is associated with cell differentiation and growth. Extracellular Growth Factors (EGFs) bind to a receptor on the cell surface membrane, leading to a series of kinase phosphorylations that induce chromatin remodeling and transcription of target genes.ref.77.31 ref.77.31 ref.77.32

Neural signaling pathways, specifically the sympathetic nervous system (SNS) pathways, have been shown to impact cancer progression. Activation of SNS pathways promotes tumor growth and metastatic dissemination, while inhibition of β-adrenergic signaling with β-blockers prevents these adverse effects.ref.107.7 ref.107.2 ref.107.10

These dysregulated signaling pathways contribute to various hallmarks of cancer, such as sustaining proliferative signaling, activating invasion and metastasis, inducing angiogenesis, and resisting cell death.ref.107.6 ref.107.7 ref.107.6

Increased Segregation Errors during Mitosis and Chromosomal Mis-segregation

The consequences of increased segregation errors during mitosis can lead to chromosomal mis-segregation, which can result in cancer and other serious diseases. Dysregulation of the DNA damage response pathway can contribute to increased segregation errors during mitosis.ref.72.1 ref.72.2 ref.72.1

The DNA damage response (DDR) is largely suppressed during mitosis, but there is increasing evidence to suggest tight control of mitotic progression in the incidence of DNA damage. Mitotic cells are capable of DNA repair, and repair mechanisms during mitosis include error-prone nonhomologous end joining (NHEJ), alternative end joining (Alt-EJ), and single-strand annealing (SSA) pathways.ref.72.2 ref.69.1 ref.72.1

The DDR and the spindle assembly checkpoint (SAC) have crosstalk and play a role in controlling mitotic progression and cell fate in the presence of mitotic DNA damage. The DDR senses mitotic DNA damage and activates the SAC, leading to cell cycle arrest. The exact mechanism by which the DDR communicates with the SAC to instigate cell cycle arrest is still unclear.ref.72.9 ref.72.18 ref.72.17

The ATM and ATR pathways are involved in the DDR and can impact mitotic progression. The ATM and ATR pathways have been shown to affect centrosome-driven spindle assembly, and the phosphorylation of XCEP63 by ATM and ATR delays spindle assembly. This pathway may contribute to regulating DNA repair or mitotic cell survival in the presence of chromosomal breakage.ref.24.0 ref.24.3 ref.24.6

Targeting the Dysregulated Mitotic Checkpoint for Killing Tumor Cells

The dysregulation of the mitotic checkpoint can affect the timing of exit from mitosis. The mitotic checkpoint, also known as the spindle assembly checkpoint (SAC), is responsible for ensuring that all chromosomes are properly attached to the mitotic spindle before the cell progresses to anaphase. When the checkpoint is dysregulated, cells may exit mitosis prematurely, leading to chromosome missegregation and aneuploidy, which can contribute to the development of cancer.ref.7.14 ref.63.2 ref.59.2

Targeting the dysregulated mitotic checkpoint for killing tumor cells is a potential anti-cancer strategy. Inhibition of the mitotic checkpoint can lead to increased segregation errors during mitosis, which can induce cell death in tumor cells. By inactivating essential checkpoint components, such as Mps1, the mitotic checkpoint can be disrupted, resulting in increased cell death in tumor cells.ref.79.8 ref.79.0 ref.79.0

In conclusion, dysregulation of mitotic signaling pathways in cancer and other diseases can disrupt mitosis, increase segregation errors, and alter cell fate. The dysregulation of the Wnt pathway, DNA damage response pathway, and mitotic checkpoint can all contribute to these effects. Understanding the molecular mechanisms and potential therapeutic targets associated with these dysregulations is crucial for the development of effective treatments.ref.72.1 ref.72.1 ref.24.3 Further research is needed to fully elucidate the complexities of these pathways and their interactions in order to advance our knowledge and improve patient outcomes.ref.24.3 ref.72.1 ref.79.1

Evolutionary Conservation of Mitotic Mechanisms

How conserved are the molecular mechanisms of mitosis across different organisms?

Introduction to the Molecular Mechanisms of Mitosis

Mitosis is a fundamental process in cell division, ensuring the accurate segregation of replicated chromosomes into two daughter cells. The molecular mechanisms underlying mitosis have been extensively studied and are known to be highly conserved across different organisms. However, the degree of conservation varies, and ongoing research aims to further understand the molecular bases of the cell response to mitotic-targeting molecules.ref.40.2 ref.87.1 ref.7.2 This essay will discuss the conserved molecular mechanisms of mitosis, the evolutionary advantages of their conservation, and current areas of ongoing research in this field.ref.40.2 ref.72.1 ref.32.2

Conserved Molecular Mechanisms of Mitosis

Numerous studies have identified specific examples of conserved molecular mechanisms of mitosis across different organisms. One example is the differential deployment of conserved biochemical modules. For instance, midzone pushing and braking via inter-polar microtubule (ipMT) crosslinking, cortical pulling, ipMT plus end dynamics/net polymerization, and minus end depolymerization/poleward flux have been observed in various organisms, including the nematode Caenorhabditis elegans.ref.28.63 ref.28.9 ref.28.24 These mechanisms contribute to the proper assembly and function of the mitotic spindle, which is crucial for chromosome segregation.ref.28.52 ref.28.30 ref.28.51

The regulation of anaphase B, another essential process in mitosis, is also an area of interest for future studies. Anaphase B involves the separation of spindle poles, mediated by the sliding of overlapping anti-parallel microtubules. The conservation of these mechanisms across different organisms indicates their fundamental role in ensuring accurate chromosome segregation during mitosis.ref.28.1 ref.28.45 ref.86.7 These conserved molecular mechanisms have likely been maintained throughout evolution due to their vital functions in cell division.ref.28.63 ref.28.63 ref.28.63

