Apoptosis Flashcards

Question

Describe the formation of the apoptosome.

Answer 1

The **apoptosome** *('wheel of death')* is made up of: * APAF-1 *(apoptotic activating factor-1)* * Cyctochrome c * ATP * Procaspase 9 APAF-1 structure * CARD domain *(at its N-terminus)* * CARD = caspase recruiting domain * ATPase domain * WD-40 repeats * *Made up of repeats of a sequence of approx. 40 AAs* * Mediate protein-protein interactions Apoptosome formation: * When cytochrome c binds to the WD-40 repeats of APAF-1 , the APAF-1 proteins come together to form a **heptamer** * ****Also requires ATP *(binding to the APAF-1)* * The CARD domains of the heptamer can then interact with **procaspase 9** molecules via their CARD domains * So each Apaf-1 in the heptamer can potentially bind a procaspase 9 * Interaction between the APAF-1 heptamer and procaspase 9 molecules → procaspase 9 oligomerisation * Oligomerisation → transcleavage of procaspase 9 → active tertramer released *(2LS + 2SS)*

Answer 2

Initiate a caspase cascade *(proteolytic cleavage cascade)*

Answer 3

**Energy levels** in the cell determines whether it will undergo apoptosis OR necrosis * Apoptosis requires energy because the **apoptosome requires ATP**

Answer 4

* Bid links the receptor-mediated and mitochondrial death pathways * Caspase 8 from the receptor-mediated pathway cleaves Bid, activating it * BID enhances release of mitochondrial proteins (including **cytochrome c**), thus engaging the intrinsic pathway * The major difference between the intrinsic and extrinsic is that the **intrinsic pathway requires ATP** * So, whichever pathway is stimulated initially, the intrinsic pathway is involved in apoptosis * Therefore, **apoptosis** is an **active process -** *requires ATP*

Answer 5

These are **intrinsic modulators of apoptosis** *(i.e. modulate the intrinsic pathway)* There are 3 main groups of proteins in the Bsl-2 family * They **all** contain a **BH3** domain * BH3 is a **dimerisation motif** * Allows proteins in the Bcl-2 family to associate and dimerise with each other * Some of the proteins contain other domains including a transmembrane domain * The transmembrane domain attaches these proteins to the outer mitochondrial membrane * Group 3 is the BH3 only family * *Can have a transmembrane domain but no other BH domains*

Answer 6

**Anti-apoptotic** * These include: * Bcl-2 * Bcl-xL * They _inhibit_ apoptosis * They are localised to the mitochondria * *Specifically the outer mitochondrial membrane* **Pro-apoptotic** * These include: * Bid * Bad * Bax * Bak * They _promote_ apoptosis * They move between the cytosol and mitochondria * *Specifically the outer mitochondrial membrane*

Answer 7

**Pathway 1** * Growth factor *(ligand)* binds to growth factor receptor *(tyrosine kinase receptor)* * This leads to receptor dimerisation → cross-phosphorylation → docking of Grb2 *(adaptor protein)* → activates Ras → ERK cascade → cell growth (and proliferation) **Pathway 2** * Growth factor *(ligand)* binds to growth factor receptor *(tyrosine kinase receptor)* * This leads to receptor dimerisation → cross-phosphorylation → docking of a **different** protein: **PI3-K** → activates PI3-K pathway → cell survival *(i.e. anti-apoptotic effects)* and proliferation **

Answer 8

PI3-K = phosphatidylinositol 3-kinase * A **lipid** kinase (NOT a protein kinase) * *i.e. phosphorylates lipids not proteins* * Involved in: * Control of cell growth * Cell survival Pathway: * PI3-K binds to the receptor → PI3-K activated * Once activated PI3-K phosphorylates **PIP₂ → PIP₃** * PIP₂ is a phospholipid present in the plasma membrane * PIP₃ activates **protein kinase B (PKB)** * ****PKB is also known as **Akt** * PKB is **anti-apoptotic** IMPORTANT: * PI3-K has 2 subunits * p85 - adaptor * p110 - kinase *(i.e. catalytic)* * PI3-K binds to **phosphorylated tyrosines** on the tyrosine kinase receptor via its **adaptor subunit** * *p85 subunit has SH2 domains which allow this* * This activates the **kinase subunit** which catalyses **PIP₂ → PIP₃**

