L5 apoptosis II part 2 Flashcards
Extrinsic pathway?
Extrinsic Pathway of Apoptosis:
Involves circulating lymphocytes, specifically T cells.
Granzyme and Perforin Mechanism:
T cells release granzymes (proteases) and perforin (a pore-forming protein).
Perforin creates pores in the target cell membrane, allowing granzymes to enter.
Granzymes cleave initiator caspases, triggering the apoptotic cascade in the target cell, thus activating caspases.
Death Receptor Pathway:
Target cells express death receptors, and T cells express death ligands that interact with these receptors.
One example of this interaction is FasL/Fas:
FasL (Fas ligand) on the T cell binds to Fas (a death receptor) on the target cell.
Fas is a member of the TNF receptor family, all of which contain ‘death’ domains.
The Fas/FasL interaction:
Important for T cell control and activation-induced cell death.
Helps limit the duration of the immune response.
Fas-mediated death of B cells is caused by FasL on activated T cells.
What happens when FasL binds to Fas?
FasL/Fas Interaction:
FasL (Fas ligand) on the T cell membrane is a trimer, while the Fas receptor on the target cell is a monomer.
When FasL binds to Fas, it causes Fas to trimerize, exposing sequences in its death domain.
Death Domain Activation:
The exposed death domain of Fas recruits FADD (Fas-associated death domain protein), which acts as an adaptor protein.
FADD bridges the Fas receptor and the initiator caspase, typically caspase-8 in its pro form.
This recruitment leads to the activation of caspase-8, which then cleaves effector caspases.
Caspase Cascade:
The activated initiator caspase (caspase-8) cleaves and activates effector caspases.
These effector caspases cleave ICAD (inhibitor of CAD), releasing CAD, which enters the nucleus to fragment DNA.
Death-Induced Signaling Complex (DISC):
The Fas/FADD/caspase-8 complex forms the Death-Induced Signaling Complex (DISC), a multi-protein complex that drives the apoptotic signaling cascade.
other receptors that can induce death or signalling?
TNF and the TNF Receptor Family:
FasL and Fas are part of the TNF receptor family.
TNF (Tumor Necrosis Factor) can induce apoptosis or other immune cell functions depending on which adaptor proteins the cell expresses.
TNF Receptor Activation:
The TNF ligand on the T cell binds to the TNF receptor on the target cell.
Upon binding, TNF proteins associated with the death domain are recruited to the trimeric TNF receptor.
This recruitment leads to the activation of early caspases, which in turn recruit subsequent caspases to initiate apoptosis.
Alternate Signaling Pathways (Without FADD):
In cell types that do not express the FADD adaptor protein, alternate molecules can bind to the TNF receptor and initiate signaling instead.
TRAF-2 (TNF receptor-associated factor-2) can be recruited, which then initiates the NF-κB (and jun) signaling pathway, leading to immune responses rather than apoptosis.
Conclusion:
TNF can either induce apoptosis or activate signaling pathways based on which adaptor proteins are expressed in the target cell.
intrinsic pathway?
activation of caspases is under tight control by bcl-2 proteins. mitochondria are central to apoptosis induction. Another way to initiate apoptosis is when cells sense something wrong, like withdrawal of trophic signals or accumulation of DNA damage. This activates the intrinsic pathway, which involves the mitochondria and the release of cytochrome c.
bcl-2 family proteins?
The Bcl-2 family regulates the intrinsic apoptotic pathway by either promoting or preventing apoptosis.
They act on the mitochondria.
Named after the B-Cell-Lymphoma gene, which was involved in a chromosomal translocation leading to lymphoma.
Anti-apoptotic proteins like Bcl-2 prevent apoptosis and give cells immortality and resistance to apoptosis (pro-survival proteins). Pro-apoptotic proteins like Bax and Bad promote apoptosis, even though they share a similar domain structure to anti-apoptotic proteins.
There is a delicate balance between pro- and anti-apoptotic expression. In cancer, pro-survival genes are often overexpressed, suppressing apoptosis, while pro-apoptotic proteins are lost, enhancing cell survival.
bcl-2 family proteins and cytochrome C release?
cythcrome c: component of ETC
APAF-1: apoptotic protease activating factor. In a normal cell cytochrome c is present only in mitochondria.when apoptosis is induced by the intrinsic pathway, cytochrome c is released from the mitochondria through pores as the mitochondria begin to swell. Cytochrome c binds to Apaf-1. apaf-1/cytochrome c complex activates initiator caspases. These initiator caspases then activate downstream caspases, which cleave caspase substrates like ICAD, leading to DNA fragmentation.
bcl-2 binds to mitochondrial membranes, blocking swelling and so blocking process that leads to cell death.
The Bcl-2 family regulates the permeability of the mitochondrial membrane, controlling the release of cytochrome c during apoptosis.
apoptosis regulation at mitochondrial outer membrane?
