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#anaphase

21 public questions tagged with this topic.

Cell Division

What is the function of the spindle assembly checkpoint?

Reliability of chromosome segregation depends on surveillance mechanism preventing anaphase until every chromosome correctly attached to spindle, because single missegregation yields aneuploid daughter prone to tumor development or cell death. Spindle assembly checkpoint monitors occupancy and tension at kinetochores, large protein structures assembled on CENP-A containing centromeric chromatin. Unattached kinetochores recruit Mad1-Mad2 complex that catalyzes conversion of cytosolic open Mad2 to closed form bound to Cdc20. Together with BubR1, Bub3, Mps1-phosphorylated Bub1 they assemble mitotic checkpoint complex MCC that diffuses to inhibit APC/C-Cdc20 ubiquitin ligase activity, blocking degradation of securin and cyclin B. Aurora B kinase at inner centromere phosphorylates Ndc80 complex under low tension destabilizing erroneous syntelic attachments, recreating unattached kinetochore that re-engages checkpoint. Upon biorientation where sister kinetochores attach to opposite poles generating tension, Mad1-Mad2 recruitment ceases, MCC disassembles, APC/C activates separase, and synchronous segregation proceeds, preventing errors. This circuitry is highly conserved across eukaryotes, integrating growth factor signals, DNA damage surveillance, and developmental cues, and its disruption frequently underlies oncogenesis, providing targets for checkpoint inhibitors and cancer therapeutics.

Ref: Musacchio & Salmon, Nature Reviews Mol Cell Biol 2007, SAC Mechanism. Alberts 7th ed., Chapter 17.

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Cell Division

The separation of sister chromatids occurs during:

Separation of sister chromatids underlies chromosome segregation during anaphase and relies on coordinated dissolution of cohesin and activation of motor mechanisms. Throughout S phase and G2, cohesin ring comprising Smc1, Smc3, and kleisin Rad21 embraces sister DNAs, establishing cohesion resisting spindle pulling until timely release. Shugoshin-PP2A protects centromeric cohesion specifically until anaphase onset. Upon satisfaction of spindle assembly checkpoint, APC/C-Cdc20 ubiquitinates securin and cyclin B with K11 chains for proteasomal destruction. Securin degradation frees separase endopeptidase that cleaves Rad21 at conserved EXXR sites, irreversibly opening ring along chromosome arms and centromeres. Loss of cohesion allows stored elastic energy release, kinetochore microtubules shorten through depolymerization at kinetochore and pole during anaphase A, while antiparallel interpolar microtubules elongate pushing poles apart during anaphase B involving Eg5 kinesin. Phosphatases reverse CDK1 phosphorylations facilitating chromatin remodeling after separation, ensuring aneuploidy avoidance. This circuitry is highly conserved across eukaryotes, integrating growth factor signals, DNA damage surveillance, and developmental cues, and its disruption frequently underlies oncogenesis, providing targets for checkpoint inhibitors and cancer therapeutics.

Ref: Alberts et al., Molecular Biology of the Cell, 7th ed., Chapter 17, Anaphase Mechanisms. Nasmyth, Annu Rev Genet 2009.

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Cell Cycle Regulation

Which protein is degraded by APC to trigger chromosome separation?

Chromatid cohesion must be maintained until synchronous separation at anaphase, requiring protection against premature cleavage. Securin, also called pituitary tumor transforming gene PTTG, functions as dual inhibitor and chaperone of separase. Securin sequence contains D-box and KEN motifs for APC/C recognition and binds separase HEAT repeats, occluding catalytic triad. Accumulation during S and G2 concentrates at centromeres. At metaphase, APC/C-Cdc20 polyubiquitinates securin, triggering rapid proteasomal degradation within five to ten minutes, concentration dropping precipitously. Simultaneously cyclin B destruction inactivates CDK1 that also phosphorylates separase inhibitory sites, fully unleashing protease. Liberated separase translocates to chromosomes and cleaves Rad21/Scc1 kleisin at conserved EXXR motifs after glutamate, generating N- and C-terminal fragments unable to maintain ring closure. This opens cohesin, allowing kinetochore microtubules to pull sisters to opposite poles. Expression of non-degradable securin mutant with mutated D-box prevents cohesion loss and arrests cells in metaphase with intact cohesion. This circuitry is highly conserved across eukaryotes, integrating growth factor signals, DNA damage surveillance, and developmental cues, and its disruption frequently underlies oncogenesis, providing targets for checkpoint inhibitors and cancer therapeutics.

Ref: Uhlmann et al., Nature 1999, Separase Cohesin Cleavage. Alberts 7th ed., Chapter 17, Securin.

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Cell Cycle Regulation

Which regulatory factor is crucial for the metaphase-to-anaphase transition?

