Activation of p115-RhoGEF requires direct association of Gα13and the Dbl homology domain

Zhe Chen, Liang Guo, Jana Hadas, Stephen Gutowski, Stephen R. Sprang, Paul C. Sternweis

Research output: Contribution to journalArticle

27 Scopus citations

Abstract

RGS-containing RhoGEFs (RGS-RhoGEFs) represent a direct link between the G12 class of heterotrimeric G proteins and the monomeric GTPases. In addition to the canonical Dbl homology (DH) and pleckstrin homology domains that carry out the guanine nucleotide exchange factor (GEF) activity toward RhoA, these RhoGEFs also possess RGS homology (RH) domains that interact with activated α subunits of G12 and G13. Although the GEF activity of p115-RhoGEF (p115), an RGS-RhoGEF, can be stimulated by Gα13, the exact mechanism of the stimulation has remained unclear. Using combined studies with small angle x-ray scattering, biochemistry, and mutagenesis, we identify an additional binding site for activated Gα13 in the DH domain of p115. Small angle x-ray scattering reveals that the helical domain of Gα13 docks onto the DH domain, opposite to the surface of DH that binds RhoA. Mutation of a single tryptophan residue in the α3b helix of DH reduces binding to activated Gα13 and ablates the stimulation of p115 by Gα13. Complementary mutations at the predicted DH-binding site in the αB-αC loop of the helical domain of Gα13 also affect stimulation of p115 by Gα13. Although the GAP activity of p115 is not required for stimulation by Gα13, two hydrophobic motifs in RH outside of the consensus RGS box are critical for this process. Therefore, the binding of Gα13 to the RH domain facilitates direct association of Gα13 to theDHdomain to regulate its exchange activity. This study provides new insight into the mechanism of regulation of the RGS-RhoGEF and broadens our understanding of G protein signaling.

Original languageEnglish (US)
Pages (from-to)25490-25500
Number of pages11
JournalJournal of Biological Chemistry
Volume287
Issue number30
DOIs
StatePublished - Jul 20 2012

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ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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