title: RGS12 and RGS14 GoLoco motifs are G alpha(i) interaction sites with guanine nucleotide dissociation inhibitor Activity.
abstract: The regulators of G-protein signaling (RGS) proteins accelerate the intrinsic guanosine triphosphatase activity of heterotrimeric G-protein alpha subunits and are thus recognized as key modulators of G-protein-coupled receptor signaling. RGS12 and RGS14 contain not only the hallmark RGS box responsible for GTPase-accelerating activity but also a single G alpha(i/o)-Loco (GoLoco) motif predicted to represent a second G alpha interaction site. Here, we describe functional characterization of the GoLoco motif regions of RGS12 and RGS14. Both regions interact exclusively with G alpha(i1), G alpha(i2), and G alpha(i3) in their GDP-bound forms. In GTP gamma S binding assays, both regions exhibit guanine nucleotide dissociation inhibitor (GDI) activity, inhibiting the rate of exchange of GDP for GTP by G alpha(i1). Both regions also stabilize G alpha(i1) in its GDP-bound form, inhibiting the increase in intrinsic tryptophan fluorescence stimulated by AlF(4)(-). Our results indicate that both RGS12 and RGS14 harbor two distinctly different G alpha interaction sites: a previously recognized N-terminal RGS box possessing G alpha(i/o) GAP activity and a C-terminal GoLoco region exhibiting G alpha(i) GDI activity. The presence of two, independent G alpha interaction sites suggests that RGS12 and RGS14 participate in a complex coordination of G-protein signaling beyond simple G alpha GAP activity.
authors.affiliation: Department of Pharmacology, Lineberger Comprehensive Cancer Center, University of North Carolina, Chapel Hill, North Carolina 27599-7365, USA.
authors.forename: R J
authors.initials: RJ
authors.lastname: Kimple
authors.forename: L
authors.initials: L
authors.lastname: De Vries
authors.forename: H
authors.initials: H
authors.lastname: Tronchère
authors.forename: C I
authors.initials: CI
authors.lastname: Behe
authors.forename: R A
authors.initials: RA
authors.lastname: Morris
authors.forename: M
authors.initials: M
authors.lastname: Gist Farquhar
authors.forename: D P
authors.initials: DP
authors.lastname: Siderovski
mesh_terms: D017607:Aluminum Compounds / Q000494:pharmacology; D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D000818:Animals; D001665:Binding Sites; D015374:Biosensing Techniques; D003001:Cloning, Molecular; D004926:Escherichia coli; D005459:Fluorides / Q000494:pharmacology; D016244:Guanosine 5'-O-(3-Thiotriphosphate) / Q000378:metabolism*; D006153:Guanosine Diphosphate / Q000378:metabolism; D020962:Heterotrimeric GTP-Binding Proteins / Q000737:chemistry / Q000378:metabolism*; D007700:Kinetics; D008954:Models, Biological; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009842:Oligopeptides; D016366:Open Reading Frames; D020710:RGS Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D051381:Rats; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D015398:Signal Transduction; D020349:Surface Plasmon Resonance
chemical_list: D017607:Aluminum Compounds; D009842:Oligopeptides; D020710:RGS Proteins; C491400:RGS14 protein, human; D011994:Recombinant Proteins; C491401:Rgs12 protein, rat; C106374:Rgs14 protein, rat; C097755:arginyl-glycyl-serine; D006153:Guanosine Diphosphate; C050992:tetrafluoroaluminate; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D020962:Heterotrimeric GTP-Binding Proteins; D005459:Fluorides