Metaphase to anaphase transition represents irreversible commitment executed by APC/C bound to Cdc20 coactivator. Throughout prometaphase chromosomes attach via kinetochores, generating unattached kinetochore signal converting Mad2 to active inhibitor forming mitotic checkpoint complex with BubR1, Bub3, and Cdc20 that binds APC/C core to block E2 recruitment. Upon achievement of biorientation of all chromosomes and establishment of tension, checkpoint protein recruitment stops, complex disassembles, and free Cdc20 associates with phosphorylated APC/C previously primed by CDK1 and Plk1. Active APC/C-Cdc20 polyubiquitinates securin and cyclin B with K11 chains for proteasomal destruction. Securin normally binds and locks separase protease in inactive conformation together with cyclin B-CDK1 phosphorylation. Degradation liberates separase that cleaves kleisin Rad21 subunit of cohesin ring embracing sister chromatids since S phase, opening ring, while cyclin B destruction drops CDK1 activity permitting phosphatase activation and mitotic exit. Blocking securin degradation prevents chromatid separation. This circuitry is highly conserved across eukaryotes, integrating growth factor signals, DNA damage surveillance, and developmental cues, and its disruption frequently underlies oncogenesis, providing targets for checkpoint inhibitors and cancer therapeutics.

Ref: Peters et al., Cell 1998, APC/C Regulation. Musacchio, Nat Rev Mol Cell Biol 2011, Anaphase.

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Cell Cycle Regulation

What ensures the proper segregation of chromosomes at anaphase?

Correct chromatid partitioning during anaphase integrates mechanical forces and biochemical resetting pathways. Physical separation depends on earlier APC/C-Cdc20 mediated securin degradation freeing separase to cleave cohesin subunit Scc1 at centromeres, allowing kinetochore microtubules to pull sister chromatids poleward via depolymerization coupled movement. However exit from high CDK state also essential. In budding yeast model, mitotic exit network monitors spindle elongation, activating Tem1 GTPase when daughter pole enters bud overcoming checkpoint inhibition by Kin4-Bub2-Bfa1 GAP complex. Tem1 triggers Cdc15 and Dbf2-Mob1 kinases leading to release of Cdc14 phosphatase from nucleolus. Cdc14 dephosphorylates CDK substrates including Cdh1 activating APC/C-Cdh1 for cyclin B and securin clearance plus spindle disassembly factors Ase1 and Fin1, Sic1 inhibitor synthesis via Swi5, and central spindle stabilization. Rising PP2A-B55 activity in mammals dephosphorylates PRC1 and ECT2 essential for central spindle bundling and RhoA activation for cytokinesis. Thus accurate segregation requires both cohesin removal and Cdc14 dependent phosphatase activation promoting low CDK environment enabling decondensation and division completion. Additional feedback loops involving polo-like kinases, phosphatases and SCF-mediated degradation reinforce irreversibility and protect against premature progression that would compromise genome integrity and viability.

Ref: Amon, Mitotic Exit Network and Cdc14 in Segregation, Current Biology; Bardin & Amon, Coordination of Anaphase and Exit.

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Cell Cycle Regulation

The metaphase-to-anaphase transition is triggered by:

The metaphase to anaphase transition is irreversible decision point governed by ubiquitin dependent proteolysis. When every chromosome achieves amphitelic kinetochore-microtubule attachment generating inter-kinetochore tension, spindle assembly checkpoint signaling through MCC production ceases. Dynein-mediated stripping of Mad1-Mad2 from kinetochores and p31comet-TRIP13 catalyzed MCC disassembly liberate coactivator Cdc20, enabling APC/C-Cdc20 ligase activation. Active APC/C polyubiquitinates securin via destruction box motif and cyclin B via D-box for rapid degradation by 26S proteasome. Securin degradation frees separase cysteine protease previously held as pseudosubstrate, while cyclin B destruction reduces CDK1 activity permitting dephosphorylation of mitotic substrates by PP1 and PP2A-B55. Free separase cleaves kleisin subunit Scc1 or Rad21 of cohesin tetramer encircling sister chromatids at centromeres rescued from earlier Wapl-mediated arm removal by Shugoshin-PP2A protection. Opening of cohesin ring dissolves linkage, allowing poleward microtubule pulling forces to separate sisters. Chromatid movement manifests as anaphase A toward poles and anaphase B spindle elongation driven by Eg5 kinesin and cortical dynein. Additional feedback loops involving polo-like kinases, phosphatases and SCF-mediated degradation reinforce irreversibility and protect against premature progression that would compromise genome integrity and viability.

Ref: Peters, Anaphase Promoting Complex Orchestrating Metaphase-Anaphase Transition, Nat Rev Mol Cell Biol 2006; Alberts et al., Chapter 18, Anaphase.

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Cell Cycle Regulation

Which enzyme cleaves cohesin at anaphase onset?

