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Y2 MCD - Cancer - Laz > Apoptosis > Flashcards

Flashcards in Apoptosis Deck (40)
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1

Define Necrosis.

Unregulated cell death associated with trauma, cellular disruption and an inflammatory response

2

Define Apoptosis.

Regulated cell death; controlled disassembly of cellular contents without disruption – no inflammatory response

3

Describe the process of necrosis.

The plasma membrane becomes more permeable – the cell swells and the membrane ruptures
Proteases are released leading to dissolution and autodigestion of thecell
There is localised inflammation

4

What are the two phases of apoptosis? Describe them.

Latent phase
 Death pathways are activated, but cells appear morphologically the same
Execution phase:
 Loss of microvilli and intercellular junctions
 Cell shrinkage
 Loss of plasma membrane asymmetry
 Chromatin and nuclear condensation
 DNA fragmentation
 Formation of membrane blebs
 Fragmentation into membrane enclose apoptotic bodies (these are then taken up by macrophages)

5

What is an important feature of apoptosis that distinguishes itfrom necrosis?

Plasma membrane remains intact – no inflammation

6

What DNA modification is seen during apoptosis?

Fragmentation of DNA ladders (seen in agarose gel)
Formation of more ends, which are labelled by adding an extra fluorescently-labelled tag in a TUNEL assay

7

What other types of cell death are there other than necrosis and apoptosis?

Apoptosis-like cell death
Necrosis-like cell death (sort of like an aborted apoptosis that ends up being necrosis)
NOTE: cell death is GRADED

8

What are caspases?

Cysteine-dependent aspartate-directed proteases
They are the executioners of apoptosis
They are activated by cleavage

9

Which caspases are effector caspases?

3, 6 and 7

10

Which caspases are initiator caspases?

2, 8, 9 and 10

11

Describe the structure of effector caspases.

They are single chain polypeptides consisting of a small and large subunit
The subunits are released by proteolytic cleavage

12

Describe the structure of initiator caspases.

They have the same two subunits found in effector caspases but they also have a targeting subunit (protein-protein interacting domain)

13

What are the two types of targeting subunit that initiator caspases can have?

CARD – caspase recruitment domain
DED – death effector domain

14

How are active caspases formed?

Cleavage of inactive procaspases is followed by the folding of 2 largeand 2 small chains to form an active L2S2 heterotetramer

15

What are the two mechanisms of apoptosis

Death by design (receptor-mediated)
Death by default (mitochondrial (intrinsic) death pathway)

16

Describe the structure of death receptors.

Cysteine-rich extracellular domain
Transmembrane domain
Intracellular tail with a death domain (DD)

17

What are the two important adaptor proteins in the death by design pathway and how are they different?

FADD – positive regulator that promotes cell death – DED + DD
FLIP – negative regulator – DED + DED

18

Describe signalling of apoptosis through Fas.

 Fas ligand binds to Fas receptor (on CTLs) and the Fas receptors undergo trimerisation, which brings the three DDs together
 The trimerised DDs recruit FADD, which binds via its own DD
 FADD then recruits and oligomerises procaspase 8 through the DED of procaspase 8
 Binding of procaspase 8 to FADD forms DISC (death-induced signalling complex)
 DISC formation results in cross-activation of procaspase 8
 Active caspase 8 is released, which then activates effector caspases

19

Describe the important of oligomerisation in this pathway.

Some initiator caspases have intrinsic low catalytic activity
Oligomerisation brings them close enough together to allow transcleavage
Also, at least 2 procaspases are required to form an active caspase

20

Describe how FLIP acts as an inhibitor of apoptosis

FLIP is evolutionarily related to caspases but has lost its catalytic activity
It has two DED domains and can compete with procaspase 8 to bind to the DED domains of FADD
It can incorporate into receptor-procaspase complexes and interfere with transcleavage

21

As an overview, describe death by default.

Cellular stress causes a change in mitochondrial membrane potential
This leads to release of cytochrome C from the mitochondrion
This stimulates formation of the apoptosome complex

22

What does the apoptosome consist of?

APAF-1 (apoptotic activating factor 1)
Cytochrome C
ATP
Procaspase 9

23

Describe the domains found within APAF-1.

CARD domain
ATPase domain
WD-40 repeats (protein-protein interactions)

24

Explain fully, how death by default leads to caspase activation.

The cytochrome C released from the mitochondria bind to the WD-40 repeats of APAF-1 and make it form a heptamer structure (apoptosome)
This requires ATP
It has 7 CARD domains in the middle, which can interact with CARD domains of procaspase 9
Seven procaspase 9 bind via their CARD domains to the apoptosome and their close contact allows them to cross-cleave each other to generate activate caspase 9

25

What pro-apoptotic protein links the death by default and death by design pathways? Explain how it works.

Bid
Caspase 8 (generated by the death by design pathway) cleaves Bid, which travels to the mitochondrion and promotes the release of cytochrome C – thus triggering the mitochondrial death pathway

26

How can energy levels of a cell show whether a cell is going through apoptosis or necrosis?

Apoptosis requires energy whereas necrosis does not

27

What is an important family of proteins that act as intrinsicmodulators of apoptosis?

Bcl-2 family

28

There are three main groups of Bcl-2 proteins. What is common toall three groups?

BH3 domain – this is a dimerisation motif, which allows members of the family to form dimers with each other

29

What are the anti-apoptotic Bcl-2 proteins and where are they found?

Bcl-2
Bcl-xL
They are found localised on the mitochondrial membrane

30

What are the pro-apoptotic Bcl-2 proteins and where are they found?

Bid
Bad
Bax
Bak
These are found in the cytoplasm and in the mitochondrial membrane