Evolutionary Advantages of Conserved Mitotic Mechanisms

The conservation of mitotic mechanisms across different organisms offers evolutionary advantages. Eukaryotes have developed diverse approaches to chromosome segregation, yet the mechanochemical mechanisms of individual motors are common threads that link seemingly divergent systems. This conservation allows for reliable and accurate chromosome segregation during mitosis.ref.91.3 ref.91.2 ref.28.63 The individual motors involved in mitotic processes, such as kinesins, play crucial roles in mediating the movements of microtubules and chromosomes. The conservation of these motors ensures the efficient and coordinated functioning of the spindle apparatus.ref.91.3 ref.91.1 ref.91.1

Recent work has highlighted the need to understand the mechanics of individual motors and subsystems of the spindle. In vitro reconstitution of kinesin-microtubule interactions has revealed unexpected complexity. This emphasizes the importance of grounding the field firmly in knowledge of the mechanics of individual motors.ref.91.28 ref.91.1 ref.91.28 The conservation of mitotic mechanisms allows for the precise coordination of motor activities, leading to successful chromosome segregation. These mechanisms have likely been conserved to ensure accurate cell division throughout evolution.ref.91.1 ref.91.28 ref.91.3

Ongoing Research in the Molecular Bases of the Cell Response to Mitotic-Targeting Molecules

Understanding the molecular bases of the cell response to mitotic-targeting molecules is crucial for developing effective cancer treatments and improving therapeutic strategies. Several areas of ongoing research are focused on this topic:ref.88.2 ref.88.1 ref.88.2

1. Impact of gene expression profiles: Researchers are investigating how gene expression profiles influence the effectiveness of mitotic-targeting agents in different cancer types and within cells from the same cancer. This knowledge can help identify specific genetic markers that contribute to drug resistance and guide personalized treatment approaches.ref.88.2 ref.88.2 ref.88.2

2. Development of resistance: The development of resistance to mitotic-targeting agents is a significant challenge in cancer therapy. Scientists are studying various mechanisms that contribute to drug resistance, such as the induction of efflux pump mechanisms and mutations or differential expression of tubulin isotypes.ref.88.2 ref.88.1 ref.88.27 Understanding these mechanisms can aid in the development of strategies to overcome resistance and improve treatment outcomes.ref.88.2 ref.88.2 ref.88.1

3. Determinants of mitotic death induction: In vivo imaging methods are being used to track mitotic cell fates at the single-cell level. This research aims to identify the determinants of mitotic death induction, which can help predict and enhance the effectiveness of mitotic-targeting agents in cancer treatment.ref.88.2 ref.24.3 ref.24.3

4. Effects of DNA damage on mitosis: Researchers are exploring the effects of DNA damage on mitosis and the complex behavior of mitotic cells challenged with different drugs. These effects can include arrest, endoreduplication of the genome, and cell death.ref.69.1 ref.72.1 ref.24.3 Understanding the molecular mechanisms underlying these responses can provide insights into the development of more targeted and effective therapies.ref.69.1 ref.69.1 ref.24.3

5. Response of cancer cell lines to mitotic-targeting agents: Characterizing the response of cancer cell lines to mitotic-targeting agents is crucial for understanding their efficacy and identifying distinct cell fate profiles. This research can uncover cytological phenotypes, MCL-1 down-regulation, caspase-3 activation, and other cellular responses that contribute to cell death during mitosis.ref.88.1 ref.88.2 ref.88.2

6. Mechanisms controlling apoptosis during mitosis: Investigating the mechanisms controlling the induction of apoptosis during mitosis is essential for understanding the relationship between mitotic progression, DNA damage, and cell fate. This research can provide insights into the development of novel strategies to selectively induce apoptosis in cancer cells during mitosis.ref.72.1 ref.68.3 ref.72.1

7. Detailed mechanisms of individual force generators in the spindle: Understanding the detailed mechanisms of individual force generators in the spindle is crucial for comprehending the emergent self-organization and complex mechanics of the spindle machinery. This research aims to elucidate how these force generators interact with each other and regulate microtubule dynamics to ensure proper chromosome segregation.ref.91.28 ref.91.1 ref.91.2

8. Control of mitosis in the event of DNA damage: Researchers are studying the control of mitosis in the presence of DNA damage. This includes investigating the activation of DNA damage checkpoints and the repair mechanisms utilized during mitosis.ref.72.1 ref.72.1 ref.72.2 Understanding how cells respond to DNA damage during mitosis can provide insights into the maintenance of genome integrity and the development of targeted therapies.ref.69.1 ref.72.1 ref.72.2

In conclusion, the molecular mechanisms of mitosis are conserved to varying degrees across different organisms. The conserved mechanisms ensure accurate chromosome segregation and successful cell division. Ongoing research aims to further understand the molecular bases of the cell response to mitotic-targeting molecules, with a focus on gene expression profiles, the development of resistance, determinants of mitotic death induction, effects of DNA damage, response of cancer cell lines, mechanisms controlling apoptosis, detailed mechanisms of force generators, and control of mitosis in the presence of DNA damage.ref.88.2 ref.72.1 ref.72.1 This research is crucial for advancing cancer treatments and improving therapeutic strategies.ref.88.2 ref.28.63 ref.88.2

What are the key similarities and differences in mitotic mechanisms between model organisms and humans?

