Cell Cycle Regulation

Cell Cycle Regulation

Which kinase is involved in regulating the initiation of DNA replication in S phase?

Initiation of DNA replication at licensed origins depends on two kinase families that separate licensing from firing. DDK kinase complex Dbf4-Cdc7 phosphorylates MCM2-7 helicase at N-terminal serine-threonine clusters, promoting Cdc45 recruitment. Second trigger is S-phase CDK activity supplied by cyclin E-CDK2 that peaks at G1/S border and cyclin A-CDK2 that sustains activity into S phase. Cyclin E-CDK2 phosphorylates Treslin/TICRR at threonine 969, MTBP, RecQL4, and orthologs of yeast Sld2 and Sld3, enabling their binding to BRCT repeats of TopBP1 and assembly of Cdc45-MCM-GINS active helica

Ref: Labib, Genes & Dev 2010, Origin Activation. Limas & Cook, Genes & Dev 2019, CDK Roles S Phase.

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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. Simultan

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

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Cell Cycle Regulation

Wee1 kinase inhibition in G2 phase leads to:

G2 progression is gated by inhibitory phosphorylation of master mitotic kinase CDK1 that prevents premature mitotic entry while last preparations complete. Nuclear kinase Wee1 and membrane-associated Myt1 phosphorylate CDK1 subunit within cyclin B-CDK1 complex at Tyr15 and Thr14 within ATP-binding loop, rendering kinase catalytically inactive despite abundant cyclin B synthesized during G2, allowing centrosome maturation, Golgi segregation, and DNA repair synthesis to proceed. At G2/M border, dual-specificity phosphatases Cdc25A/B/C remove these phosphates, causing explosive autoamplification

Ref: Harper & Elledge, Mol Biol Cell 2007, Wee1 Control. Alberts 7th ed., Chapter 17, CDK1 Regulation.

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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

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 is the function of Cdc6 and Cdt1 in DNA replication?

Pre-replication complex formation establishes replication competence before S phase, separating helicase loading from activation. Origin recognition complex ORC marks replication origins throughout cell cycle at AT-rich sequences. In late mitosis and early G1 when CDK activity minimal due to APC/C-Cdh1 mediated cyclin degradation, AAA+ ATPase Cdc6 binds ORC and recruits chaperone Cdt1 bound to single hexamer of MCM2-7 helicase. ATP hydrolysis by Cdc6 and ORC opens MCM ring, allowing encirclement of double-stranded DNA and formation of inactive double hexamer encircling DNA in head-to-head orie

Ref: Bell & Labib, Genes & Dev 2016, Pre-RC Assembly. Alberts 7th ed., Chapter 5, Replication Licensing.

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Cell Cycle Regulation

In the regulation of DNA replication, which factor ensures that origins fire only once per cycle?

Eukaryotes must replicate each DNA segment exactly once per cell cycle to avoid re-replication which causes double-strand breaks and genomic instability. Origin licensing is restricted to G1 when CDK activity low: ORC, Cdc6, and Cdt1 load double hexamer of MCM2-7 helicase. At S onset, CDK and DDK activate helicase, but re-loading must be blocked. Geminin protein accumulating from S through early mitosis directly binds Cdt1 through extended coiled-coil domain, sterically occluding MCM-interacting surface and preventing second helicase loading. Concurrently, cyclin A-CDK2 phosphorylates Cdt1 and

Ref: McGarry & Kirschner, Cell 1998, Geminin Function. Fragkos et al., Nature Rev Mol Cell Biol 2015.

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Cell Cycle Regulation

Which molecule directly targets p27 for ubiquitin-mediated degradation?

Abundance of p27 is controlled post-translationally rather than transcriptionally, illustrating CDK-regulated proteolysis that sharpens G1/S transition. Late in G1, cyclin E-CDK2 phosphorylates p27 intrinsically disordered C-terminus at threonine 187. This phosphate forms high-affinity interaction with WD40 domain of Skp2, F-box protein assembled into SCF-Skp2 E3 ligase with Skp1 adaptor, Cullin1 scaffold, Rbx1 RING finger recruiting Ubc3 E2 enzyme, plus Cks1 cofactor that bridges CDK and F-box. SCF catalyzes formation of K48-linked polyubiquitin chains on p27 lysines, marking it for rapid deg

Ref: Carrano et al., Nature Cell Biology 1999, SCF-Skp2 Targets p27. Alberts 7th ed., Chapter 3.

