Signal Transduction & Receptor Superfamilies - GPCRs 1 (2023-24) PDF

WEEK 26 MPharm Programme Signal Transduction & Receptor Superfamilies G-Protein-Coupled Receptors 1 Dr Gabriel Boachie-Ansah [email protected] Dale 113 ext. 2617 MPharm PHA112 Receptor Superfamilies - GPCRs WEEK Outline of Lectures 26 The keys steps in receptor signalli...

WEEK 26 MPharm Programme Signal Transduction & Receptor Superfamilies G-Protein-Coupled Receptors 1 Dr Gabriel Boachie-Ansah [email protected] Dale 113 ext. 2617 MPharm PHA112 Receptor Superfamilies - GPCRs WEEK Outline of Lectures 26 The keys steps in receptor signalling /signal transduction Signal transduction pathways & receptor superfamilies Structure & function of G-Protein-coupled receptors (GPCRs) Structure & role of G-proteins in GPCR signalling cAMP, IP3/Ca2+ as second messengers, and their key roles in GPCR signalling GPCR desensitisation & intracellular trafficking The role of GPCRs in the actions of neurotransmitters & hormones Slide 2 of 59 MPharm PHA112 Receptor Superfamilies - GPCRs Learning Outcomes WEEK 26 At the end of this lecture, you should be able to: Describe the keys steps in receptor signalling or signal transduction Describe the major signal transduction pathways & receptor superfamilies Describe the structure & function of G-proteins & Gprotein-coupled receptors (GPCRs) Define & name examples of second messengers, and describe their key roles in GPCR signalling Explain GPCR desensitisation & intracellular trafficking Name some of the GPCRs that mediate that actions of key neurotransmitters & hormones Slide 3 of 59 MPharm PHA112 Receptor Superfamilies - GPCRs WEEK 26 Signal Transduction How is the Drug-Receptor binding translated into a Biological Response? Slide 4 of 59 MPharm PHA112 Receptor Superfamilies - GPCRs WEEK Receptor Signalling 26 When an agonist drug binds to its receptor A drug-receptor (D-R) complex is formed The D-R complex undergoes a conformational change This triggers a chain of biochemical processes inside the cell  a biological response This process is called ‘signal transduction’ Slide 5 of 59 MPharm PHA112 Receptor Superfamilies - GPCRs WEEK Receptor Signalling 26 3 stages of the receptor signalling process Signal reception the agonist drug binds to and activates a specific ‘receptor’ on / inside the target cell Signal transduction the drug-receptor complex activates a series of relay proteins & produces 2nd messengers inside the cell Cellular response eventually a cellular or biological response to the original drug binding signal is triggered Slide 6 of 59 MPharm PHA112 Receptor Superfamilies - GPCRs WEEK 26 The 3 Stages of the Drug-Receptor Signalling Process Slide 7 of 59 MPharm PHA112 Receptor Superfamilies - GPCRs WEEK 26 Signal Transduction Pathways 4 Major Signal Transduction Pathways Activation of receptor-ion channels (Ligandgated receptors) Activation of second messenger pathways via G-protein-coupled receptors Activation of enzyme-linked receptors (e.g. Tyrosine kinase-linked receptors) Direct activation of gene transcription via Intracellular receptors Slide 8 of 59 MPharm PHA112 Receptor Superfamilies - GPCRs WEEK Receptor Superfamilies 26 Receptor Superfamily A group of receptors with a similar basic molecular structure and that use the same signal transduction pathway 4 Major Receptor Superfamilies Ligand-gated / Ion channel linked receptors G-protein-coupled receptors Kinase-linked receptors Intracellular / Nuclear receptors Slide 9 of 59 MPharm PHA112 Receptor Superfamilies - GPCRs WEEK 26 Slide 10 of 59 Receptor Superfamilies MPharm PHA112 Receptor Superfamilies - GPCRs WEEK 26 G-protein-coupled Receptors (GPCRs) (Metabotropic Receptors) Slide 11 of 59 MPharm PHA112 Receptor Superfamilies - GPCRs WEEK 26 