III. Signal transduction and cell cycle | 54. Members of the Bcl-2 superfamily and their roles in various apoptotic pathways, the "survival signal"

Question 1

I. Basics
1. What are the definition and function the Bcl-2 family of proteins?

Answer 1

• The Bcl-2 family of proteins is a major class of regulators of the intrinsic pathway of apoptosis.
• They regulate the intrinsic pathway by controlling the release of cytochrome c and other mitochondrial proteins into the cytosol.

Question 2

I. Basics
2. What are different classes of Bcl-2 families?

Answer 2

We have different classes of Bcl-2 families:
- Pro-apoptotic multidomain Bcl-2 proteins: promote apoptosis by enhancing the release of cytochrome c (Bak, Bax)
- Anti-apoptotic multidomain Bcl-2 proteins: inhibit apoptosis by blocking the release of cytochrome c (Bcl-2, Bcl-XL, MCL-1)
- Pro-apoptotic ‘’BH3 only’’ Bcl-2 proteins: (Bad, Bid, Bim, Noxa, Puma)

Question 3

I. Basics
3. What is the role of Pro-apoptotic multidomain Bcl-2 proteins?

Answer 3

promote apoptosis by enhancing the release of cytochrome c (Bak, Bax)

Question 4

I. Basics
4. What is the role of anti-apoptotic multidomain Bcl-2 proteins?

Answer 4

They inhibit apoptosis by blocking the release of cytochrome c (Bcl-2, Bcl-XL, MCL-1)

Question 5

I. Basics
5. What is the role of Pro-apoptotic ‘’BH3 only’’ Bcl-2 proteins?

Answer 5

Pro-apoptotic ‘’BH3 only’’ Bcl-2 proteins: (Bad, Bid, Bim, Noxa, Puma)

Question 6

I. Basics
7. How can proteins in the Bcl-2 family bind to each other?

Answer 6

• The pro-apoptotic and anti-apoptotic proteins can bind to each other in various combinations to form heterodimers, in which the two proteins inhibit each other’s function.
• The balance between activities of pro- and anti-apoptotic proteins largely determines whether a cell lives or dies by the intrinsic pathway of apoptosis.

Question 7

II. Bak and Bax (pro-apoptotic proteins)
1. What are Bak and Bax (pro-apoptotic proteins)?

Answer 7

Bak and Bax are the main effectors of the Bcl-2 family, forming a pore when
activated and the release of cytochrome C + other mitochondrial proteins

Question 8

II. Bak and Bax (pro-apoptotic proteins)
3. What is the mechanism of Bak and Bax (pro-apoptotic proteins)?

Answer 8

• Activation of Bak/Bax depends on the activated ‘’BH3 only’’ proteins, which are induced by apoptotic signals
• The ‘’BH3 only’’ proteins are pro-apoptotic proteins which will inhibit the anti-apoptotic Bcl-2 proteins, which usually inhibit Bak/Bax => Bak/Bax will be activated
• Bak is bound to the outer mitochondrial membrane, while Bax is mainly located in the cytosol, but translocated to the mitochondria after it is activated

Question 9

II. Bak and Bax (pro-apoptotic proteins)
4. What is the role of the ‘’BH3 only’’ proteins in case of Bak and Bax (pro-apoptotic proteins)?

Answer 9

The ‘’BH3 only’’ proteins are pro-apoptotic proteins which will inhibit the anti- apoptotic Bcl-2 proteins, which usually inhibit Bak/Bax
=> Bak/Bax will be activated

Question 10

III. Bcl-2 and Bcl-XL
1. What are Bcl-2 and Bcl-XL?

Answer 10

Bcl-2 and Bcl-XL are anti-
apoptotic proteins located in the cytosolic surface of the outer mitochondrial membrane

Question 11

III. Bcl-2 and Bcl-XL
2. What is the role of the ‘’BH3 only’’ proteins in case of Bcl-2 and Bcl-XL?

Answer 11

The ‘’BH3-only’’ proteins are the largest subclass of Bcl-2 family proteins. They are activated in response to apoptotic stimulus, and promote apoptosis by inactivating the anti-apoptotic proteins:
- Bad activated by lack of survival signals
- Bid activated by binding of death ligands
- Noxa, Puma (and Bid) are induced by p53 in response to DNA damage, stress, oncogenic insult
- Bim activated by ER stress

Question 12

IV. Bid (BH3-interacting-domain death agonist)
1. What is the role of Bid?

Answer 12

Be a part of apoptosis or caspase-independent pathway

Question 13

IV. Bid (BH3-interacting-domain death agonist)
2. What is the mechanism of Bid in case of apoptosis?

Answer 13

Bid is normally inactive, but when death receptors activate the initiator caspase 8 via the DISC, caspase 8 can cleave Bid, producing an active form (tBid) which is translocated to mitochondria and inhibits anti-apoptotic proteins
=> cytochrome c release
=> activation of intrinsic + extrinsic pathway
=> apoptosis

Question 14

IV. Bid (BH3-interacting-domain death agonist)
3. What is the mechanism of Bid in case Bid is a part of the caspase-independent pathway?