Apoptosis is controlled at the outer mitochondrial membrane (OMM).
Pro-apoptotic Bcl-2 family members oligermarize at MOM and favour MOM permeabilization. they are small cytoplasmic proteins with membrane-binding alpha helices.
When pro-apoptotic BH3-only proteins (such as BAD or BID) are triggered, they bind to pro-apoptotic proteins (like BAX).
This binding causes BAX to spread out its helices, allowing it to insert into the outer mitochondrial membrane (OMM).
BAX oligomerizes (forms a complex), and just like perforin in immune cells, BAX forms a pore in the OMM.
The BAX pore enhances the membrane’s permeability, allowing cytochrome c to leak out of the mitochondria, which is crucial for activating the intrinsic apoptotic pathway.
Pro-survival bcl-2 fam members e.g bcl-2 suppress bax oligermization and favour MOM integrity.
cytochrome c and apoptosome formation?
Cytochrome c activates the apoptosome, which is a heteromeric complex.
Cytochrome c binds to Apaf-1, causing a conformational change in Apaf-1.
This change activates the ATPase activity of Apaf-1, and when it hydrolyzes ATP, it assembles into a ring of 7 Apaf-1 molecules.
This Apaf-1 ring provides a platform for initiator caspases like caspase 9 to bind, cleave, and activate.
Once activated, caspase 9 can cleave effector caspases, continuing the apoptotic cascade. Capase 9 is the initiator which then activates caspases 3,7 (effectors) and auto-cleaves itself in the apoptosome.
This process amplifies the apoptotic signal because the release of cytochrome c triggers the assembly of 7 Apaf-1 molecules into a single complex, allowing simultaneous activation of caspases. If fewer than 7 Apaf-1 molecules are present, the complex cannot form and the apoptotic process won’t proceed.
BID?
The extrinsic pathway is typically activated by signals from outside the cell (e.g., death ligands binding to death receptors like Fas or TNF receptors).
This activation leads to the activation of caspase 8, which is an initiator caspase in the extrinsic pathway.
Caspase 8 can cleave and activate BID, a pro-apoptotic member of the Bcl-2 family.
The cleaved BID (now called tBID) then translocates to the mitochondria, where it promotes the release of cytochrome c into the cytoplasm, which is the key step in the intrinsic pathway.
The release of cytochrome c from the mitochondria activates the apoptosome and initiator caspases (like caspase 9), which then trigger the effector caspases that carry out apoptosis.
cell survival by PI-3 kinase signalling?
Bad, a pro-apoptotic BH3 only protein is sequestered by a 14-3-3 protein when phosphorylated by akt/PKB:
Without trophic factors: If trophic factors (signals promoting cell survival) are removed, pro-apoptotic proteins like BAD can promote apoptosis. BAD resides on the MOM and promotes permeability pores, which allows cytochrome c to leak out. This activates the apoptosome, which in turn activates initiator caspases like caspase 9, and these then activate effector caspases, leading to apoptosis.
With trophic factors: When trophic factors are present, they bind to their respective receptors on the cell membrane and activate a PI3-kinase (PI3K) signaling cascade. PI3K generates a lipid called PIP3 on the plasma membrane, which recruits and activates Protein Kinase B (Akt).
Akt’s role: Akt then phosphorylates BAD at a specific site. This phosphorylation prevents BAD from acting at the mitochondria.
14-3-3 proteins: After BAD is phosphorylated, it is recognized and bound by 14-3-3 proteins, which are a family of adaptor proteins in the cytosol. These 14-3-3 proteins sequester phosphorylated BAD, essentially “holding it in check” in the cytosol.
When BAD is phosphorylated by Akt, it binds to 14-3-3 proteins.
The 14-3-3 proteins sequester phosphorylated BAD in the cytosol, preventing it from interacting with the mitochondrial membrane.
When BAD is not at the mitochondria, it cannot promote pore formation in the mitochondrial membrane. As a result, cytochrome c is not released, and the intrinsic apoptotic pathway is inhibited.
So, in summary:
No trophic factors → BAD induces cytochrome c release, activating apoptosis.
With trophic factors → PI3K/Akt pathway phosphorylates BAD, sequestering it with 14-3-3, preventing apoptosis.
This is an example of post-translational regulation where the activity of BAD is controlled by its phosphorylation and sequestration, which ultimately determines whether the cell survives or undergoes apoptosis.
removal of apoptotic fragments?
Phosphatidylserine externalizes during apoptosis, signaling phagocytes to engulf and digest the dying cell. ost-engulfment consequences: Postengulfmanent consequences: This also leads to the release of anti-inflammatory cytokines (TGFB, IL-10, PGE2) to prevent excessive immune activation. + cytosheletal rearrangement
‘find me’ signals released by nucleotides such as ATP or UTP released lipids such as lysophophatidylcholine or sphingosine-1-phosphate.
