Session 7 - Receptor-Effector signalling Flashcards
Name the three superfamilies of cell surface receptor
- Ligand-gated
- Receptors with intrinsic activity
- GPCR
How do GPCRs alter cellular functions?
-Use Guanine nucleotide binding proteins (Gproteins) to activate/inhibit second-messenger generating enzymes or ion channels which then alter cellular function
What is signal transduction?
-The transfer of a signal produced by the binding of an extracellular ligand to the target effector intracellularly to generate the desired response
What is the basic structure of GPCRs?
- Single polypeptide chain with 7 TMDs
- Extracellular N terminal and intracellular C terminal (most)
- 2 ligand binding sites –> Some onto N terminal and 2-3 TMD cleft
How is the signal passed from the receptor to the G protein?
-Agonist-binding to GPCR activates the receptor and causes a conformational change which allows interaction and activation of the G protein
Why are G proteins described as heterotrimeric?
-Made of 3 subunits a,b,g
What are the two functional units of a G-protein?
- a
- bg
Which subunit of a G protein has a guanine binding site?
-a
Describe activation of the G protein
- In basal state Ga is bound to bg and GDP
- Upon activation, GDP leaves the subunit and GTP binds in its place -> (GDP/GTP exchange)
- The binding of GTP to Ga decreases Ga affinity for the receptor and Gbg causing dissociation
- Ga and Gbg are not free to interact with effectors
What facilitates GDP-GTP exchange?
-The receptor as it acts as a guanine nucleotide exchange factor
How is Gprotein signalling terminated?
- Ga has an intrinsic GTPase which slowly hydorlyses GTP to GDP
- Ga-GDP has increased affinity for Gbg and the heterotrimeric complex is reformed and becomes inactive, awaiting receptor activation
What determines the magnitude of the transduction from GPCRs?
-The timer function of GTPase of Ga which can be controlled by other proteins (RGS protein)
How is there a large range of diversity amongst GPCRs?
-20Ga, 5Gb and 12+Gg which vary in combinations producing over 1000 possible combinations
With a huge range of diversity, what ensures specificity amongst GPCR signalling?
-Each GPCR has preferential interactions with specific types f G proteins, determined mainly by Ga type
What is the effector of Gas?
-Activates adenyl cyclase=increased cAMP
What is the effector of Gai?
-Inhibits adenyl cyclase=decreased cAMP
What is the effector of Gaq?
-Activates phospholipase C= increased IP3 and DAG, regulates PKC
What is the effector of Gat?
-cGMP phosphodiesterase
How does cholera toxin work?
-Contains ADP-ribosyltransferase activity which adds an ADP-ribosyl group derived from NAD+ to Gas which inhibits GTPase activity of Gas = constitutively active
How does whooping cough occur?
-Pertussin toxin from bordella pertussis contains ADP-ribosyltransferase activity which modifies Gai to prevent receptor association and Gai protein activation
Give an example of GPCR signalling to activate and inhibit AC using the same ligand
-NA can act on B2-adrenoreceptors to activate AC via Gas or acts on a2-adrenoreceptors to inhibit AC via Gai
What is the function of AC?
-Hydrolyse cellular ATP to generate cAMP
What enzyme does cAMP regulate?
-PKA
What cellular responses does PKA regulate? (give a few examples)
-Glycogenolysis + gluconeogenesis
-Lypolysis
-Relaxation of smooth muscle
Positive inotropic/chronotropic effects in the heart
Which important receptors are linked to Gas?
- B-adrenoreceptors
- D1-dopamine receptors
- H2-histamine receptors
Which important receptors are linked to Gai?
- a2-adrenoreceptors
- D2-dopamine receptors
- H-opiod receptors
Describe the activation of PKA
- 2 regulatory domains sit in the active sites of 2 catalytic domains
- Catalytic domains have intrinsic protein kinase activity
- cAMP binds to regulatory units causing dissociation and activation as active sites are now free
What does PKA target on effectors?
