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G-protein-coupled receptors (GPCRs)
Graduate Institute of Anatomy and Cell Biology
台大醫學院 解剖學暨細胞生物學研究所
黃敏銓老師實驗室
Graduate Institute of Anatomy and Cell Biology
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The Nobel Prize in Chemistry 2012 is awarded to Brian K. Kobilka (57) and Robert J. Lefkowitz (69) for studies of GPCRs.
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G-protein-coupled receptors (GPCRs)
名稱 & 特徵
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CCR5
G protein-coupled receptors (GPCRs) Seven transmembrane receptors (7 TM receptors) Heptahelical receptors Serpentine receptors
名 稱
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Single Transmembrane Spanning Heterotrimeric GProtein-Coupled Receptors
Pharmacol Rev. 2004 Sep;56(3):371-85.
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Characteristics of GPCRs
7 transmembrane domains
3 intracellular loops
3 extracellular loopsCytoplasmic
ExtracellularNH2
HOOC
A Transmembrane Intracellular Estrogen Receptor Mediates Rapid Cell Signaling.GPR30 is uniquely localized to the endoplasmic reticulumRevankar CM et al., Science. 2005. 307:1625.
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G-protein-coupled receptors (GPCRs)
Genomics: > 800 members
wide range of cellular activities
from yeast to human
50% drug target
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G-protein-coupled receptors (GPCRs)
分類
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G-protein-coupled receptor (GPCR) families
• Class A Rhodopsin like
• Class B Secretin like
• Class C Metabotropic glutamate / pheromone
• Class D Fungal pheromone
• Class E cAMP receptors (Dictyostelium)
• Frizzled/Smoothened family
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Family classification of known GPCRs
NATURE REVIEWS | DRUG DISCOVERY VOLUME 1 | AUGUST 2002 | 599-608
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NATURE REVIEWS | DRUG DISCOVERY VOLUME 1 | OCTOBER 2002 | 808-820
The three main families of GPCRs
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GRAFS system Five main classes: Glutamate, Rhodopsin, Adhesion, Frizzled, and Secretin
Annu Rev Pharmacol Toxicol. 2013;53:127-46
Orphan receptors(unknown ligands)
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G-protein-coupled receptors (GPCRs)
訊息傳遞
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http://stke.sciencemag.org/content/vol0/issue2002/images/data/CMP_6634/DC1/science3.jpg, by R. Iyengar's laboratory
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Summary of GPCR signalling
NATURE | VOL 415 | 10 JANUARY 2002 206-212
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G protein action
GPCR
ligand1.
GDP βγα
2.
αβγ
GTP
GDP
GPCR(GEF)
second messenger
αβγ
GTP
3.
(GAP)
effectorGPCR
β
4.
αγGDP
Pi
stops
GPCR
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G protein cycle for the β2AR–Gs complex
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Nature. 2011 Jul 19;477(7366):549-55.
Kobilka’s group
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NATURE REVIEWS | MOLECULAR CELL BIOLOGY VOLUME 3 | SEPTEMBER 2002 | 639-650
A classical example of GPCR signalling
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Modified from TRENDS in Pharmacological Sciences Vol.22 No.7 July 2001, 368-376
GPCR G protein effector activities
4 大類
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Regulation of systemic functions by signaling through G protein pathways
31 MAY 2002 VOL 296 SCIENCE, 1636-1639
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NATURE REVIEWS | DRUG DISCOVERY VOLUME 1 | MARCH 2002 | 188
Heterotrimeric G-protein α-subunit subfamilies and their effects
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G-protein-coupled receptors (GPCRs)
Desensitization
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Desensitization(receptor level)
(A) Rapid: phosphorylation (sec~min)
(B) Slow: degradation or expression (hours)
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Desensitization(A) Rapid: phosphorylation (sec~min)
1. Second-messenger kinase (PKA, PKC):uncouple GPCR from G proteinligand independentheterologous desensitization
2. GPCR kinase (GRK): GRK1-GRK7binding of β-arrestin and inhibition of G protein interactionligand-activated or activated conformationhomologous desensitization
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DesensitizationGPCR kinase (GRK) phosphorylation
NATURE REVIEWS | MOLECULAR CELL BIOLOGY VOLUME 3 | SEPTEMBER 2002 | 639-650
arrestin 1-arrestin 4
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Recycling or degradation?Desensitization: (B) Degradation
SCIENCE VOL 297 26 JULY 2002
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Desensitization
a. GTPase-activating proteinsb. accelerate GTP hydrolysisc. >25 members, for selectivity
RGS (regulator of G-protein signaling)
(Post-receptor level)
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Desensitization by regulator of G-protein signalling (RGS)
NATURE REVIEWS | DRUG DISCOVERY VOLUME 1 | MARCH 2002 | 187-197
RGS2 knockout mice:• T-cell activation• Anxiety• Male aggression
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G-protein-coupled receptors (GPCRs)
Internalization
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Internalization
1. Desensitization2. Resensitization3. Signal transduction
Functions:
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Internalization
• Clathrin-coated pits–β-arrestin
• Caveolae-mediated– Lipid rafts: signalling
• Uncoated vesicles
Types:
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β-arrestin-dependent internalization of GPCRs
NATURE REVIEWS | NEUROSCIENCE VOLUME 2 | OCTOBER 2001 | 727-733
Function 1: Desensitization
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Function 2: Resensitization
GRP: GPCR specific phosphatase
Pharmacol Rev 53:1–24, 2001
P
P
P
P
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NATURE REVIEWS | NEUROSCIENCE VOLUME 2 | OCTOBER 2001 | 727-733
