S3) Intracellular Signalling Pathways

Question 1

What are G Proteins and what do they do?

Answer 1

• G-protein coupled receptors are a superfamily of receptors with an enormous diversity of cellular functions
• They alter the activities of effectors

Question 2

Describe the structure of G proteins

Answer 2

• Heterotrimeric i.e. consist of three subunits: α, β, γ
• The β and γ subunits bind tightly to each other and function as a single unit

Question 3

Describe the properties of the α-subunit of the G protein

Answer 3

• The α-subunit has a guanine nucleotide binding site
• The α-subunit possesses intrinsic GTPase activity as this site binds GTP and then slowly hydrolyses it to GDP

Question 4

Describe the activity/appearance of the G protein under basal resting conditions

Answer 4

• G-protein is present at the inner face of the plasma membrane in its heterotrimeric form
• GDP is bound to the α subunit

Question 5

Explain the effects of ligand binding on the GPCR

Answer 5

• The ligand binds with the GPCR and activates it
• The activated receptor has a high affinity for the G-protein
• GDP is later released and exchanged for GTP by the α-subunit

Question 6

What role does the GPCR take on when GDP is exchanged for GTP?

Answer 6

The receptor acts as a guanine nucleotide exchange factor (GEF)

Question 7

Describe the affinity of the GPCR for the G protein after the guanine nucleutide exchange

Answer 7

• The binding of GTP to the α-subunit decreases the affinity of the receptor for the G-α and Gβγ subunits
• Thus, both α and βγ subunits are released and can each can interact with effectors

Question 8

Explain how the GPCR cycle is reactivated after subunit interaction with effectors

Answer 8

• Effector interaction is then terminated by α-subunit’s intrinsic GTPase activity which hydrolyses GTP → GDP
• Affinity of the Gα-subunit for the Gβγ-subunit then increases
• Gαβγ hetero-trimer is reformed & awaits reactivation

Question 9

State the structure and function of Gs/Gαs

Answer 9

- Structure: Gs carries the β-adrenoceptor for adrenaline and the enzyme adenylyl cyclase

• Function: stimulates adenylyl cyclase

Question 10

State the structure and function of Gi/Gαi

Answer 10

• Structure: an inhibitory version of G protein for the activation of adenylyl cyclase
• Function: inhibits adenylyl cyclase

Question 11

Outline the mechanism of action of Gq

Answer 11

• Gq proteins preferentially interact with the membrane-bound enzyme phospholipase C
• This causes hydrolysis of a minor plasma membrane phospholipid (PIP₂) and generates 2 messengers: IP₃ and DAG

Question 12

For the Gs, Gi and Gq proteins, compare and contrast the following:

• Endogenous signalling molecule
• Receptor
• Effector
• Physiological response

Answer 12

Question 13

State the structure and function of adenylyl cyclase

Answer 13

• Structure: an integral plasma membrane protein
• Function: hydrolyses cellular ATP to generate cAMP

Question 14

State the activation and inhibition of adenylyl cyclase

Answer 14

• Activation: via Gs – noradrenaline/adrenaline at β-adrenoceptors
• Inhibition: via Gi – noradrenaline/adrenaline at α₂-adrenoceptors

Question 15

What does cAMP do?

Answer 15

Cyclic AMP interacts with protein kinase A (PKA)

Question 16

What does PKA do?

Answer 16

PKA phosphorylates a variety of cellular proteins to increase/decrease their levels of activity:

• Glycogenolysis and gluconeogenesis in the liver
• Lipolysis in adipose tissue
• Smooth muscle changes
• Inotropic and chronotropic effects in the heart

Question 17

State the structure and function of Phospholipase C

Answer 17

• Structure: an effector enzyme in the cell signalling pathway involving the hydrolysis of PIP₂
• Function: generates secondary messengers – IP₃, DAG

Question 18

What is the role of IP₃ ?

Answer 18

IP₃ interacts with intracellular receptors on the ER to release Ca²⁺ from its so it can enter the cytoplasm e.g. activation of Ca²⁺- sensitive protein kinases

Question 19

What is the role of DAG?

Answer 19

DAG interacts with a family of protein kinases (PKC), which are activated by this second messenger

Question 20

The various intracellular signalling pathways involving GPCRs can be summarised using the mnemonic QIS.

What does this mean?

Answer 20

Q = PLC pathway

I = Inhibitory AC pathway

S = Stimulatory AC pathway

Question 21

The various types of adrenergic receptors can be summarised using the mnemonic QISS.

What does this mean?

Answer 21

Q - α₁
I - α₂
S - β₁
S - β₂

(Q = PLC pathway, I = Inhibitory AC pathway, S = Stimulatory AC pathway)

Question 22

The various types of muscarinic receptors can be summarised using the mnemonic QIQ

What does this mean?

Answer 22

Q - m₁
I - m₂
Q - m₃

(Q = PLC pathway, I = Inhibitory AC pathway, S = Stimulatory AC pathway)