L4-5 apoptosis

Question 1

what is apoptosis?

Answer 1

programmed cell death, occurs in normal development and in response to problems
e.g. death of cells in tail of tadpole/death of cells with excessive DNA damage

Question 2

what is necrosis?

Answer 2

rapid accidental death of an injured cell

Question 3

what is the basic process of apoptosis?

Answer 3

1. cell shrinks
2. plasma membrane forms ‘blebs’
3. cytoskeleton collapses
4. nuclear envelope disassembles
5. nuclear DNA broken into fragments - leads to ‘DNA laddering’
6. cell surface altered leading to phagocytosis by neighbouring cell

Question 4

what are the types of degradative enzymes involved in apoptosis?

Answer 4

proteases and DNAases

Question 5

what are caspases?

Answer 5

proteases with cysteine at active site which cleave targets at aspartic acid residues

Question 6

what are the inactive precursors caspases are synthesised as?

Answer 6

procaspases, active caspases cleave procaspases

Question 7

what is the name given to when each caspase molecule can cleave many procaspase molecules?

Answer 7

caspase cascade

Question 8

what are the 2 categories of caspase?

Answer 8

1. INITIATOR caspases - activate procaspases, have extra-long prodomains through which they associate with adaptor proteins to form activation complexes.
2. EFFECTOR caspases - break down target proteins e.g. Lamin/DNase

Question 9

how do external signals initiating apoptosis work?

Answer 9

act via stimulation of death receptor proteins on cell surface, e.g. destruction of damaged/virus-infected cells by cytotoxic T lymphocytes (WB cells)

Question 10

process of recognition of infected cell by cytotoxic T cell

Answer 10

1. peptides produced by incomplete protein breakdown in proteasomes are exported to cell surface bound to MHC (major histocompatibility complex) proteins. (peptides can be foreign or endogenous)
2. target cell presents peptides bound to MHC protein on surface so the T cell receptor on cytotoxic T cell binds to MHC
3. cytotoxic T cells recognise peptides from pathogen proteins but DO NOT recognise peptides from organisms own proteins (self-tolerance)

Question 11

what is apoptosis dependent on?

Answer 11

depends on T cell recognising protein on MHC complex

Question 12

what is Fas?

Answer 12

death receptor protein on target cell surface, Fas ligand binds to it (on surface of cytotoxic t cells - holds structure together)

Question 13

what happens after Fas ligand binds to Fas?

Answer 13

1. Fas aggregates
2. ASSEMBLY OF DISC: procaspase-8/10 associates with Fas via an adaptor protein (FADD) to form death inducing signalling complex (DISC)
3. ACTIVATION & CLEAVAGE OF PROCASPASE-8/10: procaspases molecules then activate each other and initiate further activation

Question 14

what happens once the procaspase-8/10 is activated?

Answer 14

becomes active caspase-8/10

can activate executioner caspases

Question 15

what are the initiator caspases?

Answer 15

caspase-8 or -10

procaspases have death effector domain - binds to FADD to bind to Fas death receptor to form DISC

Question 16

what happens during/after apoptotic stimulation of mitochondria and how is caspase-9 activated?

Answer 16

cytochrome c released from mitochondria
cyto-c binds to and activates Apaf1 adaptor protein
ASSEMBLY OF APOPTOSOME - 7 Apaf1 structure - connected by CARD domain in ring
procaspase-9 then binds to apoptosome via its own CARD domain and becomes activated - caspase-9 (cleavage)

Question 17

what are the Bcl2 family proteins?

Answer 17

‘B cell lymphoma’ - proteins function is to control RELEASE OF CYTOCHROME C
can be pro-apoptotic and anti-apoptotic proteins - balance determines initiation of apoptosis

Question 18

what are Bak and Bax?

Answer 18

pro-apoptotic proteins
Bak = normally in membrane
Bax = translocates from cytosol into membrane when activated
form pores in outer mitochondrial membrane to allow protein channel formation or alter membrane structure

Question 19

induction of cytochrome c release

Answer 19

apoptotic stimuli cause BH3-only proteins to be produced or activated
they bind to and inhibit anti-apoptotic Bcl2 proteins
allow Bak and Bax to form pores - in outer mitochondrial membrane

Question 20

what is perforin?

Answer 20

secreted from cytotoxic T cells to form pores in membrane of target cell
protease GRANZYME B enters target cell activating Bid by proteolysis - forms tBid = causes caspase cascade

Question 21

how does tBid activate apoptosis?

Answer 21

tBid activates Bax causing cytochrome c release

Question 22

how is apoptosis prevented?

Answer 22

external signals: survival factors - e.g. IGF1
SF bind to receptors in membrane
activation of transcription of anti-apoptotic Bcl2 genes
inactivation of pro-apoptotic BH-3 proteins by phosphorylation, active Akt kinase

Question 23

role of p53?

Answer 23

DNA damage = p53 activates transcription of gene for Fas - turns on apoptosis
activates transcription of pro-apoptotic protein genes (PUMA/NOXA)