Journal
CELLULAR SIGNALLING
Volume 19, Issue 8, Pages 1681-1689Publisher
ELSEVIER SCIENCE INC
DOI: 10.1016/j.cellsig.2007.03.004
Keywords
G alpha 12; G alpha 13; G protein; GAP; GEF; RGS protein; RhoA
Categories
Funding
- NIGMS NIH HHS [GM61454] Funding Source: Medline
- NINDS NIH HHS [NS41441] Funding Source: Medline
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G alpha 12 and G alpha 13 transduce signals from G protein-coupled receptors to RhoA through RhoGEFs containing an RGS homology (RH) domain, such as p115 RhoGEF or leukemia-associated RhoGEF (LARG). The RH domain of p115 RhoGEF or LARG binds with high affinity to active forms of G alpha 12 and G alpha 13 and confers specific GTPase-activating protein (GAP) activity, with faster GAP responses detected in G alpha 13 than in G alpha 12. At the same time, G alpha 13, but not G alpha 12, directly stimulates the RhoGEF activity of p115 RhoGEF or nonphosphorylated LARG in reconstitution assays. In order to better understand the molecular mechanism by which G alpha 13 regulates RhoGEF activity through interaction with RH-RhoGEFs, we sought to identify the region(s) of G alpha 13 involved in either the GAP response or RhoGEF activation. For this purpose, we generated chimeras between G alpha 12 and G alpha 13 subunits and characterized their biochemical activities. In both cell-based and reconstitution assays of RhoA activation, we found that replacing the carboxyl-terminal region of G alpha 12 (residues 267-379) with that of G alpha 13 (residues 264-377) conferred gain-of-function to the resulting chimeric subunit, G alpha 12C13. The inverse chimera, G alpha 13C12, exhibited basal RhoA activation which was similar to G alpha 12. In contrast to GEF assays, GAP assays showed that G alpha 12C13 or G alpha 13C12 chimeras responded to the GAP activity of p115 RhoGEF or LARG in a manner similar to G alpha 12 or G alpha 13, respectively. We conclude from these results that the carboxyl-terminal region of G alpha 13 (residues 264-377) is essential for its RhoGEF stimulating activity, whereas the amino-terminal a helical and switch regions of G alpha 12 and G alpha 13 are responsible for their differential GAP responses to the RH domain. (c) 2007 Elsevier Inc. All rights reserved.
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