Modulation of subfamily B/R4 RGS protein function by 14-3-3 proteins.
Regulator of G protein signalling (RGS) proteins are primarily known for their ability to act as GTPase activating proteins (GAPs) and thus attenuate G protein function within G protein-coupled receptor (GPCR) signalling pathways. However, RGS proteins have been found to interact with additional binding partners, and this has introduced more complexity to our understanding of their potential role in vivo. Here, we identify a novel interaction between RGS proteins (RGS4, RGS5, RGS16) and the multifunctional protein 14-3-3. Two isoforms, 14-3-3beta and 14-3-3epsilon, directly interact with all three purified RGS proteins and data from in vitro steady state GTP hydrolysis assays show that 14-3-3 inhibits the GTPase activity of RGS4 and RGS16, but has limited effects on RGS5 under comparable conditions. Moreover in a competitive pull-down experiment, 14-3-3epsilon competes with Galphao for RGS4, but not for RGS5. This mechanism is further reinforced in living cells, where 14-3-3epsilon sequesters RGS4 in the cytoplasm and impedes its recruitment to the plasma membrane by Galpha protein. Thus, 14-3-3 might act as a molecular chelator, preventing RGS proteins from interacting with Galpha, and ultimately prolonging the signal transduction pathway. In conclusion, our findings suggest that 14-3-3 proteins may indirectly promote GPCR signalling via their inhibitory effects on RGS GAP function.
Abramow-Newerly
Maria
M
Department of Physiology and Pharmacology, University of Western Ontario, London, Ontario, Canada N6A 5C1.
Ming
Hong
H
Chidiac
Peter
P
D048948:14-3-3 Proteins / Q000235:genetics / Q000378:metabolism*; D001665:Binding Sites; D001667:Binding, Competitive; D002457:Cell Extracts / Q000737:chemistry; D002460:Cell Line; D019204:GTP-Binding Proteins / Q000235:genetics / Q000378:metabolism; D020690:GTPase-Activating Proteins / Q000378:metabolism; D005982:Glutathione Transferase / Q000235:genetics / Q000378:metabolism; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D006160:Guanosine Triphosphate / Q000378:metabolism; D006801:Humans; D006868:Hydrolysis; D015151:Immunoblotting; D007700:Kinetics; D007930:Leucine / Q000378:metabolism; D018613:Microscopy, Confocal; D011485:Protein Binding / Q000502:physiology; D020033:Protein Isoforms / Q000235:genetics / Q000378:metabolism; D020710:RGS Proteins / Q000235:genetics / Q000378:metabolism* / Q000502:physiology; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D014162:Transfection; D014443:Tyrosine / Q000378:metabolism
D048948:14-3-3 Proteins; D002457:Cell Extracts; D020690:GTPase-Activating Proteins; D020033:Protein Isoforms; D020710:RGS Proteins; D011993:Recombinant Fusion Proteins; D049452:Green Fluorescent Proteins; D014443:Tyrosine; D006160:Guanosine Triphosphate; D005982:Glutathione Transferase; D019204:GTP-Binding Proteins; D007930:Leucine