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Molecular Biology of the Cell > Section 4 Lecture 2 Cell Death > Flashcards

Section 4 Lecture 2 Cell Death Flashcards

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1
Q

Bak

A

Pro-apoptotic effector Bcl2 family protein always located on the mitochondrial outer membrane.
When activated by apoptotic stimulus, oligomerizes to release cytochrome C.

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2
Q

Bax

A

Pro-apoptotic effector Bcl2 family protein located in the cytosol and translocates to mitochondrial outer membrane due to apoptotic stimulus to release cytochrome C.

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3
Q

What are the 3 Bcl 2 family proteins?

A
  1. Anti-apoptotic Bcl2 family proteins
  2. Pro-apoptotic effector Bcl2 family proteins
  3. Pro-apoptotic BH3 only proteins
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4
Q

Examples of pro-apoptotic effector Bcl2 proteins

A

Bak and Bax

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5
Q

Examples of anti-apoptotic Bcl-2 family proteins

A

Bcl2 and BclXL

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6
Q

Examples of pro-apoptotic BH3 only proteins

A

Bad, Bim, Bid, Puma, and Noxa

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7
Q

Bcl2 family proteins regulate which pathway?

A

Intrinsic pathway - stimulate or inhibit cytochrome C release from mitochondria

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8
Q

What are the 2 types of cell death?

A

Necrosis and Apoptosis

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9
Q

What are the differences between necrosis and apoptosis?

A 
Apoptosis: 
programmed cell death
cell breaks into apoptotic bodies
No inflammation
phagocytosis

Necrosis: 
Accidental cell death
Cellular Swelling
Pro-inflammatory
Enzymatic digestion and leakage of cellular contents
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10
Q

PS

A

Phosphatidylserine - on the surface of apoptotic cells

Interact with microphages to produce phagocytosis

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11
Q

Blebbing

A

hallmark of apoptosis

bulges of the plasma membrane that is filled with cytosol

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12
Q

Markers of Apoptosis

A
  1. Flipping of Phosphatidylserine - PS flips from the inner lining of the plasma membrane to the outer membrane > determined using GFP-Annexin V which recognizes PS on the outside of the cell and can be seen via microscopy which cells are committed to apoptosis.
  2. DNA fragmentation > determined via Agarose Gel
  3. Release of cyt C from mito to cytosol > determined with Cyt C-GFP visualization of where it is located
  4. Cleaved caspases and targets > Caspase 3 and PARP fragments determined by Western Blot
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13
Q

Annexin V

A

recognizes PS on the outside of the cell and binds to it

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14
Q

2 types of Caspases

A
  1. Initiator

2. Executioner

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15
Q

Caspases are…

A

proteases with cysteine at the active site and cleave targets at aspartic acids (C = cysteine; asp= aspartic acid; ases= proteases)

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16
Q

Examples of Initiator caspases

A

Caspases 8 and 9

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17
Q

Examples of Executioner caspases

A

Caspases 3, 6, and 7

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18
Q

What form is an inactive initiator caspase in?

A

Monomer

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19
Q

What form is an inactive executioner caspase in?

A

Dimer

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20
Q

What form is an active executioner caspase in?

A

Tetramer

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21
Q

Caspase Cascade

A

Apoptotic Signal > Adapter proteins assemble > Dimerization, Activation , and Cleavage of initiator procaspases > Rearrangement of protease domain = Active initiator caspase > Activate by Cleaving Executioner caspase from a dimer to a tetramer => Cleavage of multiple substrates => Apoptosis

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22
Q

Targets of caspases

A
  1. Nuclear lamins
  2. iCADs proteins > frees endonucleases > DNA fragmentation
  3. cytoskeleton (microtubules and filaments)
  4. cell-to-cell adhesion proteins
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23
Q