The Use of Model Organisms and Experimental Techniques in Understanding Mitotic Mechanisms

The study of mitotic mechanisms has greatly benefited from the use of specific model organisms such as C. elegans, Drosophila, Xenopus laevis, and yeast. These organisms have been extensively studied to investigate the molecular mechanisms and dynamics of mitosis.ref.28.63 ref.91.3 ref.91.2 By utilizing these model organisms, researchers have been able to gain valuable insights into the processes involved in chromosome segregation and spindle self-organization. Additionally, in vitro biophysical approaches and live cell microscopy have been employed to study the interactions of mitotic motors with dynamic microtubules and understand how they contribute to spindle structure and function. The combination of these model organisms and experimental techniques has significantly advanced our understanding of mitotic mechanisms.ref.91.28 ref.91.3 ref.91.28

Conservation of Mitotic Mechanisms between Model Organisms and Humans

Several specific mitotic mechanisms have been found to be conserved between model organisms and humans. One example is the DNA damage checkpoint, which serves to detect and signal the presence of DNA damage to the repair machinery. This mechanism has been studied in various model organisms, and its conservation suggests its importance in ensuring accurate chromosome segregation and cell division in humans as well.ref.14.2 ref.72.1 ref.24.2 Another conserved mechanism is the regulation of cell cycle progression through the activation and deactivation cycles of cyclin-dependent kinases (Cdks) and the synthesis and degradation of regulatory cyclins. This mechanism has also been extensively studied in different model organisms and is crucial for proper cell division in humans.ref.2.4 ref.7.2 ref.7.2

Furthermore, the regulation of anaphase B, which involves midzone pushing and braking, cortical pulling, and poleward flux, is conserved across different cell types. This mechanism has been investigated in various model organisms, including yeast, worms, and flies, and its conservation indicates its significance in ensuring accurate chromosome segregation and cell division in humans as well. The conservation of these mitotic mechanisms between model organisms and humans contributes to our understanding of human biology by providing insights into the fundamental processes involved in cell division and chromosome segregation.ref.28.63 ref.28.7 ref.28.49

Insights into Human Biology from the Conservation of Mitotic Mechanisms

The conservation of mitotic mechanisms across different species provides valuable insights into the fundamental processes that underlie cell division and chromosome segregation, which are essential for understanding human biology. The diverse solutions that eukaryotic cells have developed to solve the challenges of chromosome segregation are highlighted in the study of mitotic mechanisms. These mechanisms involve the differential deployment of conserved biochemical modules in different cell types, such as midzone pushing and braking, cortical pulling, and microtubule dynamics.ref.28.63 ref.86.1 ref.28.63 Understanding the molecular mechanisms of these processes at a high temporal resolution is crucial for unraveling the complexities of mitosis and cell division.ref.28.63 ref.28.63 ref.7.2

In vitro reconstitution of these processes using purified components allows for the investigation of their underlying molecular mechanisms. By studying the regulation of anaphase B, a key aspect of mitosis, researchers can gain a deeper understanding of the complex and dynamic nature of this process. Mathematical modeling is essential for comprehending the intricate interplay between various factors involved in anaphase B.ref.28.63 ref.28.63 ref.28.56 Additionally, the evolution of these machineries and mechanisms from the purely physical mechanisms that operated in dividing ancestral protocells on the early Earth is a fascinating unknown that holds the potential to shed light on the origins of mitosis and its significance in the evolution of life.ref.28.63 ref.28.63 ref.28.48

Overall, studying the conservation of mitotic mechanisms across different species provides valuable insights into the fundamental processes that underlie cell division and chromosome segregation, which are essential for understanding human biology.ref.28.63 ref.28.63 ref.7.2

Approaches to Study Mitotic Mechanisms and their Implications for Human Health and Disease

To study mitotic mechanisms in model organisms and gain insights into human health and disease, researchers employ various approaches. One of these approaches is the investigation of the mechanisms controlling the induction of apoptosis during mitosis. Understanding how chromosome instability is generated in cancer and how cells respond to anti-cancer chemotherapeutics can be achieved through the study of apoptosis during mitosis.ref.68.2 ref.68.3 ref.68.3 Another approach involves studying the timing of exit from mitosis, which is determined by the mitotic or spindle assembly checkpoint. This checkpoint is regulated by the mitotic checkpoint complex (MCC) and the anaphase-promoting complex or cyclosome (APC/C), which are involved in the ubiquitination and destruction of key substrates. The role of cyclin B1, a key substrate of APC/C, in regulating mitosis and apoptosis is also a subject of investigation.ref.59.2 ref.68.3 ref.68.2

Researchers also focus on understanding the evolution and divergence of spindle subsystems and the mechanochemical mechanisms of individual motors involved in chromosome segregation. In particular, they study the individual mechanisms of mitotic kinesins and cytoplasmic dynein, which play crucial roles in bipolar spindle assembly and chromosome segregation in animal cells. The structure and domain organization of mitotic kinesins, which consist of head domains and tails, are also areas of study.ref.91.3 ref.91.1 ref.91.3

Moreover, researchers explore the therapeutic potential of targeting mitosis in cancer treatment. They investigate the expression profile of genes encoding microtubule-associated and microtubule-regulatory proteins in different cancer types and within cells from the same cancer. Additionally, the mechanisms of resistance to microtubule-targeting agents, which can prevent microtubule polymerization or block their dynamic functions, are studied for their effects on mitotic cell death.ref.88.2 ref.88.1 ref.88.27

Furthermore, researchers study the control of mitosis in the event of DNA damage and the crosstalk between the DNA damage response (DDR), the spindle assembly checkpoint (SAC), and cell fate control. The existence of a mitotic DNA damage checkpoint and the proteins involved in mitotic arrest following DNA damage are subjects of investigation. The Xenopus egg extract system is utilized to study cell cycle progression and DNA damage checkpoints, providing insights into the ATM and ATR dependent checkpoint affecting centrosome-driven spindle assembly.ref.72.2 ref.72.2 ref.24.1

In conclusion, the study of mitotic mechanisms in model organisms and the application of various approaches has led to a deeper understanding of the molecular processes underlying mitosis. This understanding has important implications for human health and disease, as it provides insights into the mechanisms controlling cell division and chromosome segregation. The use of model organisms, experimental techniques, and the conservation of mitotic mechanisms across species all contribute to our understanding of the fundamental processes that underlie cell division and chromosome segregation, which are essential for understanding human biology.ref.28.63 ref.87.57 ref.28.63 Additionally, the exploration of mitotic mechanisms in the context of human health and disease opens up new avenues for therapeutic interventions and the development of novel cancer treatments.ref.28.63 ref.28.63 ref.28.63

How have these mechanisms evolved to ensure fidelity in chromosome segregation?