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Cell Cycle Regulation

The role of p27 in G1 phase is to:

p27Kip1 product of CDKN1B gene exemplifies Cip/Kip family that imposes G1 arrest and integrates nutrient availability with size control. In quiescent and early G1 cells, p27 accumulates due to low proteasomal turnover and binds cyclin E-CDK2 heterodimer, inserting 3_10 helix into ATP-binding pocket, preventing catalysis and blocking Rb phosphorylation, maintaining E2F repression. This prevents premature S-phase entry when growth factors limited. Mitogen signaling induces cyclin D-CDK4/6 complexes that sequester p27 in trimeric assembly without fully inhibiting them, lowering free p27 pool. Sub

Ref: Sherr & Roberts, Genes & Dev 2004, p27 Biology. Morgan, Cell Cycle, Chapter 4, G1 Control.

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Cell Cycle Regulation

What is the role of the Spindle Assembly Checkpoint (SAC)?

Chromosome missegregation produces aneuploidy implicated in tumorigenesis, congenital disorders, and embryonic lethality, necessitating surveillance by spindle assembly checkpoint. Sensor is kinetochore assembled on CENP-A containing centromeric nucleosomes that binds microtubule plus ends via Ndc80 complex. Unattached kinetochores act as catalytic platforms where Mad1-Mad2 heterotetramer converts soluble open Mad2 into closed conformation that binds Cdc20. Together with BubR1, Bub3, and Bub1 phosphorylated by Mps1 kinase, they generate diffusible mitotic checkpoint complex that inhibits APC/C

Ref: Musacchio, Current Biology 2015, SAC Mechanism. Alberts 7th ed., Chapter 17, Mitosis Quality.

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Cell Cycle Regulation

The protein p16 (INK4) primarily inhibits:

INK4 family represents dedicated inhibitors of CDK4 and CDK6 distinguishing them from Cip/Kip family that target cyclin E-CDK2 and cyclin A-CDK2. p16INK4a product of CDKN2A locus contains four ankyrin repeats forming elongated structure that binds CDK6 opposite cyclin-binding interface, inducing distortion of ATP-binding pocket and preventing association with cyclin D. Since cyclin D-CDK4/6 is earliest kinase phosphorylating Rb in response to mitogens, its inhibition maintains Rb in hypophosphorylated active repressor state bound to E2F, arresting cell before restriction point. Genetic inactiv

Ref: Sherr & Roberts, Genes & Dev 1999, INK4 Family Biology. Alberts 7th ed., Chapter 20.

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Cell Cycle Regulation

Which kinase activates APC/C-Cdc20 to trigger the metaphase-anaphase transition?

Assembly and activation of APC/C-Cdc20 must occur only after CDK1 has driven mitotic entry, ensuring anaphase does not initiate prematurely. During G2, APC/C exists in unphosphorylated inactive conformation with weak affinity for Cdc20, while Cdc20 itself is kept inactive by CDK inhibitory phosphorylation and binding to checkpoint protein Mad2. Upon entry into mitosis, cyclin B-CDK1, the master mitotic kinase, phosphorylates multiple subunits including APC1 at loop region, APC3, and APC6, inducing conformational rearrangement exposing C-box binding groove. Polo-like kinase Plk1 adds further ph

Ref: Lodish et al., Molecular Cell Biology, 9th ed., Chapter 20: Mitotic Activation of APC/C.

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Cell Cycle Regulation

The APC/C-Cdh1 complex is required for:

Specificity and timing of APC/C activity arise from sequential association with two adaptors, Cdc20 and Cdh1, both containing seven WD40 repeats forming beta-propeller that binds D-box and KEN motifs. During prometaphase and metaphase, APC/C-Cdc20 polyubiquitinates securin and cyclin B to trigger anaphase and mitotic exit. At telophase, cyclin B degradation reduces CDK1 activity, allowing Cdc14 phosphatase to dephosphorylate Cdh1, enabling its binding to APC/C core. APC/C-Cdh1 then operates from late mitosis through entire G1, continuously ubiquitinating mitotic cyclins A and B, mitotic kinase

Ref: Peters, Nature Reviews Mol Cell Biol 2006, APC/C Mechanisms. Alberts 7th ed., Chapter 17.

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