G-protein-coupled Receptors (GPCRs) A large & diverse superfamily of integral membrane proteins used by cells to convert extracellular signals into intracellular responses They constitute the largest receptor superfamily in humans (~800 members) They transduce a wide array of extracellular signals and regulate virtually every aspect of physiology They mediate responses to hormones, neurotransmitters and growth factors, as well as responses to vision, olfaction & taste signals They are the targets of ~40% of drugs currently on the pharmaceutical market Slide 12 of 59 MPharm PHA112 Receptor Superfamilies - GPCRs WEEK 26 G-protein-coupled Receptors (GPCRs) Key Features & Characteristics of GPCRs They all share a common structural motif of seven transmembrane (7-TM) α-helices They couple to & activate cytoplasmic heterotrimeric G-proteins upon agonist binding, leading to modulation of downstream effector proteins  biological response They also couple to cytoplasmic adaptor proteins, called -arrestins, leading to receptor desensitisation & internalization or activation of downstream effector proteins  biological response Slide 13 of 59 MPharm PHA112 Receptor Superfamilies - GPCRs WEEK Structure of GPCRs 26 A GPCR typically consists of a single polypeptide chain with 3 key regions The extracellular region – the N terminus & three extracellular loops (ECL1-ECL3) modulates ligand access to the binding site on receptor The TM region – seven transmembrane (7TM) α-helices (TM1-TM7) forms the structural core, binds to ligands & transduces this information to the intracellular regions The intracellular region – three intracellular loops (ICL1ICL3), short intracellular α-helix (H8) & the C terminus interfaces with cytosolic signalling proteins, e.g. G-proteins Slide 14 of 59 MPharm PHA112 Receptor Superfamilies - GPCRs WEEK 26 General Architecture of a G Protein-coupled Receptor (GPCR) Slide 15 of 59 MPharm PHA112 Receptor Superfamilies - GPCRs WEEK 26 Slide 16 of 59 General Architecture of a GPCR MPharm PHA112 Receptor Superfamilies - GPCRs WEEK 26 GPCR Signalling via G-Proteins & Arrestins Effector enzyme Slide 17 of 59 MPharm PHA112 Receptor Superfamilies - GPCRs WEEK 26 GPCR Signalling via Heterotrimeric G Proteins The key feature of GPCRs is their interaction with heterotrimeric GTP-binding proteins (or G-proteins) Heterotrimeric G-proteins play a pivotal role in the signaltransduction pathways initiated by G-protein-coupled receptor (GPCR) activation They are localised at the inner leaflet of the plasma membrane – convey signals from cell-surface GPCR to downstream intracellular effector proteins They act as molecular binary switches – translate agonistGPCR binding into modulation of activity of downstream intracellular effector proteins  biological response Slide 18 of 59 MPharm PHA112 Receptor Superfamilies - GPCRs WEEK Heterotrimeric G-Proteins 26 Heterotrimeric G-proteins are composed of 3 different protein subunits: ,  and  Functionally, they consist of two units: an α subunit (Gα) and a tightly associated  complex Both the G and G subunits have lipid extensions that bind & tether the G-protein complex to the plasma membrane The Gα subunit harbours a guanine nucleotide-binding site, which is occupied by GDP in the inactive resting (off) state So far, 21 G, 5 G and 12 G subunits/isoforms have been identified in the human genome  multiple permutations of distinct heterotrimeric complexes Slide 19 of 59 MPharm PHA112 Receptor Superfamilies - GPCRs WEEK 26 Heterotrimeric G-protein in the Resting GDP-bound State Gα subunit is composed of 2 domains: a Ras-like domain & an α-helical domain A nucleotide-binding pocket is located between the two domains The Ras-like domain has GTPase activity (hydrolyzes GTP to GPD) & also