G-protein linked receptors

Diversity of GPCRs

IV G12 activates chloride channel G13 activates Na+/H+ exchanger pathways

activates small G protein Rho family, leading to actin cytoskeletal reorganization

extra-cellular

G-protein-linked receptors; functionally diverse but sharea common structure

Ligand binding

G-protein-linked receptors function through trimeric G-proteins

G-protein-linked receptors mediate their intracellular actionsthrough target ion channels or enzymes. pathway always involves activation of one or more guanine nucleotide-binding regulatory proteins (trimeric G proteins).

trimeric G proteins consist of three protein subunits; alpha, beta and gamma. The G alpha binds a guanyl nucleotide.Various types, each specific for a set of serpentine receptors and for a particular downstream target, but they all operate the same way.

Ligand binding (1)-conformational change in thereceptor -separates TMs.-separation of the TMs may open a crevice for binding to G.

Receptor binds to G protein (2)

Receptor, inactive G-proteins,and adenylyl cyclase are withinshouting distance in the cellmembrane.

Receptor activation results in activation of adenylyl cyclase.-indirect-stimulates a trimeric G-protein-trimeric G-proteins dissasemblewhen activated.

Receptor binds to G-proteininduces conformational change (3)GDP is replaced by GTPG dissociates from G

G then binds to adenylyl cyclase (4),activating synthesisof cAMP

The binding site for adenylyl cyclase is unmasked.

A single hormone/receptor complexstimulates the productionof many molecules of Gs

Binding of G toadenylyl cyclasecauses a conformationchange in G andGTP is hydrolyzed toGDP. This causesG to dissociate fromadenylyl cyclaseand re-bindG

The binding of the Gs

subunit to adenylyl cyclase

activates the enzyme to

produce many molecules of cAMP.

signal amplification

Terminating the response

The hormone/receptor complex must be deactivated to return to

the unstimulated state.

-phosphorylation events on the carboxy terminal tail of the receptor

lead to the inactivation of the receptor.Hydrolysis of GTP leads to inactivation of the trimeric G-protein.

-enhanced by RGS proteins (regulator of G-protein signaling).

Functionally couple the G-protein-linked receptors to their target enzymes or ion channels.

GTPases that function as molecular switches-flip between two states: active and inactive.-Inactive; trimer bound to GDP through G-Active; G bound to GTP

A G-protein which acts to stimulate a target enzyme is called a G stimulatory (Gs).

Gi is inhibitory.

The trimeric GTP-binding proteins act as molecularswitches

active

inactive

two conformation states

Mammalian RGS proteins activate the GTPase activities of G-protein alpha subunits

RGS proteins are GAPs (GTPase activating proteins).-no effect on the time course of nucleotide binding-but they stimulate the rate of GTP hydrolysis.

MODEL: RGS proteins accelerate GTP hydrolysis by preferentially binding to and stabilizing G proteins in their transition state for the hydrolysis reaction.

GTP hydrolysis

RGS protein

GDP

GTP

/

Inhibitory G proteins

While the beta-adrenergic

receptors are functionally

coupled to G-stimulatory

proteins, the alpha-2

adrenergic receptors are

coupled to inhibitory G

proteins.

Gi can contain the same beta/gamma subunits as Gs, but the alpha subunits are different.

Gi inhibits adenylyl cyclase in an indirect manner.

Hormone-induced activation and inhibition of adenylyl cyclase is mediated by G-s and G-i

PLC-

PIP2 DAG+IP3

Ras/Raf

MEK

MAPK

AC

ATP cAMP

PKC activation

[Ca+2]i

mobilizationproliferation

Rho

actin cytoskeletal rearrangement

cell cycle progression

AC

ATP cAMP

Gs

PI3K

AKT Rac

survival

out

in

GPCR-mediated Signaling Pathways

membrane

GPCR Classes

Class A: Rhodopsin likeClass B: Secretin likeClass C: Metabotropic glutamate / pheromoneClass D: Fungal pheromoneClass E: cAMP receptors

Frizzled/Smoothened family

Putative families: * Ocular albinism proteins * Insect odorant receptors * Plant Mlo receptors * Nematode chemoreceptors * Vomeronasal receptors (V1R & V3R) * Taste receptors T2R

Orphans: * Putative / unclassified GPCRs

non-GPCR families: * Class Z: Archaeal/bacterial/fungal opsins

GPCR Classes* Class A Rhodopsin like o Amine o Peptide o Hormone protein o (Rhod)opsin o Olfactory o Prostanoid o Nucleotide-like

o Cannabinoid o Platelet activating factor o Gonadotropin-releasing hormone o Thyrotropin-releasing hormone & Secretagogue o Melatonin o Viral o Lysosphingolipid & LPA (EDG) o Leukotriene B4 receptor o Class A Orphan/other

* Class B Secretin like o Calcitonin o Corticotropin releasing factor o Gastric inhibitory peptide o Glucagon o Growth hormone-releasing hormone o Parathyroid hormone o PACAP o Secretin o Vasoactive intestinal polypeptide o Diuretic hormone o EMR1 o Latrophilin o Brain-specific angiogenesis inhibitor (BAI) o Methuselah-like proteins (MTH) o Cadherin EGF LAG (CELSR) o Very large G-protein coupled receptor

* Class C Metabotropic glutamate / pheromone o Metabotropic glutamate o Calcium-sensing like o Putative pheromone receptors o GABA-B o Orphan GPRC5 o Orphan GPCR6 o Bride of sevenless proteins (BOSS) o Taste receptors (T1R)