Answer 14

• Bid is also a part of the caspase-independent pathway, where the cell ‘’refuses’’ to undergo apoptosis when it should.
• A cytotoxic lymphocyte can insert granzyme B into the cell and it will active Bid
  -> activate intrinsic apoptotic pathway
  -> induce formation of apoptosome
  -> activate caspases

Question 15

V. Role of p53
1. How does p53 operate?

Answer 15

Bak and Bax: the expression of Bak/Bax is regulated by p53, and is therefore involved in the p53-mediated apoptosis. p53 interacts with Bak/Bax, promoting its activation as well as its insertion into the mitochondrial membrane

Bid, Puma, Noxa: p53 activates these proteins, leading to the inhibition of Bcl-2. Bcl-2 is thereby not able to inhibit Bax, leading to cytochrome c release and apoptosis
=> p53 is therefore pro-apoptotic, leading to apoptosis when it is induced by e.g. DNA damage

Question 16

V. Role of p53
2. What is the role of p53 in case of Bak and Max?

Answer 16

Bak and Bax: the expression of Bak/Bax is regulated by p53, and is therefore involved in the p53-mediated apoptosis. p53 interacts with Bak/Bax, promoting its activation as well as its insertion into the mitochondrial membrane

Question 17

V. Role of p53
3. What is the role of p53 in case of Bid, Puma, Noxa?

Answer 17

Bid, Puma, Noxa: p53 activates these proteins, leading to the inhibition of Bcl-2. Bcl-2 is thereby not able to inhibit Bax, leading to cytochrome c release and apoptosis

Question 18

V. Role of p53
4. What are the features of p53?

Answer 18

p53 is pro-apoptotic, leading to apoptosis when it is induced by e.g. DNA damage

Question 19

VI. Survival signals
1. What are survival signals?

Answer 19

Survival signals are extracellular signal molecules that inhibit apoptosis. The default choice of the cell is to die, therefore, most animal cells require continuous signaling from other cells to avoid apoptosis.

Question 20

VI. Survival signals
2. What is the role of Survival signals?

Answer 20

Survival signals usually bind to cell-surface receptors, which activate intracellular signaling pathways that suppress the apoptotic program, often by regulating members of the Bcl-2 protein family.

Question 21

VI. Survival signals
3. What are the 3 examples of apoptosis inhibition by survival signals?

Answer 21

1) Some survival factors suppress apoptosis by stimulating the transcription of genes that encode antiapoptotic Bcl2 family proteins such as Bcl-2 itself or Bcl-XL.

2) Activation of serine/threonine protein kinase Akt, which, among many other targets, phosphorylates and inactivates the pro-apoptotic BH3-only protein Bad. When not phosphorylated, Bad promotes apoptosis by binding to and inhibiting Bcl2; once phosphorylated, Bad dissociates, freeing Bcl2 to suppress apoptosis.

3) Via the MAPK pathway, stimulating the phosphorylation of the anti IAP protein Hid. When not phosphorylated, Hid promotes cell death by inhibiting IAPs. Once phosphorylated, Hid no longer inhibits IAPs, which become active and block apoptosis.

Question 22

VI. Survival signals
3A. 1 of the Examples for apoptosis inhibition by survival signals is that some survival factors suppress apoptosis
=> Explain

Answer 22

Some survival factors suppress apoptosis by stimulating the transcription of genes
that encode antiapoptotic Bcl2 family proteins such as Bcl-2 itself or Bcl-XL.

Question 23

VI. Survival signals
3B. 1 of the Examples for apoptosis inhibition by survival signals is
“ Activation of serine/threonine protein kinase Akt”
=> Explain

Answer 23

• Activation of serine/threonine protein kinase Akt, which, among many other targets, phosphorylates and inactivates the pro-apoptotic BH3-only protein Bad.
• When not phosphorylated, Bad promotes apoptosis by binding to and inhibiting Bcl2; once phosphorylated, Bad dissociates, freeing Bcl2 to suppress apoptosis.

Question 24

VI. Survival signals
3C. 1 of the Examples for apoptosis inhibition by survival signals is
“Via the MAPK pathway”
=> Explain

Answer 24

• Via the MAPK pathway, stimulating the phosphorylation of the anti IAP protein Hid.
• When not phosphorylated, Hid promotes cell death by inhibiting IAPs.
• Once phosphorylated, Hid no longer inhibits IAPs, which become active and block apoptosis.