Apoptosis Flashcards
How does apoptosis proceed
Initiates a program, at the end of the program the cell in engulfed by a macrophage.
Steps in apoptosis
- DNA is condensed
- Shrinkage of the cytoplasm
- Membrane blebbing
- DNA cleavage
- Phosphotidyleserine is transported to the outer membrane (macrophage eat me signal)
- Tight regulaiton
- No inflammation
- Tight control
- Energy is required
Steps in necrosis
- Cytolysis: membrane destruction
- Inflammation
- Random DNA cleavage
Two types of apoptosis
What is similar and different about them
Extrinsic and intrinsic, the only difference is where do you get the signal from.
In both of the mechanism the cells activate caspases which are enzymes that eat proteins. There is also MOMP (mitochondria outer membrane permealization) which permealizes the outer membrane of the mitochondria
Explain extrinsic apoptosis
When a particular ligand is secreted, our cells have what we call the death receptor. The death domain on the death receptor recruits scaffolding proteins which then recruit caspase 8 from the cytosol. Caspase 8 is then activated (self activated), releases from the receptor and activates MOMP.
There are precaspases and effector caspases. Initiator caspases are the ones that are recruited to the receptor
What is a zymogen
An inactivated protein
Zymogen version of a caspase
Procaspase
How are procaspases activated
They have an alpha subunit and a beta subunit. They are first sequestered around the death receptor (alot of Caspase 8 is sequestered) and then they are cut. They form dimers in their mature form and then they go on to mature effector caspases.
Biochemical basis of cutting the caspases
They have dimerized structures and floppy loops coming out of them. These loops are need to be cut to activate the caspases
They are always cut at the aspartate residue
Common things between all caspases
They have a common cystiene residue at the active site.
What is done to regulate the caspases in a cell
There are proteins called the Inhibitor Apoptosis Protein (IAP). What they do is that when they see a caspase, either active or inactive, they bind to it and close the active site
How is apoptosis initiated in the presence of caspase regulation
IAP mediated inhibition is abrogated by smac/DIABLO during apoptosis
What are over expressed in cancers
Death receptors
What happens downstream of Caspase
- The kinases are cut, now they are more active and they will phosphorylate whatever they can
- There is deterioration of the cell cytoskeleton
- PARP destruction, DNA repair mechanism is now impaired. Capase cleaves PARP
- DNA fragmentation by caspase activated DNAase
- Nuclear envelope is degraded by the destruction of the lamin in the envelope
- Finally phosphotidylserine from the inside of the cell membrane is flipped to be exposed outside for the macrophage to phagocytose the cell
How is the DNA cut in apoptosis and how this is avoided in a functioning cell
This is avoided by the protein called Inhibitors of Caspase Activated DNAse (ICAD).
During apoptosis Caspase 3 cleaves ICAD and so now the Caspase activated DNA is turned on and cleaves the DNA
What does caspase do to the nucleas during apoptosis
It degrades nuclear lamins
What are the 2 proteins associated with activating or deactivating eat me signal
Scrambalase flips this signal out, activating englufement whereas flippase puts this signal back in. During apoptosis flippase is cleaved.
Explain intrinsic apoptosis
Due to stress there is activation of a transcription factor called p53. This transcription factor leads to the activation of gene transcription, this leads to PUMA. This is a BH3 binding domain.
Also increased stress can lead to activation of the BH3 proteins
What are the 2 kinds of BCL-2 protein
How do these proteins work
They are either pro-apoptopic or anti-apoptopic
So basically we have anti-apoptopic family members that are bound to the pro-apoptopic family members. When BH3 is activated it binds to the anti-apoptopic family members and takes them away from the pro-apoptopic genes that codes for pro-apoptopic proteins. Now these genes are transcribed and they cause MOMP leading to apoptosis
What mechanism is a commonality between the extrinsic and intrinsic apoptosis
The activation of pro-apoptopic BCL 2 proteins
How does the pro-apoptopic BCL-2 proteins act
They form a pore and incorporate themselves into the mitochondria forming pores.
Know what MOMP causes death of mitochondria
Proton gradient is not formed, there are more ROS.
Cytochrome c leaks out in the cytoplasm, it activates caspases, it is a super activator, t forms an apoptosome
What process is involved in ubiquitinilation
E1, E2 and E3 proteins are involved (3 is most in number and most specific, followed by E2 and then E1). E1 is activating enzyme, E2 is conjugating enzyme and E3 is ligaing enzyme (it is the one that trasnfers the ubiquitin on the protein that needs to be degraded).
What does p53 do
It binds to DNA (since its a transcription factor) leads to shuttind down the cell cycle or if the signal is very strong, it will lead to cell death.
How is p53 regulated
MDM2 is bound to p53. MDM2 is itself an E3 so it facilitates in p53 degradation by proteosome
What does cancer cells do regarding death receptor
They over express those (well some of the cancer cells over express them)
Important thing to remember about p53 and death receptors
Death receptors cause apoptosis in an p53 independent manner
What are the causes of lupus
- Increases apoptosis of monocytes and keratinocytes mediated by increased p53 activity
- Increased T cell necrosis
- Impaired mitochondrial function
- Loss of apoptopic cell clearence
- Increase in auto-antigen presentation
