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Important Checkpoint proteins
- Regulatory proteins:
- 1. Cyclin
- 2. Cyclin-Dependent kinases (CDKs)
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End of G1phase
- Cells INCREASE in size
- New organelles produced
- New proteins produced
- Cdk inhibitor DEGRADED
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Entry to S-phase Requirements
Formation of: Cyclin E and cdk2
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Role of cdk2 and Cyclin E
- CDK2 phosphorylates cyclin E
- Phosphorylated CyclinE/CDK2 complex Phosphorylates a tumor suppressor protein RB1
- This causes RELEASE of EF2
- Once Cyclin E duties are done, it is ubiquitinated and degraded via proteasome
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Role of CAK
- Phosphorylates T loop
- Needed to Activate Cyclin E and cdk2
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Role of E2F
- PRODUCES Cyclin E
- Activates genes whose protein products are required for transition from G1>> S phase
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Role of cyclin D/Cdk4,6,
- Involved with INITIATION of signals prior to ENTRY of S phase
- This complex Phosphorylates Rb, which ALLOWS EF2 to activate genes required for transition from G1 >> S phase
- Also with Cmyc
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G2 phase
Rapid Cell growth and Protein synthesis, prior to Mitosis
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G2M Checkpoint
Ensures DNA replication has completed during S phase
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Role of MPF
- Maturing-promoting factor: Cyclin-cdk complex that triggers cell passage PAST G2 checkpoint
- Consists of Cyclin B complex with cdk1 or 2
- 3 residues must be activated on this complex for G2M to occur
- 1. Cdk1 must be Phosphorylate threonine by Cyclin activating kinase (CAK)
- 2 and 3. To ACTIVATE cyclin B/cdk1, cdc25 must remove phosphate on threonine and tyrosine and ADD hydroxy group
- Activity HIGH during M phase
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Metaphase/Anaphase Checkpoint
On each kinetichore, once microtubules are captured and tension detected, Mad2 is released
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Role of CENP-E
- Mitotic motor protein: Uses ATP hydrolysis to drive transport of cellular cargo along microtubules
- 1. essential for attachment of chromosomes (via kinetichore) to spindle microtubules
- 2. Helps chromosomes CONGREGAGE @ equatorial plane and STABILIZE
- 3. Helps develop TENSION across aligned chromosomes
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How CENP-E aids in increasing the tension across bi-orientated kinetichore pairs
- BubR1 (checkpoint kinase) and Bub3 are partiall RELEASED
- Mad2 is blocked: SILENCES Spindle assembly checkpoint upon proper microtubule kinetichore attachment
- This PREVENTS INHIBITION of Anaphase-promoting complex (APC)
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Role of Mad2
- Spindle checkpoint PROTEIN
- Provides biochemical signal to proceed to ANAphase, by being released
- Mad2 required to activate APC/C to go into Anaphase
- Mad2 bound to CDC20 will NOT activate APC/C
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Mad2 and cancer
- Mutations in Mad2 can cause DISRUPTIONS in Spindle Assembly Checkpoint (SAC)
- Decrease in Mad2: Inactivation, can result in aneuploidy, which results in TUMOR formation (via unregulated cell growth)
- Mad2 OVERexpression: Stabilizing proteins DELAY exit from mitosis, cause aneuploidy
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ProphaseKinetichore
assembly Recruitment of CENP-E and Mad2
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Separase
enzyme that Degrades COHESION, allowing ANAPHASE to progress
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Phases of mitosis and meiosis
Look at diagram ONLINE
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Metaphase
- + ends of microtubules attach to sister kinetichores and ALIGN sister chromatids on plane
- Creates TENSION
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Anaphase
Microtubules elongate, helping PUSH spindle poles apart
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