Topic 11: Apoptosis

Question 1

What is the purpose of apoptosis?

Answer 1

remove diseased or damaged tissue

allows tissue remodeling and maintenance

must be tightly controlled

Question 2

What are some general characteristics of apoptosis?

Answer 2

nucleus is dismantled (nuclear envelope vesiculates, nucleus shrinks)

DNA is degraded by DNAses into ~200bp fragments

cytoplasm shrinks

mitochondria breakdown and release cytochrome c

cells release ATP and UTP into extracellular to recruit macrophages

phagocytic cells secrete inhibitory cytokines to reduce inflammation

cell cytoskeleton dismantles (causes “blebbing”)

cell vesiculates (gets dismantled into vesicles)

Question 3

What are the three main steps of apoptosis?

Answer 3

as a cell begins to undergo apoptosis, its chromosomes condense and its cytoplasm shrinks

eventually, the nucleus becomes fragmented, its DNA is digested at regular intervals (“laddering”), the cytoplasm becomes fragmented, and the cell extends numerous blobs

ultimately the remnants of the dead cell (apoptotic bodies) are ingested by phagocytic cells

Question 4

What is phosphatidylserine (PS)?

Answer 4

normally on intracellular leaflet

PS flips to extracellular face and acts as an “eat me” signal

Question 5

What is the balance of signals in controlling apoptosis?

Answer 5

the balance between loss of survival signals and reception of death signals (intracellular and extracellular)

Question 6

How does the withdrawal of positive signals control apoptosis?

Answer 6

the continued survival of most cells requires that they receive continuous stimulation from other cells and, for many, continued adhesion to the surface on which they are growing

example: anchorage dependent growth (signals via integrins
they tell the cell it is healthy and to stay alive, therefore loss of this signal should trigger apoptosis

Question 7

How does the receipt of negative signals control apoptosis?

Answer 7

internal signal: DNA damage or oxidative stress

external signal: FasL and TNF (tumor necrosis signal) trigger apoptosis via caspase 8

Question 8

What is TNF?

Answer 8

TNF produced during inflammation can trigger TNFR

Question 9

What is FasL?

Answer 9

FasL (Fas ligand) binds Fas receptors and triggers apoptosis

Question 10

What are the three different mechanisms by which a cell commits suicide by apoptosis?

Answer 10

1. signals arising from within the cell
2. oxidative damage (cause accumulation of reactive oxygen species, ROS = do widescale cell damage)
3. receipt of death signals

Question 11

What are caspases?

Answer 11

a family of over a dozen caspases

they are all proteases

they get their name because they cleave proteins, mostly each other, at aspartic acid (Asp) residues

Question 12

What is Caspase 9?

Answer 12

protease that cleaves over caspases

caspases are produced as inactive precursors (pro-proteins) that require proteolytic cleavage for activation

called the initiator caspase

Question 13

What is Caspase 3?

Answer 13

called the effector caspase

activated by caspase 9

cleaves many downstream targets: rMLC (causes cell blebbing), GRASP6S (induce Golgi fragmentation), MST1 (promotes nuclear fragmentation), and others

cleaves adhesion proteins involved in cell-cell junctions

Question 14

What is the role of the mitochondria in apoptosis?

Answer 14

highly metabolic, involves reactions involving oxygen, can create toxic oxygen species

stores cytochrome C (component of ETC), cyt. c is also a signaling molecule that activates caspase 9

Question 15

What is apoptosis triggered by internal signals, the intrinsic or mitochondrial pathway?

Answer 15

in a healthy cell, the outer membranes of its mitochondria display the protein Bcl-2 on their surface, Bcl-2 inhibits apoptosis

Bcl-2 mediated internal damage to the cell

cell stress (from mitochondria and p53 that monitors genome health) can trigger downstream response

sensors of cell stress –| Bcl2 –| Bax –> cyt C

Bax is in mitochondria and responds to the loss of inhibition by Bcl-2, inserts into MOM allowing release of cyt C

Apaf-1: when activated by cyt C, polymerizes into apoptosome, “wheel of death”, apoptosome cleaves and activates caspase 9

Question 16

What is apoptosis triggered by external signals, the extrinsic or death receptor pathway?

Answer 16

FasL and TNF –> death signals

receptors for death ligands (FasLR and TNFR) are constitutively expressed in all cells

binding of death signal activates caspase 8

active caspase 8 –> target caspase 3

active caspase 3 –> apoptosis proceeds in the same manner as intrinsic

Question 17

What are the steps in the receipt of death signals pathway to apoptosis?

Answer 17

cytotoxic T cell produces death signals

FasL binds to FasLR

activates procaspase 8 which cleaves and activates caspase 3, which triggers apoptosis

Question 18

What are the steps in the loss of survival signals pathway to apoptosis?

Answer 18

as sensors of cell stress increase, Bcl2 is inhibited

since Bcl2 levels decrease, levels of Bax increase

as Bax levels increase so does the level of cyt. C

Apaf-1 is activated by cytc. C and polymerizes into apoptosome (wheel of death)

the apoptosome activates caspase 9, which in turn activates caspase 3, which then causes apoptosis

Question 19

What is the intrinsic pathway to apoptosis?

Answer 19

ROS damage to DNA

this signal is received by p53 (gossip queen), which senses many forms of cell damage

from here, the pathway follows the same steps as the loss of survival signals pathway

Question 20

What is the Apoptosis-Inducing Factor (AIF)?

Answer 20

neurons, and perhaps other cells, have another way to self-destruct that, unlike the two paths described above, does not use caspases

AIF is a protein that is normally located in the intermembrane space of the mitochondria

when the cell receives a signal telling it that is is time to die, AIF is released into cytoplasm

caspase-independent

AID is released from mito and migrates to nucleus (bind and degrade DNA)

Question 21

What is the relationship between apoptosis and cancer?

Answer 21

some viruses associated with cancers use tricks to prevent apoptosis of the cells they have transformed

defects in apoptotic pathways contribute to immortality of cancer cells

Bcl-2 is overexpressed in B cell lymphoma, gain of function = promoted cell survival by enhancing apoptotic inhibition

Question 22

What are the ways cancer cells avoid apoptosis?

Answer 22

upregulating Bcl-2 (oncogene)

downregulating p53 (TSG), 50% of all cancers contain a p53 mutation, most frequent mutation in all cancers

decrease Apaf-1 expression (TSG)

secreting decoy death signals (bind and block TNFR and FasLR to prevent activation of receptor)