Mechanisms of Chromosome Segregation during Mitosis

The accurate segregation of chromosomes during mitosis is essential for the maintenance of genomic stability and the proper distribution of genetic material to daughter cells. This process involves a series of coordinated events and interactions between various molecular players. The main mechanisms involved in chromosome segregation during mitosis include homologous pairing and synapsis, recombination, cohesion, kinetochore-microtubule attachment, and spindle organization.ref.87.0 ref.87.1 ref.87.2

Homologous pairing and synapsis are crucial steps in meiosis I that ensure the correct segregation of homologous chromosomes. These processes involve the alignment and physical connection of homologous chromosomes, forming structures called bivalents. The formation of bivalents allows for the proper segregation of homologous chromosomes during the first meiotic division.ref.87.2 ref.87.7 ref.87.1 The molecular mechanisms underlying homologous pairing and synapsis have been extensively studied in model organisms such as Arabidopsis, maize, and rice. These studies have identified key proteins involved in these processes, such as the ZMM proteins in Arabidopsis, which are essential for the stabilization of chromosome pairing and synapsis.ref.87.5 ref.87.7 ref.87.4

Recombination, also known as crossing over, is another important event during meiosis that promotes genetic diversity and ensures proper chromosome segregation. Recombination involves the exchange of genetic material between homologous chromosomes, resulting in the formation of chiasmata. These chiasmata help hold homologous chromosomes together and facilitate their proper segregation during meiosis I.ref.87.8 ref.87.2 ref.87.1 The formation and resolution of chiasmata are tightly regulated processes involving several proteins, including the meiotic recombination proteins like MutS, MutL, and RPA.ref.87.8 ref.87.0 ref.87.2

Cohesion, mediated by the cohesin complex, plays a critical role in holding sister chromatids together until anaphase onset. The cohesin complex is composed of multiple subunits, including the Smc1, Smc3, and Rad21 proteins. These subunits form a ring-like structure that encircles sister chromatids, ensuring their proper alignment and segregation.ref.87.13 ref.30.2 ref.3.8 The regulation of cohesin dynamics during mitosis is tightly controlled by various factors, including the phosphorylation of cohesin subunits by protein kinases such as Polo-like kinase 1 (Plk1) and Aurora B kinase. The timely release of cohesin from chromosomes is essential for the separation of sister chromatids during anaphase.ref.3.8 ref.30.2 ref.87.13

Kinetochore proteins and microtubules are responsible for establishing proper spindle attachment and bi-orientation of sister kinetochores, ensuring the correct segregation of chromosomes in meiosis II. The kinetochore is a multi-protein complex that assembles on the centromere region of each sister chromatid. It serves as the attachment site for microtubules emanating from the spindle poles.ref.92.4 ref.64.3 ref.87.39 The dynamic interactions between kinetochore proteins and microtubules allow for the capture and movement of chromosomes during mitosis. The bi-orientation of sister kinetochores ensures that each chromatid is attached to microtubules from opposite spindle poles, promoting their equal segregation.ref.65.3 ref.87.39 ref.86.7

The regulation of these processes and the interaction between the proteins involved are areas of ongoing research. Various protein kinases, phosphatases, and ubiquitin ligases play crucial roles in modulating the activities of the proteins involved in chromosome segregation. For example, the Aurora B kinase is known to phosphorylate kinetochore proteins, contributing to the correction of improper microtubule attachments and the establishment of bi-orientation.ref.76.1 ref.65.41 ref.76.1 Additionally, spindle assembly checkpoint mechanisms monitor the proper attachment of kinetochores to microtubules, delaying anaphase onset until all chromosomes are correctly aligned and attached.ref.86.8 ref.65.41 ref.30.2

Chromosome Segregation during Plant Meiosis

The accurate segregation of chromosomes during meiosis is particularly important in plants, as it is crucial for the formation of haploid gametes and the maintenance of genetic diversity. Failure in proper chromosome segregation can lead to the formation of imbalanced gametes and aneuploid or polyploid progeny. The processes involved in accurate chromosome segregation during plant meiosis include synapsis, recombination, and the coordination of cell-cycle-dependent release of cohesin and attachment of sister kinetochores to microtubules.ref.87.1 ref.87.0 ref.87.1

Synapsis, the alignment and connection of homologous chromosomes, is a critical step in meiosis I. In plants, synapsis is mediated by the formation of the synaptonemal complex (SC), a proteinaceous structure that holds homologous chromosomes together. The SC is composed of two lateral elements and a central element, which are connected by transverse filaments.ref.87.7 ref.87.2 ref.87.18 The assembly of the SC is tightly regulated and involves the action of various proteins, including the ZYP1 protein in Arabidopsis. ZYP1 is essential for the stabilization and formation of the SC, ensuring the proper alignment and segregation of homologous chromosomes.ref.87.18 ref.87.17 ref.87.7

