005 Hydrophilic Hormones and Enzymatic Cascades

Question 1

Which pathway does Gs alpha subunit activate?

Answer 1

adenylate cyclase –> increases cAMP

Question 2

Which pathway does Gi alpha subunit activate?

Answer 2

Inhibits adenylate cyclase –> decreases cAMP

Question 3

Which pathway does Gt alpha subunit activate?

Answer 3

activates retinal cyclic GMP phosphodiesterase

Question 4

Which pathway does Gq alpha subunit activate?

Answer 4

activates PLC –> increases DAG, IP3 –> increases Ca2+

Question 5

Describe the cAMP-PKA pathway that is activated by adrenaline

Answer 5

1. Adrenaline binds to Gs protein
2. Conformational change in 3,4,5,6 alpha helices
3. GDP switches for GTP
4. alpha subunit dissociates from BY subunit
5. alpha subunit activates adenyl cyclase, converting ATP to cAMP.
6. 4 x cAMP binds to regulatory region of PKA
7. This causes dissociation of regulatory region from catalytic region.
8. Catalytic unit of PKA phosphorylates other proteins.

Question 6

Describe the structural changes of switch II during activation of the G protein

Answer 6

1. Switch II is hidden by BY subunits
2. Extra phosphate from GTP on alpha subiuntis stabilises helical switch II
3. Switch II activates adenylyl cyclase
4. Hydrolysis of GTP causes switch iI to become disordered again.

Question 7

How many alpha helices does each G protein have on the membrane?

Answer 7

7

Question 8

How many isoforms does G alpha, G beta and G gamma have?

Answer 8

G alpha: 4
G beta: 5
G gamma: 6

Question 9

How does cholera toxin disrupt the G protein?

Answer 9

Prevents hydrolysis of GTP causing continuous activity –> affects osmosis.

Question 10

Describe the mechanism of epidermal growth factor receptor activation

Answer 10

1. EGF binds to EGFR, causing dimerisation
2. X-phosphorylation of cytosolic PTK sites.
3. PTK phosphates causes Grb recruitment and Sos attached to Grb.
4. Sos causes exchange of GDP with GTP on Ras G monomeric protein.
5. GTP-Ras binds with Raf, which is a membrane bound protein kinase.
6. GTP-Ras-Raf causes cascade of phosphorylation in the MAPK pathway.

Question 11

How does cetuximab work?

Answer 11

Prevents the dimerisation of EGFR receptors, preventing the cascade of growth and division to occur. This is used in colerectal cancers.

Question 12

Describe the mechanism of insulin receptor activation

Answer 12

1. Insulin binds to dimeric receptor causing X-phosphorylation
2. Recruits IRS-1, which is bound to PI3K
3. Presence of PI3K at the membrane causes phosphorylation of PIP2 to PIP3.
4. PIP3 allows both PDK1 and PKB to associate with membrane via PH domains.
5. Phosphorylated PKB dissociates from membrane and causes phosphorylation of the target proteins.

Question 13

Describe the mechanism of transforming growth receptor beta activation?

Answer 13

1. TGF-Beta binds to TBRII (which has constitutive serine/threonine activity)
2. TBRII binds to TBR-1 and phosphorylates target site glycine-serine rich domain.
3. This activates TBR-I serine/threonine activaty
4. TBR-I phosphorylate sR-Smads
5. Two R-Smads and Co-Smad form heterotrimer.
6. Nuclear localisation signals exposed, interacting with transcription factors.

Question 14

What are some roles of TGF beta?

Answer 14

Inhibiting cell proliferation
Cell differentiation
Tissue organising - promoting expression of ECM

Question 15

Describe the mechanism of cytokine receptor activation e.g. erythropoiten

Answer 15

1. Epo simultaenously binds to two EpoR
2. JAK kinases in cytosolic domain are close enough to cross phosphorylate (lowering Km)
3. Phosphorylated JAK kinases allow STAT5 to bind via SH2 domains.
4. Phosphorylation of STAT5 causes dissociation from receptor.
5. Dimerisation of STAT5 exposes nuclear localisation sequence.

Question 16

Which molecule can switch off the JAK/STAT pathway and describe how it works?

Answer 16

SHP-1 - phosphotyrosine phosphatase that binds phosphorylated receptors. These dephosphorylates JAK kinases, inhibiting pathway when cytokines are no longer binding the receptor.

Question 17

What happens in mutation of TBR-1 or TBR-II in TGFbeta receptor?

Answer 17

Mutations of this will cause disruption in the TGF-beta effects, causing resistance to growth inhibition by TGFBeta.

Question 18

What cancer is mutation/deletion of Smad-4 found in, and what physiological effect does this have?

Answer 18

Pancreatic cancer

This disrupts the TGFBeta pathway, preventing inhibition of cell differentiation.

Question 19

What implication of the erythropoietin pathway does mutations in SHP have?

Answer 19

Higher levels of RBC despite lower erythropoietin levels.