Answer 9

* Phosphorylates and inactivates **Bad** * Phosphorylates and inactivates **caspase 9** * Inactivates **FOXO** transcription factors * FOXOs promote expression of apoptosis-promoting genes * *Promotes the intrinsic and extrinsic pathway* * Other - e.g. stimulates ribosome production and protein synthesis * *Impaired ribosome biogenesis → p53-mediated apoptosis* * *So stimulating ribosome production prevents this* * *Also, in general, you need ribsome production to allow protein synthesis* * *And you need sufficient protein synthesis, so the cell has enough resources (e.g. enzymes) to carry out its normal function and survive*

Answer 10

Cell survival *(picture A)* * The anti-apoptotic proteins *(Bcl-2. Bcl-xL)* form a **heterodimer** with the the pro-apoptotic proteins *(Bax, Bak)* * Dimerisation via **BH3 domains** * *Both pro- and anti-apoptotic proteins have BH3 domains* * **Heterodimer on outer mitochondrial membrane * **PKB** phosphorylates **Bad** *(pro-apoptotic)* * This results in Bad forming a heterodimer with the protein 14-3-3 * REMEMBER: This requires presence of a **growth factor** stimulating the PI3-K pathway * The pro-apoptotic proteins are held **inactive** in their heterodimers Apoptosis *(picture B)* * Growth factor **absent →** no PI3-K pathway → PKB does **not** phosphorylate Bad → **Bad is active** * Bad binds to Bcl-2 and Bcl-xL, displacing the pro-apoptotic proteins (Bax, Bak) * *i.e. Bax and Bak released from their heterodimer → **active*** * Bax and Bak then form a pore in the outer mitochondrial membrane → cytochrome C released from mitochondria * *Bax and Bak oligomerise to form a pore complex* * **Once cytochrome C is in the cytosol, it can stimulate the formation of the apoptosome → apoptosis

Answer 11

* It's a lipid kinase which dephosphorylates PIP₃ back to PIP₂ * This turns off activation of PKB and the PI3-K pathway * This pathway promotes cell survival and protein synthesis *(and proliferation)* * So by turning off this pathway, it **promotes apoptosis** * By turning off a pathway that promotes cell survival and proliferation, PTEN acts as a **tumour suppressor** * *You need this **negative regulation** to prevent the cell from dividing uncontrollably*

Answer 12

IAP = **I**nhibitor of **A**poptosis **P**roteins IAPs inhibit *(regulate)* the **extrinsic** pathway of apoptosis by: * Binding to procaspases and preventing activation * Binding to active caspases and inhibiting their activity

Answer 13

NOTE: Inhibiting the apoptotic pathways is **cytoprotective** because it protects the cell from death **Intrinsic pathway inhibition:** * Bcl-2 * Bcl-xL **Extrinsic pathway inhibition:** * FLIP * IAPs **Both pathways:** * Growth factor pathways via PI3’-K and PKB/Akt

Answer 14

**Proto-oncogenes:** * Bcl-2 * PKG/Akt EXPLANATION (extra): * These are anti-apoptotic genes → promote cell survival * So, if these genes mutated, you get cell survival even when apoptosis should occur *(e.g. DNA is damaged)* * Cell survival allows cell proliferation, which could lead to tumour formation **Tumour suppressor:** * PTEN * *It inhibits the PI3-K pathway which promotes cell survival and proliferation* * *This negative regulation prevents the cell from dividing uncontrollably*

Answer 15

* Stimulating apoptosis in harmful cells * e.g. Cells with viral infection or DNA damage * These cells can be oncogenic *(can potentially cause cancer)* * Chemotherapeutic killing of tumour cells * e.g. Dexamethasone stimulates DNA cleavage * *DNA fragmentation is a key feature of apoptosis*

Apoptosis Flashcards

(39 cards)