Apoptosis Flashcards
Apoptosis
Programmed cell death
Routine controlled cell death that minimizes spread of damage and or inflammation
Eliminates unwanted cells (tadpole tail)
Necrosis
Accidental cell death
Eg- carrying Petri dish of cells, accidentally drop and step on them
Phenotype of Apoptosis
- overall shrinkage of cell volume and nucleus
- loss of adhesion to neighboring cells
- formation of blebs on surface
- DNA fragmentation
- cytoskeleton collapses
- Nuclear envelope disassembles
- rapid engulfment of dying cell by phagocytosis (macrophages)
What roles do apoptosis play/what is the importance of apoptosis?
- programmed cell death important for certain cells: abnormal, non functional, potentially dangerous
- eliminate lymphocytes after they destroy and ingest microbes
- organs can be kept correct size (liveR)
DNA damaged cells are destroyed if there is enough damage
Biochemistry of Apoptosis
An endonuclease cleaves DNA into a ladders of fragments in distinctive sizes
Cleavages occur in linker regions of chromosome
Agarose gel will show this pattern (a smear vs ladder indicates random cleavage)
Cytochrome C is release from mitochondria during apoptosis (apop marker)
Caspases (Cysteine ASPartyl specific proteASE)
Proteases that mediate the apoptotic cascade intracellularly
Activation is key
Cysteine in active site
Targets proteins and cleaves them in sequence where Asp AA occurs
Pro caspases
Caspases synthesized first as inactive prescursors
Becomes activated by protease cleavage
Cleaves at specific sites to form large and small subunits. These form a heterodimer which can be activated to a caspase by another caspase
Initiator Caspase
Initiates apoptosis
Caspase-8 and Caspase-9
Executioner Caspases
Destroy actual targets- execute process of apoptosis (caspase-3)
Cleaves downstream proteins, inactive endonuclease
Targets cytoskeleton
Attacks cell adhesion proteins- cells roll up in a ball
The Caspase Cascade
Irreversible
There specific caspases: knockout of caspase-3 in mice reduces apoptosis in developing brain and fetus dies with deformation
The machinery for this process is always there
Initiator caspase auto-activates itself
Executioner caspases cleaves cellular targets
Differentiate the two Apoptotic Pathways
Internal and External
Depend on where the stimuli come from
I- Mitochondrial dependent
E- mito independent
Intrinsic Pathway
Cells active apoptosis from inside cells
In response to injury, DNA damage and lack of O2, nutrients, or extra cellular survival signals
Translocation of cytochrome C from intermediate space of mito
It is release to cytosol and binds to adaptor protein (Apaf-1) to activate procaspases
Apoptosome forms (by of Apaf1) activating caspase-9. 9 activates downstream executioner caspase-3
About the Extrinsic Pathway
Extracellular signals bind to cell surface death receptors and trigger pathway (transmembrane proteins)
3 domains:
- extracellular domain
- single transmembrane domain
- intracellular death domain
Receptors are homotrimers
Activates downstream executioner caspases, but there are inhibitory proteins that retrain it. Decoy receptors: FLIP
Homotrimers
Three proteins of the same type (members of tumor necrosis factor family of proteins)
Decoy Receptors
Have ligand binding domain but no death domain.
can bind death ligand but does not trigger apoptosis
FLIP
Protein that resembles initiator procaspases with no proteolytic domain
Competitive inhibitor against procaspases-8 and procaspase-10 (prevents apoptosis)
Describe the process of the Extrinsic Pathway
Fans binds to Fas Death Receptor - ligand form homotrimers
Adaptor proteins are recruited: FADD adaptor (Fas associated death domain) and Procaspase-8 with death effector domain
Formation of trimers: brings death domains together to form DISC (death inducing signal complex)
Activates caspase-8 or -10
Activates downstream executioner caspases - caspase-3 **
Bcl2 family of proteins
Regulate intrinsic apop pathway
Bcl2 controls release of cytochrome C into cytosol
2 types of Bcl2: anti and pro apoptotic
(3 classes, anti, pro BH123, pro BH3)
Anti-apoptotic (prosurvival) Bcl2
Pro-apoptotic Bcl2
- blocks release of cytochrome C (Bcl2 example)
- 4 domains within Bcl2 BCl Homology domains
- Promotes release of cytochrome C
- BH123 protein + BH-3 only protein
What are the three classes of Bcl2 proteins?
1) anti-apoptotic Bcl2 protein
2) pro-apoptotic BH123 protein
3) pro-apoptotic BH3 protein
BH123 protein
Pro-apoptotic
Stimulus triggers intrinsic pathway
Become activated in outer mitochondrial membrane and induce release of cytochrome C.
Then apoptosome is formed by binding to Apaf1
Anti-apoptotic Bcl2 Proteins
Bcl2 and BCl-XL
Mainly located on cytosolic surface of outer mitochondrial membrane
These proteins prevent apoptosis by binding to pro-apoptotic proteins (BH123) and prevent aggregation into active form
Inactive Intrinsic Pathway/Inhibition
Active anti-apoptotic Bcl2 protein is stopping BH123.
Cytochrome C trapped in outer mitochondrial membrane space
Can be stopped by BH3-only
Prevents spread out BH123 proteins from coming together to form allow passage of cyto c and intermembrane proteins
BH3 only protein
Pro-apoptotic, cytosolic protein
Translocated into mito after apoptotic signals activate it.
Inhibits Bcl2 protein from inhibiting aggregation to release cytochrome c (allows BH123 to come together and form channel)
IAPs (inhibitors of apoptosis)
Bind to and inhibit caspases
Some IAPs add ubiquitin to caspases marking them for proteosomal destruction
Block apoptosis by binding caspases
This is GOOD because it can stop the caspases from auto-activating!
Anti-IAPs
Proapoptotic
If apoptotic stimulus is present anti-IAPs are released from mitochondria to block IAP activity so that executioner caspases can be activated
Neutralize IAPs, free/liberate caspases
Disease related to insufficient apoptosis
B Cell Lymphoma
Chromosome translocation causing excessive Bcl2 to be made can over-inhibit apoptosis. This means even severely damaged cells are not rid of.
This can result in cancer.
P53 mutation
P53 is a tumor suppressor gene.
Mutation of p53 means it can no longer arrest cell cycle
It becomes insufficient and does not promote apoptosis
Cells with DNA damage stick around, thus cancer can be generated
Excessive apoptosis
Heart attacks and strokes
Excessive cell death leading to organ failure
