Apoptosis Flashcards
Is apoptosis or necrosis the most common form of cell death?
Apoptosis
True or False:
In apoptosis, cell contents spill out; in necrosis, cells shrink and condense.
False - apoptosis, cells shrink and condense (contents never leak out); necrosis, cell contents spill out.
Give an example of apoptosis being important in development of mice and humans.
Apoptosis (cell death) sculpts hands and feet during embryonic development. Thus, keeping hands and feet from being webbed.
List the phenotype of apoptosis (characteristics).
- overall shrinkage in volume of cell and its nucleus.
- loss of adhesion to neighboring cells.
- formation of bless on surface.
- DNA fragmentation
- cytoskeleton collapses
- nuclear envelope disassembles.
- rapid engulfment of dying cell by phagocytosis (macrophages).
What type of certain cells is programmed cell death important for?
- abnormal cells
- non-functional cells
- potentially dangerous cells
- eliminating lymphocytes after destroying and ingesting microbes
- keep organs the correct size (liver)
- DNA damaged cells are destroyed if bad enough damage
When running an agarose gel to look at the characteristics of apoptotic cells, where does the endonuclease the DNA resulting in a ladder of fragments in distinctive sizes?
Endonuclease cleaves in linker regions of nucleosomes.
What is the key event to initiate apoptosis?
Activation of caspases.
Caspase = Cysteine Aspartyl specific proteASE
What amino acid is the active site in caspases?
Cysteine in active site.
Where does caspase cleave target proteins?
Caspase targets proteins and cleaves them in their sequence where an aspartic amino acid residue occurs.
What inactive precursor is caspases synthesized first as?
Procaspase
To activate procaspase, what is it cleaved to form?
Procaspases cleaved at specific sites to form a large and small subunit which form a heterodimer - caspases activates procaspase.
What are the two major classes of caspases?
- Initiator caspase
- executioner caspase
What do initiator caspases do?
Initiate apoptosis (including caspase-8 and caspase-9)
What do executioner caspases do?
Destroys actual targets - executes apoptosis (includes caspase-3).
True or False:
The caspase cascade is irreversible.
True
What do activated caspases do?
- cleaves downstream proteins
- cleaves inactive endonuclease
- targets cytoskeleton
- attacks cells adhesion proteins (cells roll up in ball)
True or False:
Executioner caspases auto-activates itself.
False - Initiator caspase auto-activates itself.
executioner caspases cleaves cellular targets
What are the two apoptotic pathways?
- internal pathway
- external pathway
Give an example of an internal stimuli that activates the internal apoptotic pathway.
Abnormalities in DNA.
Give an example of an external stimuli that activates the external apoptotic pathway.
Removal of survival factors and proteins of tumor necrosis factor family.
Is the internal apoptotic pathway or external apoptotic pathway mitochondrial dependent?
- Intrinsic pathway is mitochondrial dependent.
- Extrinsic pathway is mitochondrial independent.
What are the 3 domains that compose death receptors in the extrinsic apoptotic pathway?
- extracellular binding domain
- single transmembrane domain
- intracellular death domain
What is apoptosis?
A routine controlled cell death that minimizes spread of damage and/or inflammation.
Is cell death under physiological conditions.
Clean way of dying.
Apoptosis is programmed cell death.
Apoptosis is an intracellular proteolytic cascade mediated by proteases called caspases.
True or False:
To initiate the extrinsic apoptosis pathway, extracellular signals bind to cell surface death receptors and trigger the extrinsic pathway.
True
Are the death receptors of the extrinsic pathway homotrimers or heterotrimers?
homotrimers - 3 proteins of same type (members of TNF family of proteins)
In the apoptosis extrinsic pathway, what ligand binds to Fas death receptors?
Fas
Once Fas binds to Fas death receptors in he extrinsic apoptosis pathway, what adaptor proteins are recruited?
- FADD adaptor (Fas associated death domain)
- Procaspase-8 with death effector domain
What forms the DISC complex in the extrinsic apoptosis pathway?
- FADD adaptor protein (binds to Fas death receptor)
- Procaspase-8 or 10 (binds to death effector domain, which is a FADD adaptor domain)
In the extrinsic apoptosis pathway, what does activated caspase-8 or 10 activate?
Executioner caspases (e.g., caspase-3)
What is the role of decoy receptors in the extrinsic apoptosis pathway?
- Decoy receptors have ligand binding domain but no death domain.
> bind death ligand but does not activate apoptosis
> thus, they are inhibitory proteins that restrain the extrinsic pathway
In the extrinsic apoptosis pathway, what is the function of FLIP proteins?
