Apoptosis

Question 1

What is the function of apoptosis

Answer 1

removes damaged or dysfunctional cells through controlled, programmed cell death (cell suicide)

Question 2

What is necrosis

Answer 2

Uncontrolled cell death – cell homicide

Question 3

Is necorsis pathological or physiological

Answer 3

Is a pathological process when compared to apoptosis which is physiological

Question 4

What does necrosis involve including cell death

Answer 4

Involves lots of cells, inflammation, acidosis and liquefaction

Question 5

How does necrosis affect other cells in the environment

Answer 5

Cell contents released which effect normal tissue – this causes secondary tissue damage

Question 6

What is the Hayflick limit

Answer 6

The capacity for a cell to divide – is limited

Dictated by the telomeres on chromosomes – these shorten by 60-80 bp per cell cycle

Question 7

What does the Hayflick limit cause when exceeded

Answer 7

Genomic instability which results in apoptosis

Question 8

What is telomerase

Answer 8

nzyme that maintains the telomeres – present in germ cells (like stem cells)

Is activated in certain diseases like cancer (cancer cells can overcome the Hayflick limit)

Question 9

What mediates apoptosis

Answer 9

The caspases

Question 10

What does the term caspase stand for

Answer 10

Cysteinyl-aspartic-specific proteases (caspases)

Question 11

What processes are caspases involved in

Answer 11

DNA fragmentation (apoptosis)
Cell shrinking (apoptosis)
T cell proliferation
Tumour metastasis
Tumour suppression
Cell migration
Inflammatory response

Question 12

What residue is important in the active site of caspases

Answer 12

Cysteine

Question 13

What do caspase enzymes do to aspartic residues

Answer 13

They cleave the peptide bond on the C terminal side of an aspartate residue on their target (targets differ)

Question 14

Why can caspase enzymes be found in a “pro” form

Answer 14

As caspase play a critical role in cell survival, they must be in an inactive “pro” form

Question 15

How many domains does pro-caspase have

Answer 15

3 or 4 domains

Question 16

What happens to the pro domain in pro-caspase

Answer 16

Pro domain is cleaved off and released – large and small subunits which come together (heterotetramer) —> functional caspase

Question 17

Where is cytochrome-c found and what does it do

Answer 17

Cytochrome-c sits between complex-III and complex-IV on the inner membrane surface of the mitochondria

Shuttles electrons from the complex-III and complex-IV during electron transport

Question 18

What happens to mitochondria membrane integrity during apoptosis

Answer 18

Mitochondria membrane integrity is compromised

Question 19

What happens as a result of mitochondria membrane integrity being compormised

Answer 19

The release of cytochrome-c

This activates pro-caspase-9

Question 20

What other proteins mediate apoptosis that are released from mitochondria

Answer 20

Endonuclease G (DNA fragmentation)

Smac/Diablo

AIF (apoptosis inducing factor)

Question 21

How does BCL-2 protect mitochondria

Answer 21

prevents the release of Cyt-c and Endo-G

Question 22

How does BCL-2 contribute to cancer

Answer 22

BCL-2 is an oncogene so can contribute to cancer if mutated

Question 23

How is the intrinsic pathway of apoptosis driven

Answer 23

Driven by mitochondria and cytochrome-c

Question 24

What factors can cause permeability in mitochondria membrane

Answer 24

DNA damage, topoisomerase inhibition, cytoplasmic stress (ROS etc) among other stimuli cause permeability in the mitochondrial membrane

Question 25

What does DNA damage, topoisomerase inhibition, cytoplasmic stress (ROS etc) cause to be released

Answer 25

This releases Cyt-c, endonucelase G, apoptosis activating factor (AIF)

Question 26

How is mitochondrial permeabilisation mediated

Answer 26

Mitochondrial permeabilization is mediated by the BCL-2 protein family which have pro-apoptotic and anti-apoptotic effects

Question 27

What proteins does BCL-2 inhibit to prevent apoptosis

Answer 27

Bax and Bak

Question 28

How is apoptosis measured using cell markers

Answer 28

Phosphatidyl serine on the outside of cell membrane (detected by FACS) recognition by phagocytic cells

Question 29

What does it mean if FITC (a dye) is present when measuring apoptosis

Answer 29

The cell is apoptotic as phosphatidyl serine has flipped to the outside membrane

Question 30

How is late apoptosis shown

Answer 30

If PI is present then membrane integrity is lost, showing late apoptosis If only PI is present necrosis is indicated as membrane integrity will be lot

Question 31

How is the extrinsic pathway mediated

Answer 31

Involves the activation of receptors at the cell membrane --> signal trandsuction --> apoptosis initiation

Question 32

What receptors are responsible in this pathway

Answer 32

Death receptors are a class of membrane receptors

Question 33

What is an important subgroup of death receptors and what are some members

Answer 33

the tumour necrosis factor (TNF) receptor family Death receptor 3 (DR3) TNF-R1 TNF-related apoptosis inducing ligand receptor 2 (Trail-R2) FAS Programmed death protein 1 (PD1) Apoptosis is triggered by ligand binding to DR at cell surface

Question 34

What is the topology of a death receptor

Answer 34

Has cysteine rich domains which are involved in ligand binding Ligands are usually trimers and associated with the membrane of a neighbouring cell – can be soluble factors too

Question 35

What domain does signal transduction which results in apoptosis occur

Answer 35

The death domain

Question 36

What occurs upon ligand binding in death receptor 3 (DR3)

Answer 36

ligand binding causes receptor trimerization - receptor adaptor proteins also contain death domains

Question 37

What do adaptor proteins invovled in DR signalling include

Answer 37

the Fas-associated protein with death domain (FADD)

Question 38

What protein interaction does the FADD contain

Answer 38

a second protein interaction domain known as the dead effector domain (DED)

Question 39

What does the DED of FADD interact with

Answer 39

The DED of FADD interacts with the DEDs on initiator caspases

Question 40

What is the complex formed known as

Answer 40

The complex formed is known as death induced signalling complex (DISC)

Question 41

What survival advantage do cancer cells possess

Answer 41

induce apoptosis in tumour reactive immune cells

Question 42

How do cancer cells suppress apoptosis themselves (intrinsic pathway)

Answer 42

BCL-2 suppresses the release of Cyt-c from the mitochondria and up-regulation of BCL-2 is associated with pathogensis of many cancers Cancer cells can detect DNA damage and are associated with various stress (oxidative etc) which initiates the intrinsic apoptosis pathway BCL-2 blocks this pathway leading to the survival of the cancer cells

Question 43

What does mTOR stand for

Answer 43

mechanistic target of rapamycin Rapamysin is an anti-fungal agent

Question 44

What is mTOR central to

Answer 44

Overall, mTOR plays a central role in growth – proliferation and cell survival

Question 45

What are the 2 complexes of mTOR

Answer 45

, mTORC1 , mTORC2

Question 46

What does autophagy refer to and what does it allow for

Answer 46

Refers to "self-eating" - allows degradation and recycling of cellular components

Question 47

What is apoptosis associated with forming

Answer 47

This is associated with the formation of autophagosomes (packaging the cell --> protects the release of tissue damaging proteases etc --> necrosis)

Question 48

How does apoptosis and autophagy differ to necrosis

Answer 48

Apoptosis (caspase initiated) and autophagy (suppressed by mTOR pathway) mediates cell death This packages the cell and its contents and preserves membrane integrity

Question 49

What is necrosis associated with

Answer 49

cell lysis and the release of intracellular contents Necrosis also mediates inflammation by the activation of macrophages – this can damage healthy tissues