Apoptosis, Autophagy, Necrosis Flashcards
(37 cards)
two major pathways of apoptosis (programmed cell death)?
intrinsic: triggered by intracelular stress (oxidative damage, ischemia)
extrinsic: triggered by death ligand (tumor necrosis factor, TNF)
what is the basic progression of intrinsic pathway of apoptosis (naming molecules involved)?
- intracelular stress
- mitochondria cytochrome c
- APAF1
- caspase 9 (this step inhibited by Bcl-2)
- caspase 3
- apoptosis execution by DNA fragmentation
what is the basic progression of extrinsic apoptosis pathway? (naming molecules involved)
- extrinsic signal (death ligand such as TNF)
- FADD
- DISC formation
- caspase 8
- caspase 3
- apoptosis execution by DNA fragmentation
how do pro-apoptotic signals affect mitochondria in intrinsic apoptosis pathway?
pro-apoptotic stimuli increase permeability of outer membrane and promote mobilization of Cytochrome C (usually bound to cardiolipin, CL)
in cytosol, cytochrome C activates APAF1 and proteolytic maturation of caspase-9 and caspase-3
[reminder - cyt c functions as electron shuttle in ETC, but also functions as apoptosis signal]
how do Bcl-2 family proteins influence apoptosis in intrinsic pathway
B-cell lymphoma-2 (Bcl-2) superfamily:
pro-apototic: Bax, Bak - promote permeability of mitochondria and Cyt C release
anti-apoptotic: Bcl-2, Bcl-XL - bind Bax/Bak to inhibit
which Bcl-2 proteins are pro-apoptosis, which are anti-apoptosis?
Bax and Bak are PRO-apoptosis
Bel-2 and Bcl-XL are ANTI-apoptosis and bind/inhibit Bax/Bak
how is Bcl-2 implicated in cancer?
in follicular lymphoma, a chromosomal translation commonly occurs which leads to high levels of Bcl-2 expression
this causes decreased propensity to undergo apoptosis —> increased cancer growth
Bcl-2 overexpression linked to poor disease outcomes in several cancers
how does p53 function to prevent cancer formation? give 3 ways
initiates apoptosis (blocks anti-apoptosis Bcl-2 and Bcl-XL while promoting pro-apoteosis Bak/Bax)
activates DNA repair proteins
arrests growth by holding cell cycle at G1/S checkpoint
how does APAF1 function in intrinsic pathway of apoptosis
APAF1 = apoptotic protease activation factor 1
cytochrome C (released from mito) activates APAF1 by displacing it from CARD (CAspase Recruitment Domain)
CARD domains form cluster of 7 —> apoptosome
*apoptosome cleaves procaspase 9 to form caspase 9 (initiator caspase), which activates cas3/6/7 (executioner caspases)
the molecular “hitmen” are
caspases - enzymes that give rise to morphological and biochemical changes arising form apoptosis
must be CLEAVED to be active
regulated at post-translational level so they can be rapidly activated
group these into apoptosis initiators and effectors:
caspase
2, 3, 6, 7, 8, 9, 10
caspase initiators: 2, 8, 9, 10
caspase effectors: 3, 6, 7
what are the pro-apoptotic receptors in the extrinsic pathway? what do they bind?
DR4 and DR5
bind endogenous Apo2L (apoptosis-inducing ligand 2) and TRAIL (TNF-related apoptosis-inducing ligand)
after activation, what do DR4 and DR5 recruit? (extrinsic apoptosis) What steps occur next?
recruit FADD: Fas-associated death domain
FADD recruits Caspase 8 and/or 10 (initiator caspases) to the DISC (death-inducing signaling complex)
cas-8 and 10 are released into cytoplasm, activate effectors (cas3/6/7)
what are the ligands of each of these TNF-family death receptors of extrinsic apoptotic pathway?
a. FAS
b. DR5
c. TNFR1
d. DR3
FAS binds FasL (Fas ligand)
DR5 binds TRAIL/Apo2L
TNFR1 binds TNFalpha
DR3 binds Apo3L
T/F: extrinsic pathway stimulates apoptosis independently of p53 and mitochondria
TRUE
fill in the blanks of extrinsic apoptotic pathway:
1. ligands bind _____
2. ______
3. caspases _____
4. caspases _____
- ligands bind DEATH RECEPTORS
- FADD/DISC
- caspases 8, 10 (initiators)
- caspases 3, 6, 7 (effectors)
where do extrinsic and intrinsic apoptotic pathways converge?
activation of effector caspases 3, 6, 7
what is a distinctive feature of DNA degradation by caspase-activated DNase(CAD) when detecting apoptosis?
DNA laddering - biochemical marker of apoptosis
what are 2 biochemical markers of apoptosis?
- DNA laddering
- caspase activation (can be measured via expression levels) - must be cleaved to become active
how can TUNEL assay be used to detect apoptosis
TUNEL = Terminal Transferase dUTP Nick-End Labeling)
measurement of DNA fragmentation
TdT (terminal deoxynucleotidyl transferase) adds dNTPs to 3’ end of DNA molecules in absence of a template
in imaging, apoptotic cells appear as blebs
macroautophagy vs. microautophagy vs. chaperone-mediated autophagy
macroautophagy: cytosolic components are sequester within double membrane organelle or autophagosome, which fuses with lysosome (forms autolysosome) for degradation
microautophagy: lysosomes DIRECTLY engulf cytosolic components via invagination
chaperone-mediated autophagy: chaperone protein and cytosolic protein form complex that is recognized by LAMP-2A which translocates it into lysosome
what recognition protein is required for chaperone-mediated autophagy
LAMP-2A: translocates chaperone protein/ cytosolic protein complex to lysosome
4 stages of autophagy
- induction
- autophagosome formation
- autophagosome-lysosome fusion
- autophagosome breakdown
autophagy is inhibited by ___ conditions and stimulated by ____ conditions
starvation STIMULATES autophagy
nutrient-rich conditions INHIBIT autophagy