Recombination, similar to mitosis, is essential for the proper segregation of chromosomes during plant meiosis. It promotes genetic diversity by facilitating the exchange of genetic material between homologous chromosomes. The molecular mechanisms underlying recombination in plants are similar to those in other organisms and involve the actions of various proteins, including the MutS, MutL, and RPA proteins.ref.87.1 ref.87.3 ref.87.8 These proteins play crucial roles in the recognition and repair of DNA double-strand breaks, which are intermediates of recombination.ref.87.8 ref.87.5 ref.87.8

The coordination of cohesin release and kinetochore-microtubule attachment is essential for the accurate segregation of chromosomes during plant meiosis. The timely release of cohesin from chromosomes is required for the separation of sister chromatids during anaphase. In plants, the regulation of cohesin release is mediated by the protein kinase CDKB1 and the anaphase-promoting complex/cyclosome (APC/C).ref.30.2 ref.3.8 ref.25.2 CDKB1 phosphorylates cohesin subunits, promoting their dissociation from chromosomes. The APC/C, on the other hand, targets specific cohesin subunits for proteasomal degradation, leading to the release of cohesin from chromosomes.ref.3.8 ref.58.2 ref.58.1

Checkpoint mechanisms acting at kinetochores also control the accuracy of kinetochore-microtubule attachment during plant meiosis. These mechanisms monitor the proper attachment of kinetochores to microtubules and delay anaphase onset until all chromosomes are correctly aligned and attached. The spindle assembly checkpoint (SAC) is a conserved surveillance mechanism that ensures the fidelity of chromosome segregation.ref.87.3 ref.63.2 ref.86.8 The SAC is activated when kinetochores are not properly attached to microtubules, leading to the recruitment and activation of proteins that inhibit the activity of the APC/C, thus delaying anaphase onset.ref.87.42 ref.22.0 ref.63.2

The molecular factors involved in these processes have been identified and characterized through studies in model plants such as Arabidopsis, maize, and rice. However, more research is needed to understand the regulation of these factors in accordance with the cell cycle and their role in the broader context of chromosome segregation. The precise coordination of these processes is crucial for the accurate segregation of chromosomes and the maintenance of genomic stability during plant meiosis.ref.87.3 ref.87.57 ref.87.0

Conservation and Variations in Mitotic Mechanisms

The document excerpts do not explicitly state whether the mechanisms of mitosis are conserved across different species. However, they do mention that the eukaryota have developed a wide variety of approaches to chromosome segregation, some of which work differently than the mainstream model systems studied. This suggests that while there may be some conservation in mitotic mechanisms across different species, there are also variations and complexities that need to be further explored.ref.87.3 ref.89.5 ref.28.63

Indeed, there are known variations and differences in mitotic mechanisms among different organisms and cell types. The document mentions that there is diversity in mitotic mechanisms even within the same organism, such as differences in rates of spindle elongation in cultured S2 cells. This indicates that mitotic mechanisms can vary depending on the specific cell type and environmental conditions.ref.28.63 ref.28.54 ref.28.63

Furthermore, there are evolutionary differences between yeast and human mitoses, suggesting variations in mitotic mechanisms. Yeast cells undergo an open mitosis, where the nuclear envelope remains intact throughout the process, while human cells undergo a closed mitosis, where the nuclear envelope breaks down during prometaphase. These differences in mitotic mechanisms likely reflect the specific requirements and adaptations of different organisms.

The document also mentions that there is great diversity in mitotic mechanisms among vertebrate cells, with different mechanisms observed in different cell lines. For example, the mitotic mechanisms in epithelial cells may differ from those in neuronal cells. These variations may be related to the specific functions and characteristics of different cell types.ref.28.63 ref.28.63 ref.28.63

Furthermore, the document discusses the conserved role of certain proteins in meiotic chromosome segregation across different kingdoms, indicating variations in meiotic mechanisms. For example, the cohesin complex and kinetochore proteins play similar roles in meiosis across different organisms, but their regulation and interactions may vary.ref.87.4 ref.87.57 ref.87.3

In conclusion, while there may be some conservation in mitotic mechanisms across different species, there are also variations and complexities that need to be further explored. The ongoing research in this field aims to understand the molecular mechanisms regulating protein functions and the interaction between proteins to define their role in the broader context of chromosome segregation. By studying a wide range of organisms and cell types, researchers can gain a comprehensive understanding of the diversity and commonalities in mitotic mechanisms, contributing to our overall knowledge of cellular processes and genomic stability.ref.87.57 ref.87.3 ref.28.63

How can the study of mitotic mechanisms in model organisms contribute to our understanding of mitosis in humans?

Introduction

The study of mitotic mechanisms in model organisms has provided valuable insights into the process of mitosis in humans. By examining the diverse approaches to chromosome segregation and spindle dynamics in model organisms such as Xenopus laevis, Drosophila melanogaster, Caenorhabditis elegans, Schizosaccharomyces pombe, Saccharomyces cerevisiae, and mammalian cells, scientists have been able to uncover the mechanochemical mechanisms of mitotic motors and the coordination of various mitotic events. This knowledge is crucial for understanding the regulation of mitosis and developing potential therapeutic strategies for diseases related to mitotic dysfunction.ref.91.3 ref.91.2 ref.28.63

Insights from studying mitotic kinesins in model organisms

Understanding the mechanochemical mechanisms of mitotic kinesins

One of the specific insights gained from studying the modular structure and domain organization of mitotic kinesins in model organisms is a deeper understanding of their mechanochemical mechanisms. Mitotic kinesins interact with dynamic microtubules and play a crucial role in driving spindle assembly and chromosome segregation. By studying these mechanisms, researchers have identified the different roles of kinesins in spindle assembly and chromosome segregation.ref.91.1 ref.91.3 ref.87.48 These roles include walking directionally along microtubules, anchoring and crosslinking microtubules, and modulating microtubule dynamics.ref.87.48 ref.91.3 ref.28.51

