Apoptosis Flashcards
What are the three types of cell death?
1 - Apoptosis (active, no inflammatory response)
2 - Autophagy
3 - Caspase-independent (Necrosis)
What is apoptosis important for?
- Normal development (sculpting body parts and the immune system)
- Deleting damaged cells due to infection or genetic abnormalities to stop propagation.
- Tissue homeostasis (maintain appropriate cell numbers)
What are the cellular events in apoptosis?
SIGNAL - cytotoxic drugs, chemo, radiation, survival factor removal
TRANSDUCTION - biochemical pathways
EXECUTION - caspase protease cleavage and activation to cause apoptosis.
Phosphatidylserine markers act as signals so apoptotic cells can be cleared by macrophages.
DNA is cleaved by CAD (caspase activated DNAse) to cleave DNA into small fragments.
What is caspase and what is its structure?
Cys-dependent aspartic acid specific proteases.
Prodomain, large, small subunit structure.
Cleaved between large and small and dimerise to activate.
What is the general mechanism pathway for caspase activation? What is one substrate example for this process?
Initial trigger > INITIATOR caspase (8,9) > EFFECTOR caspase (3,6,7) > Cleave downstream target and provide feedback amplification loop to initiators.
ICAD - Inhibitor of Caspase activated DNAse, Degradation leads to DNA cleavage.
What is the mechanism of extrinsic caspase activation?
Cell surface death receptors (Fas, TRAIL) activated by cell surface DR-ligands bound to other cells.
Adaptor protein FADD dimerises and activates pro-caspase 8 and forms DISC, which fully activates Caspase-8.
What is the mechanism of intrinsic caspase activation?
Triggered by the release of cytochrome C release from the mitochondria.
Caused by steroids, DNA damages, p53, ROS
Cytochrome C inactivates Apaf-1 regulatory domain so it can oligomerise to form the APOPTOSOME.
Can then interact with pro-caspase 9 via CARD domains to fully activate caspase 3.
What are the three subfamilies of the BCL-2 family of apoptosis-regulating proteins?
- BCL-2 (Bcl-2, Bcl-XL) (pro-survival, reduces CytC)
- BAX (Bax) (pro-apoptotic, CytC release)
- BH3-only (Noxa, Puma, Bim, pro-apoptotic)
Draw structures of BCL-2, BAX and BH3-only
Draw
Describe the mechanism of CytC release in apoptosis.
BCL-2 restrains BAX.
After an apoptotic signal, there is an increase in BH3-only proteins that block the anti-apoptotic BCL-2 proteins, releasing BAX.
BAX undergoes conformational change and inserts into the mitochondrial membrane, creating a pore that releases Cytochrome C for caspase activation.
How is p53 regulation involved in apoptosis?
p53 can activate BH3-only molecules (Noxa, Puma).
ATM kinase (DNA repair gene) phosphorylates Mdm2 (p53 regulator), blocking p53 degradation.
Causes cell cycle arrest and apoptosis through p53 activation of Noxa/Puma - block BCL-2, induce BAX.
Cells with a mutant p53 do not undergo apoptosis following DNA damage and accumulate mutations.
How do BCL-2 mutations cause cancer?
Give examples
Overexpression of BCL-2 suppresses apoptosis in many forms, contributes to the resistance of cancer therapies.
- Follicular lymphomas - chromosome translocation (14:18) = fusion gene to Ig that causes inappropriate overexpression.
- Chronic Lymphocytic Leukaemia (CLL) - deletion at chromosome 13Q = no miRNA that reduces BCL-2, causing overexpression.
How do BAX mutations cause cancer?
Give examples
If BAX is mutated, it cannot cause apoptosis.
- HNPCC = truncated BAX mutation, cannot cause apoptosis.
Why have agents that triggered apoptosis failed? What therapies are being produced instead and what can they treat?
Apoptosis triggering agents fail due to easily becoming resistant / quick cancer evolution.
- VENETOCLAX (ABT-199) - mimics BH3-only proteins by binding BCL-2 groove, meaning any BH3-only proteins produced, go straight to cause the conformational change in BAX and cause apoptosis.
Only works for BCL-2, not BCL-XL.
- Cerdulatinib = multi-therapy with venetoclaxto target multiple apoptotic pathways.
Treats CLL and follicular lymphoma.
