G-protein Linked Receptors - FFP1-62 PDF

RCSI Royal College of Surgeons in Ireland Coláiste Ríoga na Máinleá in Éirinn FFP1-62 Introduction to Receptors: G protein-coupled receptors Prof Will Ford 337 [email protected]...

RCSI Royal College of Surgeons in Ireland Coláiste Ríoga na Máinleá in Éirinn FFP1-62 Introduction to Receptors: G protein-coupled receptors Prof Will Ford 337 [email protected] Dr. Roger Preston  Learning Outcomes Outline the concept and nature of receptor signalling Explain the structure of G-protein-coupled receptors Explain the nature of the signalling cascades G- protein coupled proteins can generate Describe the mechanisms by which G- proteins regulate the effector enzymes Receptor response theory Four types of receptor – others are available  G-protein coupled receptor (GPCR) structure Monomeric proteins MW 35K- 70K Pass through the membrane 7 times At least 500 different receptors Include light, taste and smell Overview of how GPCRs work Signal amplification  Key points about G-proteins 1. An enzyme composed of 3 subunits: , ,  2. Bind to and hydrolyse GTP to GDP 3. Inactive when GDP bound 4. Active when GTP bound 5. Acquires high What does each part of the G- protein do? The activity status of G proteins is determined by the  subunit 4 families of G proteins based on structural similarities – Gs, Gi, Gq and G12 Main purpose is to regulate amplifier or effector protein activity – βγ exist as dimers – 6 different β and 11 different γ – Can also exert signalling activity FYI  GPCR signalling The  and  of the G-protein anchor it to the membrane in its “inactive” or “unbound” state G-protein is not linked to the receptor G-protein has GDP-bound Lodish et al. Molecular Cell Biology  Ligand binding and activation Agonist-induced activation induces conformational change Conformational change reveals binding site for - subunit Lodish et al. Molecular Cell Biology  G-protein binding to receptor G-protein binds to receptor Lodish et al. Molecular Cell Biology  Signal transduction begins GDP is replaced with GTP GTP-Gα dissociates from Gβγ Lodish et al. Molecular Cell Biology  Second messengers are produced GTP-Gα and / or Gβγ activate the effector Meanwhile the agonist has dissociated Lodish et al. Molecular Cell Biology  The system resets GTPase activity returns the system to resting The cycle can start again Lodish et al. Molecular Cell Biology G-protein-linked effectors Liu et al, 2024, Circ Res 135(1). https://doi.org/10.1 161/CIRCRESAHA.1 24.323067 Regulatory control of GPCRs Guanine-nucleotide exchange factors (GEF) GDI Ligand-bound receptor acts GEF (Accelerates signalling) Guanine nucleotide dissociation inhibitor (GDI) βγ acts as GDI preventing GDP release (Inhibits signalling) GTPase-accelerating proteins (GAPs) Stimulate GTPase activity (Turn off signalling) GPCR desensitisation FYI 1) Uncoupling Phosphorylation uncouples receptor stopping recruitment of G-protein. P-receptor has lower affinity for agonist (dissociation) 2) Internalisation Sustained stimulation allows binding of β- arrestin leading to receptor internalization and activation other transduction pathways. 3) Downregulation Continued stimulation of receptor traffics receptor to lysosomes where it is Albert et al. (2005). The Neuroscientist 10. 575-93. Activation of adenylyl cyclase AC generates cAMP, an important ‘second messenger’ in cells Activated by Gs Inhibited by Gi Cell Signalling Biology - Michael J. Berridge - www.cellsignallingbiology.org - 2009 Ga subunits control AC activation Gαs activates adenylyl cyclase Activated by cholera toxin Inhibited by pertussis toxin Gai inhibits adenylyl cyclase Receptors regulating adenylyl cyclase Lodish et al. Molecular Cell Biology Chapter 13  Ga subunits control AC activation 160,00 deaths / year  Role of cyclic adenosine monophosphate cAMP activates protein kinases A (PKAs) Protein kinases phosphorylate proteins* cAMP-dependent kinases have many substrates ‐ ion channels and ‐ metabolic pathways cAMP is metabolised When AC activity is disrupted E. coli toxin – E. coli ‘traveller’s diarrhoea’ toxin Covalent modification of Gαs - can’t hydrolyse GTP (locked ‘ON’) Elevated cAMP levels in colonic epithelium cause efflux of water and ions Severe diarrhea and dehydration Treatment for this disruption Loperamide/Imodium E. coli Loperamide* acts as a μ-opioid toxin receptor agonist in large intestine Treatment – opiate receptor coupled to Gi Another example of functional antagonism  Activation of phospholipase C Classically activated by Gαq/11 Hydrolyses phosphoinositide (PIP2) from the membrane 1. Diacylglycerol (DAG) 2.Produces second Inositol 1,4,5-trisphosphate (IP3) messengers PLC-generated signal transduction Ca2+ calmodulin Activation of kinases Phosphorylation cellular proteins Change in cellular function  FYI Diversity of GPCRs Gs Gi Gq/11 5-HT4, 5-HT7 5-HT1, 5-HT5 5-HT2 ACh M2, M4 ACh M1, M3, Adenosine A2 Adenosine A1, A3 M5 Adrenergic β1, Adrenergic α2 2, 3 Cannabinoid CB Adrenergic α1 Dopamine D2, 3, 4 Dopamine D1, Glutamate D5 mGlu2, 3, 4, 6, 7, 8 Glutamate Histamine H3, 4 mGlu1, 5 Opioid δ, κ, μ Histamine H1 Histamine H2 Prostanoid EP3 Vasopressin Vasopressin V V1  FYI Diversity of GPCR signalling  FYI Even more signaling pathways!!!!! Pharmacology, Rang & Dale p 67-8  Further reading and viewing https://www.nature.com/scitable/topicpage/gpcr-1404747 1 Katzung Rang Medical Pharmacology at a glance https://portlandpress.com/pages/cell_signalling_biology  What we have learned The concept and nature of receptor signalling The structure of G-protein-coupled receptors The nature of the signalling cascades G- protein coupled proteins can generate The mechanisms by which G-proteins regulate the effector enzymes – adenylate cyclase and phospholipase C