Regulation of the AGS3·G{alpha}i signaling complex by a seven-transmembrane span receptor.
G-protein signaling modulators (GPSM) play diverse functional roles through their interaction with G-protein subunits. AGS3 (GPSM1) contains four G-protein regulatory motifs (GPR) that directly bind Gα(i) free of Gβγ providing an unusual scaffold for the "G-switch" and signaling complexes, but the mechanism by which signals track into this scaffold are not well understood. We report the regulation of the AGS3·Gα(i) signaling module by a cell surface, seven-transmembrane receptor. AGS3 and Gα(i1) tagged with Renilla luciferase or yellow fluorescent protein expressed in mammalian cells exhibited saturable, specific bioluminescence resonance energy transfer indicating complex formation in the cell. Activation of α(2)-adrenergic receptors or μ-opioid receptors reduced AGS3-RLuc·Gα(i1)-YFP energy transfer by over 30%. The agonist-mediated effects were inhibited by pertussis toxin and co-expression of RGS4, but were not altered by Gβγ sequestration with the carboxyl terminus of GRK2. Gα(i)-dependent and agonist-sensitive bioluminescence resonance energy transfer was also observed between AGS3 and cell-surface receptors typically coupled to Gα(i) and/or Gα(o) indicating that AGS3 is part of a larger signaling complex. Upon receptor activation, AGS3 reversibly dissociates from this complex at the cell cortex. Receptor coupling to both Gαβγ and GPR-Gα(i) offer additional flexibility for systems to respond and adapt to challenges and orchestrate complex behaviors.
Oner
Sukru Sadik
SS
Department of Cell and Molecular Pharmacology & Experimental Therapeutics, Medical University of South Carolina, Charleston, South Carolina 29425, USA.
An
Ningfei
N
Vural
Ali
A
Breton
Billy
B
Bouvier
Michel
M
Blumer
Joe B
JB
Lanier
Stephen M
SM
D000818:Animals; D002352:Carrier Proteins / Q000737:chemistry / Q000378:metabolism*; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism*; D019206:GTP-Binding Protein alpha Subunits, Gi-Go / Q000737:chemistry / Q000378:metabolism*; D005786:Gene Expression Regulation; D020730:Guanine Nucleotide Dissociation Inhibitors; D006801:Humans; D008565:Membrane Proteins / Q000378:metabolism; D008954:Models, Biological; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D018341:Receptors, Adrenergic, alpha-2 / Q000737:chemistry / Q000378:metabolism; D017450:Receptors, Opioid, mu / Q000378:metabolism; D048790:Renilla; D015398:Signal Transduction
D002352:Carrier Proteins; C525615:GPSM1 protein, human; D020730:Guanine Nucleotide Dissociation Inhibitors; D008565:Membrane Proteins; D018341:Receptors, Adrenergic, alpha-2; D017450:Receptors, Opioid, mu; D019206:GTP-Binding Protein alpha Subunits, Gi-Go