Persistent cohesion between sister chromatids depends on cohesin, tetrameric ring of Smc1, Smc3 ATPases, kleisin Scc1/Rad21 and stromalin SA1/2 subunits embracing replicated DNA strands. During prophase most arm cohesin dissociates via Wapl-dependent opening after Plk1 and Aurora B phosphorylation of SA2, while centromeric pool protected by Shugoshin recruiting PP2A-B56 counteracting phosphorylation remains for biorientation. Final dissolution at anaphase onset requires proteolytic cleavage. Separase, 230 kDa cysteine protease of CD clan, executes this. Kept inactive by dual mechanisms including pseudosubstrate binding of securin chaperone covering conserved active site and phosphorylation by CDK1-Cyclin B1 promoting cyclin B interaction, separase activates only after APC/C-Cdc20 polyubiquitinates securin and cyclin B for proteasomal destruction. Freed separase recognizes conserved separase cleavage motif Glu-X-X-Arg in Scc1 and cleaves after Arg arginine 172 and 450 in human Rad21, opening ring structure and allowing spontaneous dissociation. Cleaved cohesin fragments removed, permitting microtubule pulling forces to drive disjunction. Separase also cleaves itself, cohesin-independent roles in FEAR activation and centriole disengagement. Additional feedback loops involving polo-like kinases, phosphatases and SCF-mediated degradation reinforce irreversibility and protect against premature progression that would compromise genome integrity and viability.

Ref: Nasmyth & Haering, Cohesin and Separase Regulation, Annu Rev Genet 2009; Uhlmann, Separase in Anaphase and Beyond.

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Cell Cycle Regulation

During mitosis, which event is triggered by APC/C activation?

Transition into anaphase is driven by activation of anaphase promoting complex cyclosome bound to Cdc20, a large 1.2 MDa cullin-RING E3 ubiquitin ligase. When SAC is satisfied, MCC disassembly through p31comet and TRIP13 releases Cdc20 permitting association with APC/C core. Active APC/C-Cdc20 recognizes destruction box degrons on two pivotal substrates: securin and Cyclin B. Polyubiquitination of securin triggers rapid proteolysis via 26S proteasome, discharging separase cysteine protease previously held inactive as pseudosubstrate. Free separase cleaves Scc1 or Rad21 kleisin subunit of cohesin rings at conserved E-x-x-R- site, opening tripartite ring and dissolving sister chromatid cohesion at centromeres remaining after Wapl-mediated arm removal. Simultaneously, cyclin B ubiquitination and degradation reduces CDK1 activity, permitting phosphatases PP1 and PP2A-B55 to dephosphorylate mitotic substrates, soften cortex and allow anaphase A chromatid-to-pole movement and anaphase B spindle elongation driven by Eg5 kinesin and cortical dynein. APC/C activation therefore couples cohesion destruction to mitotic exit preparation ensuring ordered segregation and genome stability. Additional feedback loops involving polo-like kinases, phosphatases and SCF-mediated degradation reinforce irreversibility and protect against premature progression that would compromise genome integrity and viability.

Ref: Pines, APC/C Regulation of Anaphase Onset, Nat Rev Mol Cell Biol 2011; NCBI Bookshelf, Anaphase Triggered by APC/C.

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Cell Cycle Regulation

Which factor ensures proper chromosome segregation at anaphase?

Sister chromatid cohesion, mediated by the ring-shaped cohesin complex composed of Smc1, Smc3, kleisin Scc1 or Rad21 and SA proteins, must be abruptly destroyed only after bioriented attachment. Separase provides the decisive enzymatic activity. This large cysteine endopeptidase recognizes a conserved E-x-x-R motif within the kleisin subunit and cleaves after arginine, effectively opening the ring. During early mitosis separase remains constrained by two inhibitory locks: stoichiometric binding of securin which occupies the catalytic groove as pseudosubstrate and chaperone, and CDK1-Cyclin B1 dependent phosphorylation that promotes cyclin B association. When the spindle assembly checkpoint is satisfied, free APC/C-Cdc20 polyubiquitinates securin and cyclin B through D-box and KEN-box degrons for rapid proteasomal degradation. Liberated separase becomes active, cleaves centromeric cohesin that escaped Wapl-mediated removal from chromosome arms earlier in prophase. Cleavage allows kinetochore microtubules to generate pulling forces unopposed, driving chromatids toward opposite poles. Failure to regulate separase causes premature separation, aneuploidy and tumorigenesis, underscoring tight cell cycle control by APC/C and securin balance. Additional feedback loops involving polo-like kinases, phosphatases and SCF-mediated degradation reinforce irreversibility and protect against premature progression that would compromise genome integrity and viability.

Ref: Alberts et al., Molecular Biology of the Cell, 6th ed., Chapter 17: Cell Cycle, Sister-Chromatid Separation and Separase Regulation.

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