provides binding sites for the Gβ subunits The N-terminus of Gα is myristoylated or palmitoylated, which results in the attachment of the G protein to the plasma membrane Slide 20 of 59 MPharm PHA112 Receptor Superfamilies - GPCRs WEEK 26 Signalling via Heterotrimeric G-Proteins Interaction with agonist-bound GPCR  conformational change  exchange of GDP for GTP on the Gα subunit  dissociation of G-GTP subunit from the G dimer Both G-GTP & the freed G proceed to interact with, and regulate the activity of, unique downstream effector proteins  biological response Ultimately, the activated G-protein returns to the inactive resting (off) state GTPase activity in the Gα subunit hydrolyses bound GTP to GDP Hydrolysis of GTP to GDP is accelerated by regulators of G-protein signalling (RGS) proteins or GTPase-accelerating proteins (GAPs) G-GDP then re-assembles with the G dimer to form the inactive G-protein Slide 21 of 59 MPharm PHA112 Receptor Superfamilies - GPCRs WEEK 26 Mechanism of G-Protein Activation By GPCRs Slide 22 of 59 MPharm PHA112 Receptor Superfamilies - GPCRs WEEK 26 Slide 23 of 59 The Guanine Nucleotide Cycle of Heterotrimeric G-Proteins MPharm PHA112 Receptor Superfamilies - GPCRs WEEK 26 Slide 24 of 59 The Guanine Nucleotide Cycle of Heterotrimeric G-Proteins MPharm PHA112 Receptor Superfamilies - GPCRs WEEK 26 Diversity of GPCR Signalling Mechanisms Signalling via Gα Proteins G-proteins are classified based on their Gα subunits Gα proteins are grouped into 4 families based on their peptide sequence & functional similarities – Gαs, Gαi, Gαq/11, and Gα12/13 protein families Each Gα family can relay GPCR signals to multiple downstream effectors  triggering of different signalling pathways Gαs family – stimulate adenylate cyclase   cAMP Gαi family – inhibit adenylate cyclase   cAMP Gαq/11 family – stimulate phospholipase C-β   IP3 & DAG Gα12/13 family – activate the Rho family of GTPases Slide 25 of 59 MPharm PHA112 Receptor Superfamilies - GPCRs WEEK G-Protein Families & Effectors 26 Gα family Gα family members Tissue expression Gαs Gαs, Gαolf Gαi Gαi1, Gαi2, Gαi3, most cell types; high in Gαo, Gαt, Gαg, neurons (Gαo); rod & cone Gαz cells of the eye (Gαt); taste receptor cells (Gαg); neuronal tissues & platelets (Gαz) Gαq, Gα11, most cell types (Gαs); Gα14, Gα16 olfactory neurons (Gαq, Gα11); kidney, lung & liver (Gα14); haematopoietic cells (Gα16) Gα12, Gα13 most cell types Gαq Gα12/13 Slide 26 of 59 MPharm most cell types (Gαs); olfactory neurons (Gαolf) PHA112 Effector proteins & Effect Activate adenylate cyclase Inhibit adenylate cyclase Activate cGMP phosphodiesterase Activate phospholipase C-β Modulate Rho & RasGEFs Receptor Superfamilies - GPCRs WEEK 26 GPCR-mediated Signalling Pathways Slide 27 of 59 MPharm PHA112 Receptor Superfamilies - GPCRs WEEK 26 GPCR-mediated Signalling Pathways Slide 28 of 59 MPharm PHA112 Receptor Superfamilies - GPCRs WEEK 26 GPCR-mediated Signalling Pathways Slide 29 of 59 MPharm PHA112 Receptor Superfamilies - GPCRs WEEK 26 G-mediated Signalling Pathways Most G proteins mediate GPCR signalling by regulating the levels of intracellular regulatory molecules, called second messengers The second messengers regulate the activity of multiple downstream effector proteins  biological response Key second messengers include: cAMP, IP3 and Ca2+ The cAMP signalling pathway Gs-GTP activates Adenylyl Cyclase (AC)   cAMP Gi/o-GTP inhibits Adenylyl Cyclase (AC)   cAMP The Inositol 1,4,5-trisphosphate/calcium signalling pathway Gq/11-GTP activates Phospholipase C-β (PLCβ)   IP3 & DAG   Ca2+ Slide 30 of 59 MPharm PHA112 Receptor Superfamilies - GPCRs WEEK 26 G-mediated Signalling Pathways IP3 cAMP PATHWAY Slide 31 of 59 MPharm PHA112 IP3/Ca2+ PATHWAY Receptor Superfamilies - GPCRs