Functional antagonism between oppositely-directed mitotic kinesins

Another insight gained from studying mitotic kinesins is the functional antagonism between pairs of oppositely-directed kinesins. Deleting one motor activity affects spindle function, while the deletion of an oppositely-directed motor rescues function. This finding highlights the importance of a delicate balance between the activities of different mitotic kinesins for proper spindle function.ref.91.1 ref.91.2 ref.91.3

Force-balance and force-integration mechanisms

The understanding of mitotic kinesins in model organisms has also shed light on the force-balance and force-integration mechanisms in the spindle. Researchers have investigated how mitotic kinesins generate and balance forces within the spindle, contributing to the overall stability and functionality of the mitotic machinery.ref.91.3 ref.91.2 ref.91.1

Potential of mitotic kinesins as targets for anti-cancer therapeutics

The insights gained from studying mitotic kinesins in model organisms have also led to the exploration of their potential as targets for anti-cancer therapeutics. By understanding the detailed mechanisms of these molecular motors, researchers can identify specific vulnerabilities in cancer cells and develop strategies to selectively target and disrupt mitosis in diseased cells.ref.91.2 ref.91.29 ref.91.29

Understanding motorized self-organization in eukaryotic life

Studying the detailed mechanisms of cytoskeletal molecular motors, including kinesins, in model organisms has provided a deeper understanding of motorized self-organization in eukaryotic life. By dissecting the positions and motions of kinesin molecules and understanding their interactions with different mitotic motors, researchers can gain insights into the collective behavior of these motors and their role in cellular processes.ref.91.2 ref.91.3 ref.91.1

Variety and diversity of mitotic mechanisms

Model organisms have also been instrumental in studying the variety and diversity of mitotic mechanisms in different cells and organisms. By examining these mechanisms, researchers can better understand how they originated and evolved, providing valuable evolutionary insights.

Biophysical approaches and live cell microscopy

In vitro biophysical approaches and live cell microscopy have been used to measure local forces, dissect the positions and motions of kinesin molecules, and understand the interactions between different mitotic motors. These techniques have allowed researchers to gain a more detailed understanding of the mechanisms underlying mitosis in model organisms.ref.91.28 ref.91.28 ref.91.3

Development of explicit models

The insights gained from studying mitotic kinesins in model organisms have also facilitated the development of explicit models that predict collective behavior based on the characteristic mechanical behavior of each mitotic kinesin. These models can help researchers better understand the complex dynamics of mitosis and provide a framework for further investigation.ref.91.28 ref.91.3 ref.91.1

Advanced imaging techniques

Advanced imaging techniques such as cryo-EM tomography, next-generation sequencing, and live-cell microscopy have also been utilized to improve spatial resolution, identify membrane-based kinetochores, and provide a dynamic description of mitotic mechanisms in non-model organisms. These techniques have allowed researchers to visualize and analyze mitotic processes at unprecedented levels of detail.ref.86.27 ref.86.27 ref.86.27

Application of mitotic mechanism knowledge in therapeutic strategies

The understanding of mitotic mechanisms in model organisms has significant implications for the development of therapeutic strategies for diseases associated with mitotic dysfunction.

Based on the knowledge of mitotic mechanisms, the design of mitosis-targeting molecules can be rationalized. This can lead to the development of drugs that specifically target and disrupt the mitotic process in diseased cells, offering potential therapeutic options for diseases such as cancer.ref.88.2 ref.88.1 ref.66.4

Understanding the molecular bases of the cell response to mitosis-targeting molecules is crucial for the development of more effective therapeutic strategies. By studying how cells respond to these molecules, researchers can identify potential vulnerabilities or resistance mechanisms that can be targeted for therapeutic intervention.ref.88.2 ref.88.27 ref.88.1

The identification of genes encoding microtubule-associated and microtubule-regulatory proteins in different cancer types can inform the selection of appropriate therapeutic agents. The expression profile of these genes can determine the sensitivity or resistance of tumor cells to microtubule-targeting agents, enabling the development of personalized treatment approaches.ref.88.2 ref.88.2 ref.88.1

The identification of determinants of mitotic death induction is crucial for the development of strategies to enhance the propensity of cells to undergo mitotic cell death. By understanding the factors that influence the response of cells to microtubule-targeting agents, researchers can develop approaches to increase the effectiveness of these agents in inducing cell death in diseased cells.ref.88.1 ref.88.2 ref.88.26

Sliding filament model

The sliding filament model is a hypothesis that proposes that mitotic motors slide apart adjacent microtubules to drive the movements of the mitotic spindle during chromosome segregation. This model has been extensively studied and tested through electron microscopy and biochemical studies.ref.28.6 ref.28.4 ref.28.5

The sliding filament model can explain pole-pole separation during anaphase B spindle elongation. Electron microscopy studies have shown the organization of mitotic spindle microtubules, supporting the idea that sliding filaments contribute to spindle elongation.ref.28.5 ref.28.6 ref.28.56

However, the sliding filament model does not fully explain chromosome-to-pole movement during anaphase A. Further research is needed to understand the specific mechanisms involved in this process.ref.28.56 ref.28.6 ref.28.4

Kinesin motors, particularly kinesin-5 motors, have been identified as potential motors that can cross-link and slide apart antiparallel interpolar microtubules in the spindle midzone. These motors play a crucial role in midzonal MT-MT sliding and anaphase B spindle elongation.ref.28.36 ref.28.35 ref.47.14

