title: Regulation of the G-protein regulatory-Gαi signaling complex by nonreceptor guanine nucleotide exchange factors.
abstract: Group II activators of G-protein signaling (AGS) serve as binding partners for Gα(i/o/t) via one or more G-protein regulatory (GPR) motifs. GPR-Gα signaling modules may be differentially regulated by cell surface receptors or by different nonreceptor guanine nucleotide exchange factors. We determined the effect of the nonreceptor guanine nucleotide exchange factors AGS1, GIV/Girdin, and Ric-8A on the interaction of two distinct GPR proteins, AGS3 and AGS4, with Gα(il) in the intact cell by bioluminescence resonance energy transfer (BRET) in human embryonic kidney 293 cells. AGS3-Rluc-Gα(i1)-YFP and AGS4-Rluc-Gα(i1)-YFP BRET were regulated by Ric-8A but not by Gα-interacting vesicle-associated protein (GIV) or AGS1. The Ric-8A regulation was biphasic and dependent upon the amount of Ric-8A and Gα(i1)-YFP. The inhibitory regulation of GPR-Gα(i1) BRET by Ric-8A was blocked by pertussis toxin. The enhancement of GPR-Gα(i1) BRET observed with Ric-8A was further augmented by pertussis toxin treatment. The regulation of GPR-Gα(i) interaction by Ric-8A was not altered by RGS4. AGS3-Rluc-Gα(i1)-YFP and AGS4-Rluc-G-Gα(i1)-YFP BRET were observed in both pellet and supernatant subcellular fractions and were regulated by Ric-8A in both fractions. The regulation of the GPR-Gα(i1) complex by Ric-8A, as well as the ability of Ric-8A to restore Gα expression in Ric8A(-/-) mouse embryonic stem cells, involved two helical domains at the carboxyl terminus of Ric-8A. These data indicate a dynamic interaction between GPR proteins, Gα(i1) and Ric-8A, in the cell that influences subcellular localization of the three proteins and regulates complex formation.
authors.affiliation: Department of Cell and Molecular Pharmacology and Experimental Therapeutics, Medical University of South Carolina, Charleston, South Carolina 29425, USA.
authors.forename: Sukru Sadik
authors.initials: SS
authors.lastname: Oner
authors.forename: Ellen M
authors.initials: EM
authors.lastname: Maher
authors.forename: Meital
authors.initials: M
authors.lastname: Gabay
authors.forename: Gregory G
authors.initials: GG
authors.lastname: Tall
authors.forename: Joe B
authors.initials: JB
authors.lastname: Blumer
authors.forename: Stephen M
authors.initials: SM
authors.lastname: Lanier
mesh_terms: D000818:Animals; D060807:Bioluminescence Resonance Energy Transfer Techniques; D002458:Cell Fractionation; D019206:GTP-Binding Protein alpha Subunits, Gi-Go / Q000378:metabolism*; D020662:Guanine Nucleotide Exchange Factors / Q000737:chemistry / Q000378:metabolism*; D057809:HEK293 Cells; D006801:Humans; D051379:Mice; D050505:Mutant Proteins / Q000378:metabolism; D009474:Neurons / Q000187:drug effects / Q000378:metabolism; D037342:Pertussis Toxin / Q000494:pharmacology; D020710:RGS Proteins / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects; D013347:Subcellular Fractions / Q000187:drug effects / Q000378:metabolism; D014162:Transfection; D033921:Vesicular Transport Proteins / Q000378:metabolism; D018631:ras Proteins / Q000378:metabolism
chemical_list: D020662:Guanine Nucleotide Exchange Factors; D050505:Mutant Proteins; D020710:RGS Proteins; D011993:Recombinant Fusion Proteins; C485884:Ric8a protein, mouse; D033921:Vesicular Transport Proteins; C098799:RGS4 protein; D037342:Pertussis Toxin; D019206:GTP-Binding Protein alpha Subunits, Gi-Go; D018631:ras Proteins