Skip to content
Licensed Unlicensed Requires Authentication Published by De Gruyter January 10, 2007

Asef is a Cdc42-specific guanine nucleotide exchange factor

  • Katja Gotthardt and Mohammad Reza Ahmadian
From the journal Biological Chemistry


Asef is a member of the Dbl-family of guanine nucleotide exchange factors (GEFs) with a proposed specificity for the small GTPase Rac1. Here we investigated the specificity and regulation of Asef by measuring its GEF activity in vitro and observed hardly any activity towards Rac1, Rac2 and Rac3, or RhoA and TC10. In contrast, various purified Asef protein fragments catalyzed the nucleotide exchange reaction of Cdc42. The Cdc42GEF activity of the Dbl homology (DH) domain of Asef was significantly higher in the presence of the pleckstrin homology (PH) domain. Our data strongly suggest that Asef is a canonical Cdc42GEF, which employs its PH domain to efficiently stabilize its autoinhibited state, but also to facilitate nucleotide exchange activity of the DH domain after its activation by upstream signals.


Corresponding author


Bellanger, J.M., Astier, C., Sardet, C., Ohta, Y., Stossel, T.P., and Debant, A. (2000). The Rac1-and RhoG-specific GEF domain of Trio targets filamin to remodel cytoskeletal actin. Nat. Cell Biol.2, 888–892.10.1038/35046533Search in Google Scholar

Cherfils, J. and Chardin, P. (1999). GEFs: structural basis for their activation of small GTP-binding proteins. Trends Biochem. Sci.24, 306–311.10.1016/S0968-0004(99)01429-2Search in Google Scholar

Derewenda, U., Oleksy, A., Stevenson, A.S., Korczynska, J., Dauter, Z., Somlyo, A.P., Otlewski, J., Somlyo, A.V., and Derewenda, Z.S. (2004). The crystal structure of RhoA in complex with the DH/PH fragment of PDZRhoGEF, an activator of the Ca2+ sensitization pathway in smooth muscle. Structure12, 1955–1965.10.1016/j.str.2004.09.003Search in Google Scholar

Dvorsky, R. and Ahmadian, M.R. (2004). Always look on the bright site of Rho: structural implications for a conserved intermolecular interface. EMBO Rep.5, 1130–1136.10.1038/sj.embor.7400293Search in Google Scholar

Erickson, J.W. and Cerione, R.A. (2004). Structural elements, mechanism, and evolutionary convergence of Rho protein-guanine nucleotide exchange factor complexes. Biochemistry43, 837–842.10.1021/bi036026vSearch in Google Scholar

Etienne-Manneville, S. and Hall, A. (2002). Rho GTPases in cell biology. Nature420, 629–635.10.1038/nature01148Search in Google Scholar

Fiegen, D., Haeusler, L.C., Blumenstein, L., Herbrand, U., Dvorsky, R., Vetter, I.R., and Ahmadian, M.R. (2004). Alternative splicing of Rac1 generates Rac1b, a self-activating GTPase. J. Biol. Chem.279, 4743–4749.10.1074/jbc.M310281200Search in Google Scholar

Fukuhara, S., Murga, C., Zohar, M., Igishi, T., and Gutkind, J.S. (1999). A novel PDZ domain containing guanine nucleotide exchange factor links heterotrimeric G proteins to Rho. J. Biol. Chem.274, 5868–5879.10.1074/jbc.274.9.5868Search in Google Scholar

Fukuhara, S., Chikumi, H., and Gutkind, J.S. (2000). Leukemia-associated Rho guanine nucleotide exchange factor (LARG) links heterotrimeric G proteins of the G12 family to Rho. FEBS Lett.485, 183–188.10.1016/S0014-5793(00)02224-9Search in Google Scholar

Guo, Z., Ahmadian, M.R., and Goody, R.S. (2005). Guanine nucleotide exchange factors operate by a simple allosteric competitive mechanism. Biochemistry44, 15423–15429.10.1021/bi0518601Search in Google Scholar PubMed

Haeusler, L.C., Blumenstein, L., Stege, P., Dvorsky, R., and Ahmadian, M.R. (2003). Comparative functional analysis of the Rac GTPases. FEBS Lett.555, 556–560.10.1016/S0014-5793(03)01351-6Search in Google Scholar

Hart, M.J., Sharma, S., el Masry, N., Qiu, R.G., McCabe, P., Polakis, P., and Bollag, G. (1996). Identification of a novel guanine nucleotide exchange factor for the Rho GTPase. J. Biol. Chem.271, 25452–25458.10.1074/jbc.271.41.25452Search in Google Scholar

Hemsath, L. and Ahmadian, M.R. (2005). Fluorescence approaches for monitoring interactions of RhoGTPases with nucleotides, regulators and effectors. Methods37, 173–182.Search in Google Scholar

Hoffman, G.R. and Cerione, R.A. (2002). Signaling to the Rho GTPases: networking with the DH domain. FEBS Lett.513, 85–91.10.1016/S0014-5793(01)03310-5Search in Google Scholar

Karnoub, A.E., Worthylake, D.K., Rossman, K.L., Pruitt, W.M., Campbell, S.L., Sondek, J., and Der, C.J. (2001). Molecular basis for Rac1 recognition by guanine nucleotide exchange factors. Nat. Struct. Biol.8, 1037–1041.10.1038/nsb719Search in Google Scholar PubMed