-Serine/threonine residues
What is the main function of PLC?
-Cleavage of PIP2 into IP3 and DAG
What are IP3 and DAG?
-Second messengers
What is the function of IP3?
-Bind to IP3 receptors on SR opening a ligand-gated Ca channel, allowing Ca to enter cell down its conc gradient
How can PKC be activated?
- Increasing intracellular calcium
- Interaction with DAG
Which Gprotein activates PLC?
-Gaq
What are some cellular changes regulated by PLC?
- Vascular, GI tract and airway smooth muscle contraction
- Mast cell degranulation
- Platelet aggregation
Which GPCRs associate with Gaq?
- a1-adrenoreceptors
- M1-muscarinic receptors
Describe the activation and function of cGMP phosphodiesterase, in relation to the eye?
- In the dark, levels of cGMP sufficient to open ion channels allowing Ca and Na to enter
- Gat becomes activated by a photon of light
- Activated Gat activates cGMP phosphodiesterase which breaks down cGMP -> channel closure and membrane hyperpolarisation
- Alters the signal input to CNS
How is signal amplification achieved?
1) Activted GPCR causes GDP/GTP exchange on multiple Gproteins
2) Activates Ga-GTP/Gbg activates multiple effectors
3) Effector molecules act catalytically ie 1 activates 100-1000s
4) Cellular targets of second messengers are often enzymeswhich act catalytically causing an enzyme cascade
What mechanisms control deactivation of signal transduction from GPCRs?
1) Agonist-receptor dissociation
2) Activated GPCr becomes phosphorylated preventing activation of further G proteins (receptor desinsitization)
3) GTPase of a-GTP
4) Intrinsic enzyme inhibitors of effector molecules
How is chronotropy of the heart decreased through Gproteins?
- Ach released from parasympathetic NS
- acts on M2-muscarinic receptors
- Leads to the opening of K+ channels by Gbg causing hyperpolarisation and increased K+ conductance should increase HR but….
- Decreased cAMP due to inhibition of AC by Gai
- Decreased activation of HCN channels -> HR slows
How is inotropy of the heart increased through GPCRs?
- Increased sympathetic innervation -> NA and A activates B-adrenoreceptors
- > Gas dissociates and activates AC -> increased cAMP
- > Increased cAMP increases PKA activation
- > PKA phosphorylates Ca channels causing then to open and increase intracellular Ca during plateau phase
- B-adrenoreceptors also cause Increased SR uptake of Ca and increase sensitivity of contractile machinery to Ca
How is arteriolar vasoconstriction activated through GPCRs?
- Increased sympathetic activity = increased NA
- NA acts on a1-adrenoreceptors on smooth muscle which associate with Gaq
- Gaq activates PLC -> cleaves PIP2 to IP3 and DAG
- IP3 causes Ca release from SR -> initiates contractile response
How is bronchoconstriction activated through GPCRs?
-Ach acts on M3-muscarinic receptor which associates with Gaq -> PLC-> IP3-> increased intracellular calcium -> contraction of smooth muscle
How can neurotransmitter release be modulated through u-opioid receptor?
- Pre-synaptoc u-opiod receptors can be stimulated by endogenous opiod (encephalin) or by analgesic such as morphine
- GPCR associated with Gai; activated Gai which dissociate from Gbg
- Gbg interact with VOCCs and reduce entry of Ca2+
- > decrease in Ca2+ influx to synaptic bouton inhibits the release of neurotransmitter
B-adrenoreceptors are coupled to which Gprotein?
-Gas
a-adrenoreceptors are coupled to which G-protein?
- a1 coupled to Gaq
- a2 coupled to Gai
-Muscarinic ach receptors are coupled to which Gproteins?
- M1, M3 and M5 coupled to Gaq
- M2 and M4 coupled to Gai
Describe a pneumonic for remembering receptors and their associated G proteins
Qu - a1 Q - M1
I - a2 I - M2
S - b1 Q - M3
S - b2