Caspase Cascade Activation Pathways

Study These Flashcards
A
  1. Extrinsic Pathway

2. Intrinsic Pathway

24
Q

Extrinsic Pathway

Study These Flashcards
A

Triggered by cell surface receptors receiving an apoptotic signal

25
Intrinsic Pathway
Activated by release of Cytochrome C from mitochondria into cytosol
26
Parts of the Extrinsic Pathway
1. TNF family (i.e. TNF and Fas ligands) extracellular signal are received by cell surface death receptors. 2. Fas receptor recruits FADD adapter protein through death domain 3. FADD recruits initiator procaspases through death effector domain 4. DISC complex formed 5. Activating the interior caspases through induced proximity 6. Caspase Cascade activation 7. Apoptosis
27
DISC
Death-inducing signaling complex
28
Inhibitors of extrinsic pathway
1. Decoy receptors that lack death domain > does not induce DISC formation 2. FLIP proteins that lack cleavage domain
29
FADD
Fas-associated death domain
30
Cell surface death receptors are...
trimeric
31
3 domains of trimeric death receptors
1. Extracellular ligand- binding domain 2. Single transmembrane domain 3. Intracellular death domain
32
Parts of Intrinsic Pathway
1. Apoptotic stimulus from inside of the cell (i.e. DNA damage) 2. Cytochrome C released from mitochondria 3. Cyt C binds to Apaf 1 4. Apaf 1 structural change that exposes CARD 5. Apoptosome complex forms through exposed CARDs to form wheel-like heptomer 6. CARD recruits initiator Procaspase 9 to each exposed CARD 7. Induced proximity induces proteolytic cleavage of Procaspase 9 8. Activation of executioner caspases 9. Caspase cascade > Apoptosis
33
Apaf 1
Apoptotic protease activating factor 1
34
CARD
Caspase Recruitment Domain
35
Apoptosome
Heptomer complex that forms in the Intrinsic pathway due to exposed CARD on the Apaf 1 - Cyt C complex Recruits Procaspase 9
36
Bcl 2 proteins
Regulate the intrinsic pathway | Stimulate or inhibit release of cyt C from mito
37
3 types of Bcl 2 proteins
1. Anti-apoptotic Bcl 2 2. Pro-apoptotic effector Bcl 2 3. Pro-apoptotic BH3-only protein
38
Bax
Pro-apoptotic effector Bcl 2 | In cytosol, oligomerizes in the outer mitochondria membrane upon apoptotic stimulus
39
Bcl 2 and BclXL
Examples of Anti-apoptotic Bcl 2
40
Largest Bcl 2 subfamily
Pro-apoptotic BH3-only proteins
41
Bad
Pro-apoptotic BH3-only protein Binds to anti-apoptotic Bcl 2 survival factor to inhibit it and allow apoptosis to take place. Akt pathway phosphorylates Bad and deactivates it so that Bcl 2 can prevent apoptosis
42
Bim, Puma, Noxa, Bid, and Bad
Pro-apoptotic BH3-only proteins
43
Bid
Pro-apoptotic BH3-only protein that links the extrinsic to the intrinsic pathway to amplify the apoptotic signal 1. Initiator caspase 8 of extrinsic pathway cleaves cytosolic Bid 2. tBid (truncated Bid) goes to mito and inhibits Bcl 2 3. Bax and/ or Bak are allowed to oligomerize and release cyt C which will lead to apoptosis
44
Which Bcl 2 BH domain in shared by all?
BH3
45
Bac
Pro-apoptotic effector Bcl 2 Always bound to the other mitochondrial membrane to oligomerize and allow cytochrome C to exit into the cytosol upon apoptotic signal
46
Pro-apoptotic effector Bcl2 proteins
trigger release of cyt. C from mitochondrial intermembrane | i.e Bak and Bax
47
Anti-apoptotic Bcl 2 proteins
inhibit oligomerization Bak and Bax effector proteins | located on cytosolic surface of outer mito membrane
48
Pro-apoptotic BH3-only proteins
inhibit anti-apoptotic Bcl 2 proteins to allow Bax and Bak to function Binds to Bcl 2 and neutralizes activity Produced/ activated from apoptotic signal
49
Apoptotic stimuli to activate BH3-only proteins
1. Deprivation of survival signals > Activate JNK > transcription of Bim 2. Irreparable DNA damage > activates p53 > transcription of Puma and Noxa
50
IAPs
Inhibitors of Apoptosis | Control caspase activity
51
Anti-IAPs
inhibit IAPs = Promote apoptosis | released by mitochondria with cyt C to promote intrinsic pathway
52
Extracellular survival factors
Inhibit apoptosis usually cell-to-cell signals in tissue cells i.e. nervous system requires continuous signaling to avoid apoptosis ensures cell survives on when and where needed
53
Nervous system cell survival
Nerve cells produced in excess. Apoptosis regulated by abundance of survival factor that is released by target cells. If there are too many nerve cells they will not receive enough survival factor and undergo apoptosis
54
3 Mechanisms of Action for Survival Factors
1. Increased production of anti-apoptotic Bcl2 family protein through transcription 2. Inactivation of pro-apoptotic BH3-only protein (Akt -Bad pathway) 3. Inactivation of anti-IAPs (Hid deactivation through MAP Kinase phosphorylation of Hid > IAPs released to promote survival)
55
Cancer therapies targeting apoptosis
1. Target Bcl 2 > activate intrinsic pathway 2. Pro-apoptotic receptor peptides = ligands to activate apoptosis (i.e. Fas and TNF) 3. Target IAPs
56
6 Hallmarks of Cancer
1. Evade apoptosis 2. Self sufficient growth signals 2. Insensitivity of antigrowth signals 4. Sustained angiogenesis 5. Limitless replicative potential 6. Tissue invasion and metastasis

Molecular Biology of the Cell (2 decks)

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