- is a competitive inhibitor against procaspase-8 and procaspase-10 = prevents apoptosis.
- FLIP is a protein resembling initiator procaspase with NO proteolytic domain.
List some reasons that a cell would activate apoptosis from inside of the cell via the intrinsic pathway.
- in response to injury
- DNA damage and lack of oxygen
- nutrients
- extracellular survival signals
What is the key event of the intrinsic apoptosis pathway?
The translocation of cytochrome C from the intermediate space of mitochondria is the key event.
In the intrinsic apoptosis pathway, where is cytochrome c released from and what is its function?
- cytochrome c is translocated from intermediate space of mitochondria into the cytosol.
- cytochrome c binds to procaspase-activating adaptor protein Apaf1 (apoptotic protease activating factor-1).
- Apaf1 forms apoptosome which activates caspase-9.
- Caspase-9 activates downstream executioner caspases (caspase-3, which is common to both pathways).
What triggers the assembly of the apoptosome?
Release of dADP in exchange for dATP (or ATP).
What happens during the event of cytochrome c activation of Apaf1?
Activation of Apaf1 by cytochrome c and hydrolysis of bound dATP to dADP.
Remember: The assembly of apoptosome is then triggered by the releases of the dADP in exchange for dATP (or ATP).
What protein regulates the intrinsic apoptotic pathway by controlling the release of cytochrome c into the cytosol from the intermediate space of the mitochondria?
Bcl2
(Bcl = B Cell Lymphoma)
What are the two different types of Bcl2 proteins that regulate the intrinsic apoptotic pathway?
- Pro-apoptotic
- Anti-apoptotic (pro-survival)
What does the pro-apoptotic Bcl2 protein function to do in the intrinsic apoptotic pathway?
Promotes release of cytochrome c.
What does the anti-apoptotic Bcl2 protein function to do in the intrinsic apoptotic pathway?
Blocks the release of cytochrome c.
What are the distinctive domains of anti-apoptotic protein Bcl2?
Has 4 distinctive domains - called Bcl homology domains or BH domains.
What are the domains of pro-apoptotic protein Bcl2 in the intrinsic apoptotic pathway?
BH123 protein + BH-3 only protein.
What are the 3 classes of Bcl2 proteins?
> Anti-apoptotic Bcl2 protein (BH1234)
- e.g., Bcl2, Bcl-XL
> Pro-apoptotic BH123 protein
- e.g., Bax, Bak
> Pro-apoptotic BH3- only protein
- e.g., Bad, Bim, Bid, Puma, NoxaWhat is the mechanism of BH123 protein in apoptosis?
- apoptotic stimulus triggers intrinsic pathway
- BH123 proteins become activated
- from aggregation in mitochondrial outer membrane and induce release of cytochrome c
- then apoptosome formed by binding to Apaf1
Where are anti-apoptotic Bcl proteins, like Bcl2 and Bcl-XL, mainly located?
Cytosolic surface of outer mitochondrial membrane.
What do anti-apoptotic Bcl proteins, like Bcl2 and Bcl-XL, function to do?
These proteins prevent apoptosis by binding to pro-apoptotic proteins (e.g., BH123) and prevent aggregation into active form.
What happens when BH3- only protein (pro-apoptotic) is activated?
Activated BH3-only protein is cytosolic and translocates to mitochondria after apoptotic signal activates it (pro-apoptotic).
It inhibits anti-apoptotic Bcl2 protein from inhibiting aggregation to release cytochrome c.
What do IAP proteins function to do in the regulation of apoptosis?
- IAPs = Inhibitors of Apoptosis
- bind and inhibit caspases
- some IAPs add ubiquitin to caspases to mark them for destruction by proteasome
- IAPs block apoptosis by binding to caspases
What is the role of anti-IAPs in the regulation of apoptosis?
If there is apoptotic stimuli or apoptosis signals…….then this causes release of anti-IAPs from mitochondria to block activity of IAPs.
Executioner caspases can therefore be activated with the IAPs blocked.
anti-IAPs neutralize the IAPs and liberate the caspases
Insufficient apoptosis can contribute to disease.
> Due to chromosome translocation that causes excessive Bcl2 to be made - Bcl2 is inhibitor of apoptosis.
Excess Bcl2 promotes development of cancer by inhibiting apoptosis - DNA-damaged cells will not be programmed for cell death but can go on to cause cancer.
> p53 mutation:
- mutated p53 can no longer cause cell cycle arrest
- and no longer promotes apoptosis
- cells with DNA damage stick around and cancer can be generated
excessive apoptosis can be a problem too - this occurs in heart attacks and strokes
