Eukaryotic Cell doi:10.1128/EC.00221-06
Copyright (c) 2006, American Society for Microbiology and/or the Listed Authors/Institutions. All Rights Reserved.
RacG regulates morphology, phagocytosis and chemotaxis
Baggavalli P. Somesh,
Georgia Vlahou,
Miho Iijima,
Robert H. Insall,
Peter Devreotes,
and
Francisco Rivero*
Center for Biochemistry and Center for Molecular Medicine Cologne, Medical Faculty, University of Cologne. Joseph-Stelzmann-Strasse 52, D-50931 Köln, Germany; School of Biosciences, The University of Birmingham, Edgbaston, Birmingham, B15 2TT, UK; Department of Cell Biology, Johns Hopkins University School of Medicine, 725 N. Wolfe St., Baltimore MD, 21205, USA
* To whom correspondence should be addressed. Email:
francisco.rivero{at}uni-koeln.de.
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Abstract |
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RacG is an unusual member of the complex family of Rho GTPses in Dictyostelium. We have generated a knockout strain (KO) as well as strains that overexpress the wild-type (WT), constitutively active (V12) or dominant negative (N17) RacG. The protein is targeted to the plasma membrane, apparently in a nucleotide-dependent manner, and induces the formation of abundant actin-driven filopods. RacG is enriched at the rim of the progressing phagocytic cup and overexpression of RacG-WT or RacG-V12 induced an increased rate of particle uptake. The positive effect of RacG on phagocytosis was abolished in the presence of 50 µM LY294002, a phosphoinositide 3-kinase inhibitor, indicating that generation of phosphatidylinositol 3,4,5-trisphosphate is required for activation of RacG. RacG-KO cells showed a moderate chemotaxis defect that was stronger in RacG-V12 and RacG-N17 mutants, in part due to interference with signaling through Rac1. The in vivo effects of RacG-V12 could not be reproduced by a mutant lacking the Rho insert region, indicating that this region is essential for interaction with downstream components. Processes like growth, pinocytosis, exocytosis, cytokinesis and development were unaffected in Rac-KO cells and in the overexpressor mutants. In a cell-free system RacG induced actin polymerization upon GTP
S stimulation, and this response could be blocked by an Arp3 antibody. While the mild phenotype of RacG-KO cells indicates some overlap with one or more Dictyostelium Rho GTPases, like Rac1 and RacB, the significant changes found in overexpressors show RacG plays important roles. We hypothesize that RacG interacts with a subset of effectors, in particular those concerned with shape, motility and phagocytosis.
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