Session 7 - The real cell cycle Flashcards
Outline the stages of cell cycle
G0 – Senescence
G1 – Growth of cell
S phase – DNA increases from 2n to 4n. Generates non-diploid number of chromosomses
G2 – Reorganised interphase cromatin into chromosomes.
M – Mitosis occurs, with multiple phases within (pro met anna on the telephone). DNA transitions from 4n to 2n.
What are the three main checkpoints of the cell cycle?
G1 checkpoint - pRb
G2 checkpoint - p53
M checkpoint
How can we detect the phases of the cell cycle?
Flow cytology allows us to detect the phases of the cell cycle by measuring amount of DNA in cell at any one time
What is the main protein involved at the G1 to S phase checkpoint
pRb
What stimulates cell cycle through pRb checkpoint
Hormones, growth factors and cell contact
What are the three main proteins involved in regulation of pRb?
Cyclins
Cyclin dependent serine/threonine kinase
Cyclin dependent kinase inhibitors
What does phosphorylation of pRb do?
Inhibits it
What does hypophosyphorylation of pRb do?
Activates it
What are four mechanisms that regulate cyclin activity?
- Association with cyclins
- Association with CDK inhibitors
- Inhibitory phosphorylation (P21)
- Activating phosphorylation
What cyclins act at what points?
- G1 – Cyclin D
- G1/S – Cyclin E
- S – Cyclin A
- M – Cyclin B
What are the two mechanisms of action of pRb?
Prevents cell cycle progression via production of HDAC1 which prevents DNA transcription by causing chromatin condensation, and by not releasin transcription factor E2F
What do CDKs do to pRb?
If CDKs are active, they will phosphorylate pRb (the master brake), preventing it from inhibiting the release of the transcription factor E2F. When E2F is released it stimulates transcription which allows the cell cycle to progress (by stimulating production of Cyclin E and Cyclin D, as well as DNA polymerase)
What does HDAC1 do?
HDAC1 protein activated by pRb causes deacetylation and condensation of chromatin, decreasing transcription. This occurs when pRb is not phosphorylated.
What does active pRb do?
Active pRb – E2F production inhibited, HDAC1 activated -> Cell cycle halted
What does inhibited pRb do?
Inhibited pRb – E2F activated, HDAC1 inhibited -> Cell cycle stimulated
How does p53 moderate cell cycle (with regards to checkpoint 1)
P53 also moderates cell cycle at G1, by blocking cycle progression until DNA repair is completed. It does this by triggering hypophosphorylation of pRb which prevents release of E2F and thus prevents progression of cell cycle.
What is the exact mechanism of pRb inhibition?
Requires phosphorylation via CDK4 (cyclin D attached) followed by further phosphorylation by CDK2 (cyclin E attached).
What is the exaction mechanism of CDK inhibition?
P16(INK4A) is responsible for the binding to CDKs and preventing their interaction with pRb. This effect is mimicked by p21, p15.
What is the wnt signalling path?
Growth stimulating pathway
What is the cell surface receptor for WNT?
Frizzled receptor
What is activated when WNT ligand binds with frizzled?
Phosphoprotein dishevelled activatied
What are the two pathways of WNT?
Canonical and non-canonical
What does WNT path do?
Causes accumulation of B-catenin in cell cytoplasm via inhibition of destruction pathway. B-catenin translocates to nucleus, activating cyclin D1 and D2 transcription, and represses p16. This promotes entry into S phase.
How is WNT path upregualted?
Increased expression of Wnt ligand proteins or B-catenin causes activation of canonical path and thus activation of cell cycle.
What is the purpose of the G2/M checkpoint?
DNA replication check point, found after DNA synthesis
What is G2/M regulated by?
Mediated by MPF (maturatin promoting factor). This is a protein composed of Cyclin B and CDK1.
This protein phosphorylates multiple substrates to trigger entry into mitosis.
What can MPF be inhibited by?
This can be inhibited by double strand breaks (ATM produced which activates WEE1 to phosphorylate MPF and inhibit it) or DNA damage (ATR -> WEE1 -> MPF phosphorylation).
ATM and ATR also activate P53, which activates CKI P21 and further inhibits cell cycle.
What occurs in ataxia telangictasia?
ATM and ATR can be mutated which means this checkpoint does not work, leading to replication of cells with DNA damage.
What does CDK2NA do?
Activates pRb AND p53.
What are two apoptotic pathways?
Intrinsic and extrinsic pathway
What do intrinsic and extrinsic path converge on?
Capse 3
Outline the extrinsic path of apoptosis
TNF-a, TRAIL bind to membrane (trail-R) and cause capsase activation and cell apoptosis. This is regulated by p53, which moderates productions of death receptors and capsases.
Outline the intrinsic path of apoptosis
Apoptosome is formed by cytochrome C and APAF-1 -> This can only form as a result of increased permeability of mitochondria, and a result of Bcl-2, Bak and Bax (all triggered by P53!).
What is required for the intrinsic path to proceed?
For intrinsic pathway to proceed via >mitochondrial permeability, pro-survival BCL-2 family must inhibited and BAX or BAK must be activated (both of these things done by P53!)