Other work has shown that motors located on the cell cortex can also contribute to midzonal MT-MT sliding and anaphase B spindle elongation. Cortical pulling mechanisms involve the pulling apart of the spindle poles by motors located on the cell cortex, providing an alternative or complementary mechanism for spindle elongation.ref.28.7 ref.28.27 ref.28.52

The balance between midzone pushing and cortical pulling mechanisms contributes to the sliding filament mechanism of anaphase B spindle elongation. Outward pulling forces generated by cortical dynein, along with the activity of kinesin-5 motors on the midzone, drive the slow phase of anaphase B. Forces exerted at the cortex, likely by dynein and/or aMT depolymerization, pull the poles apart while these forces are resisted by antagonistic forces generated at the midzone.ref.28.7 ref.28.52 ref.28.6

Conclusion

The study of mitotic mechanisms in model organisms has provided valuable insights into the diverse approaches to chromosome segregation and spindle dynamics. Through the study of mitotic kinesins, researchers have gained a deeper understanding of the mechanochemical mechanisms involved in mitosis, the balance between different mitotic motors, and the variety and diversity of mitotic mechanisms in different cells and organisms. This knowledge has important implications for the development of therapeutic strategies for diseases associated with mitotic dysfunction.ref.91.1 ref.91.3 ref.91.3 Furthermore, the sliding filament model has provided a framework for understanding the movements of the mitotic spindle, although further research is needed to fully elucidate the specific mechanisms involved. Overall, the study of mitotic mechanisms in model organisms has significantly advanced our understanding of mitosis and has the potential to lead to the development of novel therapeutic approaches.ref.28.6 ref.28.4 ref.28.63

What are the implications of evolutionary conservation for therapeutic interventions targeting mitosis?

Implications of Evolutionary Conservation for Therapeutic Interventions Targeting Mitosis

The implications of evolutionary conservation for therapeutic interventions targeting mitosis are significant. Mitosis remains a valuable therapeutic target, as it plays a crucial role in cell division and is essential for the growth and proliferation of cancer cells. Therefore, efforts to rationalize the design of mitosis-targeting molecules and understand the molecular bases of the cell response to these molecules are encouraged.ref.88.2 ref.88.2 ref.88.2

However, there are limitations to the therapeutic use of mitosis-targeting agents. One limitation is the expression profile of genes encoding microtubule-associated (MT-associated) and MT-regulatory proteins found in different cancer types. The expression profile of these genes can synergize with or antagonize the effect of the drugs, affecting their efficacy.ref.88.2 ref.88.1 ref.88.27 Additionally, tumor cells can develop resistance to mitosis-targeting agents through diverse mechanisms. For example, efflux pump mechanisms can be induced, leading to the expulsion of the drugs from the cancer cells. Furthermore, the presence of mutations or differential expression of tubulin isotypes can also contribute to drug resistance.ref.88.2 ref.88.26 ref.88.27

To overcome these limitations, there is a need to increase the molecular repertoire of potentially effective drugs. Tubulin-targeting agents, for example, can prevent MT polymerization or block their dynamic functions, leading to mitotic cell death. However, cells treated with these agents show variable propensity to undergo mitotic cell death.ref.88.2 ref.88.1 ref.88.26 Recent studies have sought to identify the determinants of mitotic death induction, including the expression profile of genes encoding MT-associated and MT-regulatory proteins, as well as the presence of mutations or differential expression of tubulin isotypes.ref.88.26 ref.88.2 ref.88.29

Furthermore, the timing of exit from mitosis is determined by the mitotic or spindle assembly checkpoint. The propensity of cells to undergo apoptosis in mitosis correlates with the duration of mitosis. This suggests that the duration of mitosis is a critical factor in determining the cell's fate.ref.68.3 ref.68.3 ref.72.17 Moreover, the progression of cells carrying DNA damage through mitosis and the subsequent DNA damage response (DDR) and repair in the G1 phase have been observed. There is evidence for the existence of a mitotic DNA damage checkpoint, and the repair mechanisms during mitosis include nonhomologous end joining (NHEJ), alternative end joining (Alt-EJ), and single-strand annealing (SSA) pathways. However, DNA repair in mitosis is largely suppressed, and inaccurate repair in mitosis can lead to the formation of chromatin bridges.ref.72.2 ref.72.6 ref.72.1

The control of mitosis in the event of DNA damage involves crosstalk between the DDR and the spindle assembly checkpoint. There is evidence for the existence of a mid-anaphase DNA damage checkpoint, which suggests that cell fate decisions are instigated in mitosis, even though they are not physically carried out until the G1 phase. The DNA repair proteins are involved in complexes that tether broken chromosomes, and cell fate is not fully determined until the next G1 phase.ref.72.17 ref.72.2 ref.72.1 Further research is needed to understand how DNA damage is sensed in mitosis and how it leads to a prolonged spindle assembly checkpoint.ref.72.1 ref.72.2 ref.72.18

Molecular Bases of the Cell Response to Mitosis-Targeting Molecules

The specific molecular bases of the cell response to mitosis-targeting molecules are multifaceted. The expression profile of genes encoding MT-associated and MT-regulatory proteins plays a crucial role in determining the response of cancer cells to these molecules. The presence of mutations or differential expression of tubulin isotypes can also influence the response.ref.88.2 ref.88.27 ref.88.1

The response of cancer cells to anti-mitotic drugs is influenced by the genetic background of the cancer cells and the expression profile of genes encoding MT-regulatory factors. Different drugs targeting tubulin display differential efficacy in living cells, indicating that molecules with similar tubulin polymerization inhibitory activity in vitro can have different effects in living cells. This highlights the complexity of the response and the need to consider the specific molecular context in which the drugs are acting.ref.88.27 ref.88.26 ref.88.1

