HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS		QUICK SEARCH:   [advanced] Author: Keyword(s): Year:  Vol:  Page:

J. Biol. Chem., Vol. 282, Issue 41, 29967-29976, October 12, 2007

This Article Full Text Full Text (PDF) Supplemental Data All Versions of this Article: 282/41/29967    most recent M701877200v1 Submit a Letter to Editor Alert me when this article is cited Alert me when eLetters are posted Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Request Permissions Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Barber, M. A. Articles by Welch, H. C. E. Search for Related Content PubMed PubMed Citation Articles by Barber, M. A. Articles by Welch, H. C. E. Social Bookmarking                      What's this?

Membrane Translocation of P-Rex1 Is Mediated by G Protein Subunits and Phosphoinositide 3-Kinase^*

Mark A. Barber¹, Sarah Donald, Sylvia Thelen, Karen E. Anderson, Marcus Thelen, and Heidi C. E. Welch, Recipient of a Medical Research Council Career Development Award²

From the Inositide Laboratory, Babraham Institute, Babraham Research Campus, Cambridge CB22 3AT, United Kingdom and the Institute for Research in Biomedicine, Via Vela 6, 6500 Bellinzona, Switzerland

P-Rex1 is a guanine-nucleotide exchange factor (GEF) for the small GTPase Rac that is directly activated by the subunits of heterotrimeric G proteins and by the lipid second messenger phosphatidylinositol (3,4,5)-trisphosphate (PIP₃), which is generated by phosphoinositide 3-kinase (PI3K). G subunits and PIP₃ are membrane-bound, whereas the intracellular localization of P-Rex1 in basal cells is cytosolic. Activation of PI3K alone is not sufficient to promote significant membrane translocation of P-Rex1. Here we investigated the subcellular localization of P-Rex1 by fractionation of Sf9 cells co-expressing P-Rex1 with G and/or PI3K. In basal, serum-starved cells, P-Rex1 was mainly cytosolic, but 7% of the total was present in the 117,000 x g membrane fraction. Co-expression of P-Rex1 with either G or PI3K caused only an insignificant increase in P-Rex1 membrane localization, whereas G and PI3K together synergistically caused a robust increase in membrane-localized P-Rex1 to 23% of the total. PI3K-driven P-Rex1 membrane recruitment was wortmannin-sensitive. The use of P-Rex1 mutants showed that the isolated Dbl homology/pleckstrin homology domain tandem of P-Rex1 is sufficient for synergistic G- and PI3K-driven membrane localization; that the enzymatic GEF activity of P-Rex1 is not required for membrane translocation; and that the other domains of P-Rex1 (DEP, PDZ, and IP4P) contribute to keeping the enzyme localized in the cytosol of basal cells. In vitro Rac2-GEF activity assays showed that membrane-derived purified P-Rex1 has a higher basal activity than cytosol-derived P-Rex1, but both can be further activated by PIP₃ and G subunits.

Received for publication, March 5, 2007 , and in revised form, July 17, 2007.

^* This work was supported in part by British Biological Sciences Research Council Grant C19943. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

The on-line version of this article (available at http://www.jbc.org) contains supplemental Videos 1–3.

¹ A Cambridge Commonwealth Trust postgraduate fellow.

² To whom correspondence should be addressed. Tel.: 44-1223-496-596; Fax: 44-1223-496-043; E-mail: heidi.welch{at}bbsrc.ac.uk.

CiteULike    Complore    Connotea    Del.icio.us    Digg    Reddit    Technorati    What's this?

This article has been cited by other articles:

				A. Garcia-Regalado, M. L. Guzman-Hernandez, I. Ramirez-Rangel, E. Robles-Molina, T. Balla, J. Vazquez-Prado, and G. Reyes-Cruz G Protein-coupled Receptor-promoted Trafficking of G{beta}1{gamma}2 Leads to AKT Activation at Endosomes via a Mechanism Mediated by G{beta}1{gamma}2-Rab11a Interaction Mol. Biol. Cell, October 1, 2008; 19(10): 4188 - 4200. [Abstract] [Full Text] [PDF]

				J. E. Waters, M. V. Astle, L. M. Ooms, D. Balamatsias, R. Gurung, and C. A. Mitchell P-Rex1 - a multidomain protein that regulates neurite differentiation J. Cell Sci., September 1, 2008; 121(17): 2892 - 2903. [Abstract] [Full Text] [PDF]

				Y. Ugolev, Y. Berdichevsky, C. Weinbaum, and E. Pick Dissociation of Rac1(GDP){middle dot}RhoGDI Complexes by the Cooperative Action of Anionic Liposomes Containing Phosphatidylinositol 3,4,5-Trisphosphate, Rac Guanine Nucleotide Exchange Factor, and GTP J. Biol. Chem., August 8, 2008; 283(32): 22257 - 22271. [Abstract] [Full Text] [PDF]