Agonist selective regulation of G proteins by cannabinoid CB(1) and CB(2) receptors.
We have examined the ligand regulation and G protein selectivity of the human cannabinoid CB(1) and CB(2) receptors by an in situ reconstitution technique directly measuring G protein activation. Membranes from Spodoptera frugiperda cells expressing CB(1) and CB(2) receptors were chaotrope extracted to denature endogenous GTP-binding proteins. The ability of the receptors to catalyze the GDP-GTP exchange of each G protein was then examined with purified bovine brain G(i) and G(o). Activation of CB(1) receptors produced a high-affinity saturable interaction for both G(i) and G(o). Agonist stimulation of CB(2) receptors also resulted in a high-affinity saturable interaction with G(i). In contrast, CB(2) receptors did not interact efficiently with G(o). G protein activation was then examined with a diverse group of ligands. For the interaction of CB(2) receptors with G(i), HU210 was the only compound tested that demonstrated maximal activation. In contrast, WIN55,212 (64%), anandamide (42%), and Delta(9)-tetrahydrocannabinol (Delta(9)-THC) (44%) all initiated submaximal levels of G protein activation. For CB(1) receptor-catalyzed activation of G(i), HU210, WIN55,212, and anandamide all elicited maximal activation, whereas Delta(9)-THC (56 +/- 6%) caused only partial G(i) activation. In contrast, only HU210 effected maximal CB(1) stimulation of G(o), with anandamide, WIN55, 212, and Delta(9)-THC all stimulating between 60 and 75% compared with HU210. These data demonstrate that different agonists induce different conformations of the CB(1) receptor, which in turn can distinguish between different G proteins. Our data thus demonstrate agonist-selective G protein signaling by the CB(1) receptor and suggest that therapeutic agents may be designed to regulate individual G protein-signaling pathways selectively.
Glass
M
M
Section on Signal Transduction, National Institutes of Health, National Institute on Deafness and Other Communication Disorders, Rockville, Maryland 20850, USA.
Northup
J K
JK
D000818:Animals; D002186:Cannabinoids / Q000378:metabolism; D002417:Cattle; D019206:GTP-Binding Protein alpha Subunits, Gi-Go / Q000819:agonists / Q000378:metabolism*; D016244:Guanosine 5'-O-(3-Thiotriphosphate) / Q000378:metabolism; D020962:Heterotrimeric GTP-Binding Proteins / Q000819:agonists / Q000378:metabolism*; D006801:Humans; D010880:Piperidines / Q000494:pharmacology; D011720:Pyrazoles / Q000494:pharmacology; D043882:Receptors, Cannabinoid; D011955:Receptors, Drug / Q000819:agonists / Q000037:antagonists & inhibitors / Q000378:metabolism*; D011994:Recombinant Proteins / Q000378:metabolism; D000077285:Rimonabant; D018411:Spodoptera / Q000166:cytology; D013462:Sulfur Radioisotopes; D014162:Transfection; D010566:Virulence Factors, Bordetella / Q000494:pharmacology
D002186:Cannabinoids; D010880:Piperidines; D011720:Pyrazoles; D043882:Receptors, Cannabinoid; D011955:Receptors, Drug; D011994:Recombinant Proteins; D013462:Sulfur Radioisotopes; D010566:Virulence Factors, Bordetella; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D019206:GTP-Binding Protein alpha Subunits, Gi-Go; D020962:Heterotrimeric GTP-Binding Proteins; D000077285:Rimonabant