Lecture 27 – Cell Signalling II: G Protein Coupled Receptors and cAMP

Q: What type of signalling does a G protein coupled receptor (GPCR) use?

Second messenger–based signalling.

Q: What are G proteins, and how do they function as molecular switches?

GTP-binding proteins that exist in two interchangeable states, active (GTP-bound) and inactive (GDP-bound).

Q: How does a G protein switch from active to inactive?

By hydrolysing GTP to GDP (GTPase activity).

Q: How does a G protein switch from inactive to active?

By releasing GDP and binding GTP (nucleotide exchange).

Q: What are the subunits of a heterotrimeric G protein?

Alpha (α), beta (β), gamma (γ). The α and γ subunits are linked to the membrane via lipid groups.

Q: What is the role of heterotrimeric G proteins?

They relay the signal from the ligand-bound receptor to the cytoplasm or nucleus via an effector protein.

Q: Where is the GTP binding site located on a heterotrimeric G protein?

On the α (alpha) subunit.

Q: Walk through the GPCR activation cycle (steps 1–6).

(1) Ligand binds the receptor, causing a conformational change that increases its affinity for G proteins. (2) Gα releases GDP and binds GTP (nucleotide exchange). (3) The conformational change in Gα reduces its affinity for Gβγ, and Gα attaches to the effector. (4) The effector (e.g., adenylyl cyclase) produces cAMP as a second messenger. (5) GTP hydrolysis causes a conformational change in Gα, which loses affinity for the effector. (6) Gα dissociates from the effector and reassociates with the Gβγ dimer, reforming the inactive heterotrimer. As long as the ligand remains bound, the G protein can undergo another round of activation.

Q: How is a GPCR desensitised after activation?

G protein coupled receptor kinase (GRK) phosphorylates the receptor. Arrestin then binds the phosphorylated receptor, preventing it from activating additional G proteins. The receptor is likely taken up by endocytosis. Arrestin also facilitates degradation of second messengers.

Q: What does "arrestin" refer to functionally?

"arrests" (stops) the signalling process by blocking the phosphorylated receptor from activating more G proteins.

Q: What is the effector in the GPCR–cAMP pathway?

Adenylyl cyclase, an integral membrane protein whose catalytic domain resides at the inner surface of the plasma membrane.

Q: What does adenylyl cyclase do?

It catalyses the conversion of ATP to cyclic AMP (cAMP), the second messenger.

Q: What is a second messenger, and why is it important?

A small intracellular signalling molecule produced in response to a first messenger (extracellular ligand). It allows a wider, amplified response from a single extracellular signal.

Q: What enzyme breaks down cAMP, and what is the result?

Phosphodiesterase, brings signal transduction to a halt.

Q: Name three hormones that all activate adenylyl cyclase through their own GPCRs.

Adrenocorticotropic hormone (ACTH) from the pituitary gland, glucagon from the pancreas, and epinephrine from the adrenal glands.

Q: What happens when two of these hormones (ACTH, glucagon, epinephrine) are added together to a cell?

The effects are not additive, indicating that each hormone stimulates the same population of adenylyl cyclase molecules rather than separate pools. The hormones compete for, not share, adenylyl cyclase.

Q: What is the role of glucagon in glucose metabolism?

Stimulates the breakdown of glycogen to release glucose.

Q: What is the role of insulin in glucose metabolism?

Stimulates the uptake of glucose and storage of glycogen.

Q: What is epinephrine, and what does it do in the liver?

A fight-or-flight hormone produced by the adrenal gland. Triggers the same GPCR–cAMP pathway as glucagon, leading to glucose mobilization.

Q: What does cAMP activate in the glucose mobilisation pathway?

cAMP-dependent protein kinase A (PKA).

Q: What are the three actions of PKA in liver cells?

(1) activate phosphorylase kinase, which releases glucose into the bloodstream. (2) phosphorylates the transcription factor CREB in the nucleus, promoting gluconeogenesis. (3) interacts with glycogen synthase and inhibits glycogen production.

Q: How does signal amplification occur in the GPCR–cAMP pathway?

A single hormone molecule can activate multiple G proteins, each of which activates an adenylyl cyclase, each of which rapidly produces many cAMP molecules.

Q: What is CREB, and what does it do?

(cAMP response element–binding protein) is a transcription factor. When phosphorylated by PKA in the nucleus, it binds as a dimer to CRE (cAMP response element) sequences on DNA, activating genes involved in gluconeogenesis.

Q: What is gluconeogenesis?

A metabolic pathway by which glucose is formed from intermediates of glycolysis. Several of its enzymes are encoded by genes containing CRE elements.

Q: Why do different cell types show different responses to the same cAMP signal?

Different cell types express different PKA-dependent substrates, leading to cell type–specific responses even though the upstream pathway (cAMP/PKA) is the same.

Q: What are the three core principles of GPCR signalling?

(1) The receptor undergoes a conformational change during activation or deactivation. (2) G proteins help activate specific effector molecules. (3) Signal amplification takes place at key steps in the cascade.