The response of cancer cells to tubulin inhibitors involves the induction of cytological phenotypes that trigger MCL-1 down-regulation and caspase-3 activation, ultimately leading to cell death. The timing of exit from mitosis, as determined by the mitotic or spindle assembly checkpoint, also influences the response. The propensity of cells to undergo apoptosis during mitosis correlates with the duration of mitosis, suggesting that there is a progressive increase in pro-apoptotic signaling until it reaches a threshold sufficient to initiate cell death.ref.68.2 ref.68.3 ref.68.4 Whether a cell dies in mitosis depends on whether the apoptotic threshold is breached before exit from mitosis.ref.68.3 ref.68.4 ref.68.2

The response of mitotic cells to DNA damage or spindle poisoning drugs can be complex and heterogeneous, with individual cells displaying a variety of different fates. The severity of induced MT damage and the induction of sustained mitotic arrest are linked to the capacity of tubulin inhibitors to induce cell death. The severity of MT damage is also a predictor of mitotic cell death induction by tubulin inhibitory drugs.ref.88.26 ref.24.3 ref.88.1 The response of cancer cell lines to tubulin inhibitors can vary, with different inhibitors yielding distinct cell fate profiles. This highlights the need to consider the specific molecular context and genetic background of the cancer cells when designing therapeutic interventions.ref.88.2 ref.88.1 ref.88.27

Mechanisms of Resistance to Mitosis-Targeting Agents

Cancer cells can develop resistance to mitosis-targeting agents through diverse mechanisms. The expression profile of genes encoding MT-associated and MT-regulatory proteins is one of the mechanisms that can contribute to drug resistance. Additionally, the induction of efflux pump mechanisms implicated in multidrug resistance can lead to reduced efficacy of mitosis-targeting agents in cancer therapy.ref.88.2 ref.88.27 ref.88.2 The presence of mutations or differential expression of tubulin isotypes is another mechanism that can confer resistance.ref.88.2 ref.88.1 ref.88.26

Furthermore, the ability of cells to adapt to stressful conditions through checkpoint adaptation can contribute to drug resistance. Checkpoint adaptation allows cells to bypass the mitotic or spindle assembly checkpoint and exit mitosis despite the presence of DNA damage or drug-induced spindle defects. This adaptation can occur through the inactivation or down-regulation of essential checkpoint components, such as Mps1.ref.14.16 ref.14.16 ref.14.17 Inhibition of the mitotic checkpoint represents a potential anti-cancer strategy to overcome resistance.ref.79.0 ref.79.0 ref.79.0

Given the diverse mechanisms through which cancer cells can develop resistance, there is a need to explore alternative strategies for targeting the mitotic checkpoint in tumor cells. Increasing the molecular repertoire of potentially effective drugs and considering the specific molecular context in which the drugs are acting can help overcome resistance and improve the efficacy of mitosis-targeting agents in cancer therapy.ref.88.2 ref.88.1 ref.88.2

Positive Feedback Regulation and the Timing of Exit from Mitosis

The timing of exit from mitosis is a tightly regulated process controlled by the mitotic or spindle assembly checkpoint. Positive feedback in the Cdk1-cyclin B1 regulatory network plays a crucial role in the timing of exit from mitosis. This positive feedback ensures that the duration of mitosis is short and remarkably constant, regardless of variability in earlier cell-cycle phases.ref.7.37 ref.7.22 ref.7.19

The duration of mitosis is temporally insulated from variability in earlier cell-cycle phases through the regulation of positive feedback. Perturbing positive feedback results in a sluggish and variable entry and progression through mitosis, coupling the duration of mitosis to the duration of interphase or overall cell-cycle length. This highlights the interconnectedness of the timing of exit from mitosis, the duration of mitosis, and the propensity of cells to undergo apoptosis.ref.7.9 ref.7.1 ref.7.0

The propensity of cells to undergo apoptosis correlates with the duration of mitosis, indicating a progressive increase in pro-apoptotic signaling until it reaches a threshold sufficient to initiate cell death. This further emphasizes the importance of the timing of exit from mitosis and the duration of mitosis in determining the cell's fate. Positive feedback in the networks that regulate mitosis is the molecular mechanism that insulates the duration of mitosis from variability in earlier cell-cycle events.ref.68.3 ref.7.37 ref.68.4

In conclusion, the implications of evolutionary conservation for therapeutic interventions targeting mitosis are significant. Mitosis remains a valuable therapeutic target, but there are limitations to the therapeutic use of mitosis-targeting agents, such as the expression profile of genes encoding MT-associated and MT-regulatory proteins and the development of resistance through diverse mechanisms. The specific molecular bases of the cell response to mitosis-targeting molecules include the expression profile of these genes, the presence of mutations or differential expression of tubulin isotypes, and the differential pharmacokinetics properties of different mitosis-targeting molecules at the single cell level.ref.88.2 ref.88.1 ref.88.27 Furthermore, the response of cancer cells to these molecules is influenced by the genetic background of the cancer cells and the expression profile of genes encoding MT-regulatory factors. Understanding the determinants of mitotic death induction, as well as the mechanisms of resistance, can help improve the design of therapeutic interventions targeting mitosis. Additionally, the regulation of positive feedback in the cell-cycle regulatory network plays a crucial role in the timing of exit from mitosis, the duration of mitosis, and the propensity of cells to undergo apoptosis.ref.88.27 ref.88.2 ref.88.26 Overall, further research is needed to fully elucidate the molecular mechanisms underlying the cell response to mitosis-targeting agents and to develop effective strategies for targeting mitosis in cancer therapy.ref.88.2 ref.88.1 ref.88.26

Works Cited