title: The second intracellular loop of the human cannabinoid CB2 receptor governs G protein coupling in coordination with the carboxyl terminal domain.
abstract: The major effects of cannabinoids and endocannabinoids are mediated via two G protein-coupled receptors, CB1 and CB2, elucidation of the mechanism and structural determinants of the CB2 receptor coupling with G proteins will have a significant impact on drug discovery. In the present study, we systematically investigated the role of the intracellular loops in the interaction of the CB2 receptor with G proteins using chimeric receptors alongside the characterization of cAMP accumulation and ERK1/2 phosphorylation. We provided evidence that ICL2 was significantly involved in G protein coupling in coordination with the C-terminal end. Moreover, a single alanine substitution of the Pro-139 in the CB2 receptor that corresponds to Leu-222 in the CB1 receptor resulted in a moderate impairment in the inhibition of cAMP accumulation, whereas mutants P139F, P139M and P139L were able to couple to the Gs protein in a CRE-driven luciferase assay. With the ERK activation experiments, we further found that P139L has the ability to activate ERK through both Gi- and Gs-mediated pathways. Our findings defined an essential role of the second intracellular loop of the CB2 receptor in coordination with the C-terminal tail in G protein coupling and receptor activation.
authors.affiliation: Institute of Biochemistry, College of Life Sciences, Zhejiang University, Hangzhou, Zhejiang, China.
authors.forename: Congxia
authors.initials: C
authors.lastname: Zheng
authors.forename: Linjie
authors.initials: L
authors.lastname: Chen
authors.forename: Xiaopan
authors.initials: X
authors.lastname: Chen
authors.forename: Xiaobai
authors.initials: X
authors.lastname: He
authors.forename: Jingwen
authors.initials: J
authors.lastname: Yang
authors.forename: Ying
authors.initials: Y
authors.lastname: Shi
authors.forename: Naiming
authors.initials: N
authors.lastname: Zhou
mesh_terms: D000067956:Adenylyl Cyclase Inhibitors; D000262:Adenylyl Cyclases / Q000378:metabolism; D000595:Amino Acid Sequence; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism; D017868:Cyclic AMP-Dependent Protein Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism; D004789:Enzyme Activation / Q000187:drug effects; D048049:Extracellular Signal-Regulated MAP Kinases / Q000378:metabolism; D019204:GTP-Binding Proteins / Q000378:metabolism*; D057809:HEK293 Cells; D006801:Humans; D008969:Molecular Sequence Data; D050505:Mutant Proteins / Q000737:chemistry / Q000378:metabolism; D011392:Proline / Q000378:metabolism; D011485:Protein Binding / Q000187:drug effects; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D043884:Receptor, Cannabinoid, CB1 / Q000737:chemistry / Q000378:metabolism; D043885:Receptor, Cannabinoid, CB2 / Q000819:agonists / Q000737:chemistry* / Q000378:metabolism*; D011994:Recombinant Proteins / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects; D013329:Structure-Activity Relationship
chemical_list: D000067956:Adenylyl Cyclase Inhibitors; D050505:Mutant Proteins; D047428:Protein Kinase Inhibitors; D043884:Receptor, Cannabinoid, CB1; D043885:Receptor, Cannabinoid, CB2; D011994:Recombinant Proteins; D011392:Proline; D017868:Cyclic AMP-Dependent Protein Kinases; D048049:Extracellular Signal-Regulated MAP Kinases; D019204:GTP-Binding Proteins; D000262:Adenylyl Cyclases