Function 3: Signal transduction
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TRENDS in Pharmacological Sciences Vol.28 No.8 417
Annu. Rev. Physiol. 2007. 69:511–34
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GPCR signaling via heterotrimeric G proteins from endosomes
J Biol Chem. 2015 Jan 20. [Epub ahead of print]
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G-protein-coupled receptors (GPCRs)
G-protein-independent signalling
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G-protein-independent signalling
Regulation of GPCR Signalling by Scaffold Proteins
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39Circ Res. 2002;91:672-680
Scaffold Proteins
1
2
3
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1. GPCR/PDZ scaffold interaction
Circ Res. 2002;91:672-680
(calcium channel)
PDZ: PSD-95/Discs-large/ZO-1
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β2 adrenoceptor regulates the Na+-H+ exchanger (NHE)
TRENDS in Biochemical Sciences Vol.26 No.5 May 2001, 291-297
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2. GPCR/non-PDZ scaffold interaction
Circ Res. 2002;91:672-680
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Metabotropic glutamate receptor-mediated Ca2+ release
TRENDS in Biochemical Sciences Vol.26 No.5 May 2001, 291-297
Ca2+ release
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3. GPCR/β-arrestin interaction
Circ Res. 2002;91:672-680
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β2 Adrenoceptor can mediate the Src kinase pathway
TRENDS in Biochemical Sciences Vol.26 No.5 May 2001, 291-297
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NATURE | VOL 415 | 10 JANUARY 2002 206-212
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Transactivation
NATURE REVIEWS | CANCER VOLUME 7 | FEBRUARY 2007 | 85
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48Trends Endocrinol Metab. 2006 Mar;17(2):48-54
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49Trends Endocrinol Metab. 2006 Mar;17(2):48-54
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G-protein-coupled receptors (GPCRs)
Oligomerization
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Signal amplification at the receptor level
NATURE REVIEWS | DRUG DISCOVERY VOLUME 1 | OCTOBER 2002 | 808-820
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Heterodimerization of the GABAB receptor produces a functional receptor
NATURE REVIEWS | MOLECULAR CELL BIOLOGY VOLUME 3 | SEPTEMBER 2002 | 639-650
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TRENDS in Pharmacological Sciences Vol.26 No.1 January 2005
Don’t worry ‘B’ happy!:a role for GABABreceptors in anxiety and depressionJohn F. Cryan and KlemensKaupmannNeuroscience Research, Novartis Institutes for BioMedical Research, Novartis Pharma AG, Basel, Switzerland
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EMBO reports VOL 7 | NO 11 | 2006, p. 1096
Ligand-independent functions of orphan GPCRs in heterodimers
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Bivalent and dimeric ligands
NATURE REVIEWS | DRUG DISCOVERY VOLUME 1 | OCTOBER 2002 | 808-820
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Drug discoveryGPCR Drugs >$20B annual sales
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GPCR and Diseases
• Obesity• Cardiovascular Disease• Inflammation• Cancer• Diabetes• Alzheimer’s Disaese
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G-protein-coupled receptors (GPCRs)
Drug screening
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Key requirements of a ligand screen
• Cell surface expression of GPCRs
• Cellular assays to detect ligand binding
• Libraries of putative natural and synthetic ligands
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Method for “de-orphanizing” GPCRs
NATURE REVIEWS | MOLECULAR CELL BIOLOGY VOLUME 3 | SEPTEMBER 2002 | 639-650
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FIRST™ : Functionally Immobilized Receptor Solubilization Technology
Consensus Pharmaceuticals, Inc.
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CARTConstitutively Active Receptor Technology
Arena Pharmaceuticals, Inc.
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DISEASE OVERACTIVE KNOWN GPCRs----------------------------------------------------------------Schizophrenia 5HT2A, D4----------------------------------------------------------------Depression 5HT2A----------------------------------------------------------------Hyperthyroidism Thyrotropin----------------------------------------------------------------Hypertension Angiotensin AT1A----------------------------------------------------------------Asthma Adenosine A1----------------------------------------------------------------Melanoma MC-1----------------------------------------------------------------Retinitis Pigmentosa Rhodopsin----------------------------------------------------------------
Diseases - GPCRs
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Nat Rev Drug Discov. 2011 Jan;10(1):47-60.
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Nat Rev Drug Discov. 2011 Jan;10(1):47-60.
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Representation of ‘inactive’ and ‘active’ GPCR conformations
NATURE REVIEWS | DRUG DISCOVERY VOLUME 1 | AUGUST 2002 | 599-608
inactive active
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CARTConstitutively Active Receptor Technology
Inverse agonists: block ligand-independent activity
Selective molecules: one ligand to multiple receptors
Agonists
Allosteric modulators
(Antagonist: block ligand-dependent activity)
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Summary
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Modified from TRENDS in Pharmacological Sciences Vol.22 No.7 July 2001, 368-376
Diversity of G-protein-coupled receptors (GPCRs)
Scaffold proteins
Internalization
signaling
RGS
GRKPKAPKC
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TRENDS in Pharmacological Sciences Vol.22 No.8 August 2001, 396
Treasures throughout the life-cycle of GPCRs