* Class D Fungal pheromone o Fungal pheromone A-Factor like (STE2,STE3) o Fungal pheromone B like (BAR,BBR,RCB,PRA) o Fungal pheromone M- and P-Factor

* Class E cAMP receptors

* Frizzled/Smoothened family o frizzled o Smoothened

Putative families:

* Ocular albinism proteins * Insect odorant receptors * Plant Mlo receptors * Nematode chemoreceptors * Vomeronasal receptors (V1R & V3R) * Taste receptors T2R

Orphans:

* Putative / unclassified GPCRs

non-GPCR families:

* Class Z Archaeal/bacterial/fungal opsins

GPCR Ligands Rhodopsin family: amine receptors Acetylcholine (muscarinic) Adrenaline Dopamine Histamine Serotonin Octopamine Trace amine

Rhodopsin family: peptide receptors Angiotensin Apelin Bombesin Bradykinin C5a anaphylatoxin CC Chemokine CXC Chemokine CX3C Chemokine C Chemokine Cholecystokinin Endothelin fMet-Leu-Phe Galanin Ghrelin KiSS1-derived peptide Melanocortin Motilin Neuromedin U Neuropeptide FF Neuropeptide S Neuropeptide Y Neuropeptide W / neuropeptide B Neurotensin Orexigenic neuropeptide QRFP Opioid Orexin Oxytocin Prokineticin Somatostatin Tachykinin Urotensin II Vasopressin Protease-activated (thrombin) Adrenomedullin (G10D) GPR37 / endothelin B like Chemokine receptor like Melanin-concentrating hormone Follicle stimulating hormone Lutropin-choriogonadotropic hormone Thyrotropin

Rhodopsin family: other receptors Rhodopsin Olfactory Prostaglandin Prostacyclin Thromboxane Adenosine Purine / pyrimidine Cannabinoid Platelet activating factor Gonadotropin-releasing hormone Thyrotropin-releasing hormone

Melatonin Lysosphingolipid and LPA (EDG) Leukotriene B4 receptor SREB Mas proto-oncogene & Mas-related (MRGs) RDC1 EBV-induced Relaxin LGR like Free fatty acid G protein-coupled bile acid Nicotinic acid GPR GPR45 like Cysteinyl leukotriene Putative / unclassified Class A GPCRs

Secretin family Calcitonin Corticotropin releasing factor Gastric inhibitory peptide Glucagon Growth hormone-releasing hormone Parathyroid hormone PACAP Secretin Vasoactive intestinal polypeptide EMR1 Latrophilin Brain-specific angiogenesis inhibitor (BAI) Methuselah-like proteins (MTH) Cadherin EGF LAG (CELSR) Putative / unclassified Class B GPCRs

Metabotropic glutamate family Glutamate (metabotropic) Extracellular calcium-sensing GABA-B Pheromone (V2R) Taste receptors (T1R) Orphan GPRC5 Orphan GPCR6 Bride of sevenless proteins (BOSS) Putative / unclassified Class C GPCRs

Other families Frizzled / Smoothened family Ocular albinism proteins Vomeronasal receptors (V1R) Taste receptors (T2R) Insect odorant receptors Nematode chemoreceptors Plant Mlo receptors Fungal pheromone cAMP (Dictyostelium) Bacterial rhodopsin

G protein-based disease

pituitary tumor

GHRH--Growth-hormone-releasing hormone

GH--Growth-hormone

GHRHGHRH

Pituitary

GHRH Receptor

GsαGsα(+)

cAMPcAMP GH secretionGH secretion(-)

somatostatinsomatostatin

GiGi

mechanism

Gs gene mutation

GTPase activity

Persistent activation of Gs

Persistent activation of AC

cAMP

Acromegaly or Gigantism

Pituitary proliferation and secretion

Cholera Toxin

cholera toxin enzyme that catalyzes the transfer of ADP ribose from

intracellular NAD+ to alpha s. The ADP ribosylation alters the alpha s so that it can no

longer hydrolyze its bound GTP. Thus, alpha s continues to stimulate adenylyl cyclase to produce cAMP.

The prolonged production of cAMP in the intestinal epithelial cells causes a large efflux of Na+ and water into the gut, and is responsible for the severe diarrhea that is characteristic of cholera.

Effect of choleratoxin

Persistent activation of adenylyl cyclase

Reseptor Asetilkolin Muskarinik

Nestler et al, 2001 Molecular Neuropharmaclogy

Reseptor Asetilkolin Muskarinik

Reseptor Adrenergik

Receptors and signal transduction in the ANS

Adrenergic Receptors

1A

1 2

1B 1D 2A 2B 2C 1 2 3

Direct acting adrenergic receptor agonists: 1 receptors

• Phenylephrine (Neosynephrine)

• Methoxamine (Vasoxyl)

• Oxymetazoline (Visine)

Phenyleph r in e

HO

CH CH2 NH CH3

OH

NH 3

COOH

G q

Phospho -lipase C

(+)

PIP 2

IP 3 Diacylglycerol

Increase Ca 2+ Activate ProteinKinase C

Response

Direct acting adrenergic receptor agonists: 2 receptors

• Clonidine (Catapres)• Methyldopa (Aldomet)• Guanabenz (Wytensin)• Guanfacine (Tenex)• Tizanidine (Zanaflex)

Clon id ine

NH 3

COOH

G I

(-)

ATP cAMP

Reduce cAMP -DependentProtein Kinase Activity

Response

X(+)K +

Adenylate Cyclase

N

NH

HN

Cl

Cl

Reseptor Dopamin

Nestler et al, 2001 Molecular Neuropharmaclogy,

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