The N-terminal non-RGS domain of human regulator of G-protein signalling 1 contributes to its ability to inhibit pheromone receptor signalling in yeast.
Regulators of G-protein signalling (RGS) are a family of proteins that interact with G-proteins to regulate negatively G-protein coupled receptor (GPCR) signalling. In addition to a conserved core domain that is necessary and sufficient for their GTPase activating protein (GAP) like activity, RGSs possess N- and C-terminal motifs that confer distinct functional differences. In order to identify the role of the non-RGS region of human RGS1, we have characterized a series of fusions between RGS1 and GFP in a yeast mutant lacking the RGS containing SST2 gene. Using both halo assays as well as a GPCR responsive FUS1-LacZ reporter gene, we demonstrate that a RGS1-GFP fusion inhibits GPCR signalling in yeast while GFP fusions containing either the N-terminus non RGS sequence of RGS1(1-68) or the sequence containing the RGS box of RGS1(68-197) produce proteins that retain RGS1 activity. These results suggest that both the N-terminal and the RGS box of RGS1 function to inhibit signalling. Analysis of a series of mutants spanning the entire N-terminal non-RGS region of RGS1 produced by conservative segment exchange (CSE) mutagenesis showed little loss of function in yeast. This suggests that the overall structure of the N-terminal region of RGS1 rather than specific motifs or residues is required for its function.
Somerville
Wendy
W
Department of Anatomy and Cell Biology, McGill University, Montreal, Quebec, Canada.
Song
Wei
W
Kong
Janice L
JL
Panetta
Rosemarie
R
Greenwood
Michael T
MT
D000595:Amino Acid Sequence; D020555:GTP-Binding Protein Regulators / Q000235:genetics* / Q000378:metabolism; D019204:GTP-Binding Proteins / Q000378:metabolism*; D020690:GTPase-Activating Proteins / Q000235:genetics; D017930:Genes, Reporter; D006801:Humans; D008969:Molecular Sequence Data; D009154:Mutation; D010675:Pheromones / Q000378:metabolism*; D010957:Plasmids; D012441:Saccharomyces cerevisiae / Q000235:genetics* / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics; D015398:Signal Transduction / Q000502:physiology*; D020710:RGS Proteins
D020555:GTP-Binding Protein Regulators; D019204:GTP-Binding Proteins; D020690:GTPase-Activating Proteins; D010675:Pheromones; D029701:Saccharomyces cerevisiae Proteins; C094301:SST2 protein, S cerevisiae; D020710:RGS Proteins