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Divergence of Rho residue 43 impacts GEF activity
Journal article   Open access   Peer reviewed

Divergence of Rho residue 43 impacts GEF activity

Christina M Sloan, Clancy V Quinn, Justin P Peters, Janean Farley, Chris Goetzinger, Molly Wernli, Kris A DeMali and Shawn M Ellerbroek
Small GTPases, Vol.3(1), pp.15-22
01/2012
DOI: 10.4161/sgtp.19557
PMCID: PMC3398911
PMID: 22673745
url
https://doi.org/10.4161/sgtp.19557View
Published (Version of record) Open Access

Abstract

RhoA, RhoB and RhoC GTPases are over 85% identical at the amino acid level, with RhoA and RhoC differing at only one residue (43) across the initial two-thirds of their sequences. A putative regulatory distinction between the molecules is their capacity to be uniquely activated by guanine nucleotide exchange factors (GEFs). We hypothesize that variation of amino acid residue 43 between RhoA/B (valine) and RhoC (isoleucine) impacts GEF activity. Direct participation of residue 43 in GEF-catalyzed exchange was confirmed by the observation that mutation of this position to a threonine reduced GEF-catalyzed nucleotide exchange activity in vitro (Vav2, XPLN, GEFT, Dbl and Dbs) and greatly depressed RhoA and RhoC GTP-loading profiles in cell lysates. Using a residue swap approach, substitution of RhoA Val 43 with an Ile was found to significantly promote basal nucleotide exchange activity and enhance GTP-loading in cells. Substitution of Val 43 with an Ile in RhoB negatively affected nucleotide exchange in vitro. Substitution of RhoC Ile 43 with a Val increased GEF-catalyzed exchange in vitro. In addition, RhoC-I43V was more efficacious at driving ovarian cancer cell invasion through matrigrel than wild-type RhoC, RhoC-I43T, wild-type RhoA, RhoA-V43I or RhoA-V43T GTPases. These findings suggest that a divergence between RhoA/B and RhoC at residue 43 impacts basal and GEF-stimulated nucleotide exchange activity.
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