title: Biochemical characterization of RGS14: RGS14 activity towards G-protein alpha subunits is independent of its binding to Rap2A.
abstract: RGS (regulators of G-protein signalling) modulate signalling by acting as GAPs (GTPase-activating proteins) for alpha subunits of heterotrimeric G-proteins. RGS14 accelerates GTP hydrolysis by G(ialpha) family members through its RGS domain and suppresses guanine nucleotide dissociation from G(ialpha1) and G(ialpha3) subunits through its C-terminal GoLoco domain. Additionally, RGS14 binds the activated forms of the small GTPases Rap1 and Rap2 by virtue of tandem RBDs (Raf-like Ras/Rap binding domains). RGS14 was identified in a screen for Rap2 effectors [Traver, Splingard, Gaudriault and De Gunzburg (2004) Biochem. J. 379, 627-632]. In the present study, we tested whether Rap binding regulates RGS14's biochemical activities. We found that RGS14 activity towards heterotrimeric G-proteins, as either a GAP or a GDI (guanine nucleotide dissociation inhibitor), was unaffected by Rap binding. Extending our biochemical characterization of RGS14, we also examined whether RGS14 can suppress guanine nucleotide exchange on G(ialpha1) in the context of the heterotrimer. We found that a heterotrimer composed of N-myristoylated G(ialpha1) and prenylated G(betagamma) is resistant to the GDI activity of the GoLoco domain of RGS14. This is consistent with models of GoLoco domain action on free G(alpha) and suggests that RGS14 alone cannot induce subunit dissociation to promote receptor-independent activation of G(betagamma)-mediated signalling pathways.
authors.affiliation: Department of Cell Biology and Physiology, Washington University School of Medicine, 660 South Euclid Ave., Campus Box 8228, St. Louis, MO 63110, USA.
authors.forename: Vivek
authors.initials: V
authors.lastname: Mittal
authors.forename: Maurine E
authors.initials: ME
authors.lastname: Linder
mesh_terms: D000818:Animals; D002460:Cell Line; D044385:GTP-Binding Protein alpha Subunits / Q000378:metabolism*; D044387:GTP-Binding Protein beta Subunits / Q000378:metabolism; D044388:GTP-Binding Protein gamma Subunits / Q000378:metabolism; D020690:GTPase-Activating Proteins / Q000378:metabolism; D006801:Humans; D007313:Insecta; D051379:Mice; D011485:Protein Binding; D020710:RGS Proteins / Q000378:metabolism*; D015398:Signal Transduction; D020668:rap GTP-Binding Proteins / Q000378:metabolism*
chemical_list: C118981:G-protein Beta gamma; D044385:GTP-Binding Protein alpha Subunits; D044387:GTP-Binding Protein beta Subunits; D044388:GTP-Binding Protein gamma Subunits; D020690:GTPase-Activating Proteins; D020710:RGS Proteins; C491399:Rgs14 protein, mouse; C116756:RAP2A protein, human; D020668:rap GTP-Binding Proteins