Regulation of RGS-RhoGEFs by Galpha12 and Galpha13 proteins

Methods Enzymol. 2004;390:285-94. doi: 10.1016/S0076-6879(04)90018-3.

Abstract

Three mammalian Rho guanine nucleotide exchange factors (RhoGEFs), leukemia-associated RhoGEF (LARG), p115RhoGEF, and PDZ-RhoGEF, contain regulator of G-protein signaling (RGS) domains within their amino-terminal regions. These RhoGEFs link signals from heterotrimeric G12/13 protein-coupled receptors to Rho GTPase activation, leading to various cellular responses, such as actin reorganization and gene expression. The activity of these RhoGEFs is regulated by Galpha12/13 through their RGS domains. Because RhoGEFs stimulate guanine nucleotide exchange by Rho GTPases, RhoGEF activation can be measured by monitoring GTP binding to or GDP dissociation from Rho GTPases. This article describes methods used to perform reconstitution assays to measure the activity of RhoGEFs regulated by Galpha12/13.

MeSH terms

  • Animals
  • Cell Line
  • GTP-Binding Protein alpha Subunits, G12-G13 / isolation & purification
  • GTP-Binding Protein alpha Subunits, G12-G13 / metabolism*
  • Guanine Nucleotide Exchange Factors / isolation & purification
  • Guanine Nucleotide Exchange Factors / metabolism*
  • Guanosine 5'-O-(3-Thiotriphosphate) / metabolism
  • Guanosine Diphosphate / metabolism
  • Phosphorylation
  • RGS Proteins / metabolism*
  • Rho Guanine Nucleotide Exchange Factors
  • rhoA GTP-Binding Protein / isolation & purification
  • rhoA GTP-Binding Protein / metabolism

Substances

  • Guanine Nucleotide Exchange Factors
  • RGS Proteins
  • Rho Guanine Nucleotide Exchange Factors
  • Guanosine Diphosphate
  • Guanosine 5'-O-(3-Thiotriphosphate)
  • GTP-Binding Protein alpha Subunits, G12-G13
  • rhoA GTP-Binding Protein