Karnoub, A.E., Symons, M., Campbell, S.L., and Der, C.J. (2004). Molecular basis for Rho GTPase signaling specificity. Breast Cancer Res. Treat.84, 61–71.10.1023/B:BREA.0000018427.84929.5cSearch in Google Scholar

Kawasaki, Y., Senda, T., Ishidate, T., Koyama, R., Morishita, T., Iwayama, Y., Higuchi, O., and Akiyama, T. (2000). Asef, a link between the tumor suppressor APC and G-protein signaling. Science289, 1194–1197.10.1126/science.289.5482.1194Search in Google Scholar PubMed

Kawasaki, Y., Sato, R., and Akiyama, T. (2003). Mutated APC and Asef are involved in the migration of colorectal tumour cells. Nat. Cell Biol.5, 211–215.10.1038/ncb937Search in Google Scholar PubMed

Kozasa, T., Jiang, X., Hart, M.J., Sternweis, P.M., Singer, W.D., Gilman, A.G., Bollag, G., and Sternweis, P.C. (1998). p115 RhoGEF, a GTPase activating protein for Gα12 and Gα13. Science280, 2109–2111.10.1126/science.280.5372.2109Search in Google Scholar PubMed

Kristelly, R., Gao, G., and Tesmer, J.J.G. (2004). Structural determinants of RhoA binding and nucleotide exchange in leukemia-associated Rho guanine-nucleotide exchange factor. J. Biol. Chem.279, 47352–47362.10.1074/jbc.M406056200Search in Google Scholar PubMed

Movilla, N. and Bustelo, X.R. (1999). Biological and regulatory properties of Vav-3, a new member of the Vav family of oncoproteins. Mol. Cell. Biol.19, 7870–7885.10.1128/MCB.19.11.7870Search in Google Scholar PubMed PubMed Central

Noritake, J., Watanabe, T., Sato, K., Wang, S., and Kaibuchi, K. (2005). IQGAP1: A key regulator of adhesion and migration. J. Cell Sci.118, 2085–2092.10.1242/jcs.02379Search in Google Scholar PubMed

Raftopoulou, M. and Hall, A. (2004). Cell migration: Rho GTPases lead the way. Dev. Biol.265, 23–32.10.1016/j.ydbio.2003.06.003Search in Google Scholar PubMed

Reid, T., Bathoorn, A., Ahmadian, M.R., and Collard, J.G. (1999). Identification and characterization of hPEM-2, a guanine nucleotide exchange factor specific for Cdc42. J. Biol. Chem.274, 33587–33593.10.1074/jbc.274.47.33587Search in Google Scholar PubMed

Rossman, K.L. and Sondek, J. (2005). Larger than DbI: new structural insights into RhoA activation. Trends Biochem. Sci.30, 163–165.10.1016/j.tibs.2005.02.002Search in Google Scholar PubMed

Rossman, K.L., Worthylake, D.K., Snyder, J.T., Cheng, L., Whitehead, I.P., and Sondek, J. (2002a). Functional analysis of Cdc42 residues required for guanine nucleotide exchange. J. Biol. Chem.277, 50893–50898.10.1074/jbc.M208580200Search in Google Scholar PubMed

Rossman, K.L., Worthylake, D.K., Snyder, J.T., Siderovski, D.P., Campbell, S.L., and Sondek, J. (2002b). A crystallographic view of interactions between Dbs and Cdc42: PH domain-assisted guanine nucleotide exchange. EMBO J.21, 1315–1326.10.1093/emboj/21.6.1315Search in Google Scholar PubMed PubMed Central

Rossman, K.L., Cheng, L., Mahon, G.M., Rojas, R.J., Snyder, J.T., Whitehead, I.P., and Sondek, J. (2003). Multifunctional roles for the PH domain of Dbs in regulating rho GTPase activation. J. Biol. Chem.278, 18393–18400.10.1074/jbc.M300127200Search in Google Scholar PubMed

Rossman, K.L., Der, C.J., and Sondek, J. (2005). GEF means go: Turning on Rho GTPases with guanine nucleotide-exchange factors. Nat. Rev. Mol. Cell Biol.6, 167–180.10.1038/nrm1587Search in Google Scholar PubMed

Schiller, M.R. (2006). Coupling receptor tyrosine kinases to Rho GTPases–GEFs what's the link. Cell. Signal.18, 1834–1843.10.1016/j.cellsig.2006.01.022Search in Google Scholar PubMed

Schmidt, A. and Hall, A. (2002). Guanine nucleotide exchange factors for Rho GTPases: turning on the switch. Genes Dev.16, 1587–1609.10.1101/gad.1003302Search in Google Scholar PubMed

Snyder, J.T., Worthylake, D.K., Rossman, K.L., Betts, L., Pruitt, W.M., Siderovski, D.P., Der, C.J., and Sondek, J. (2002). Structural basis for the selective activation of Rho GTPases by Dbl exchange factors. Nat. Struct. Biol.9, 468–475.10.1038/nsb796Search in Google Scholar

Stam, J.C. and Collard, J.G. (1999). The DH protein family, exchange factors for Rho-like GTPases. Prog. Mol. Subcell. Biol.22, 51–83.10.1007/978-3-642-58591-3_4Search in Google Scholar

van Horck, F.P.G., Ahmadian, M.R., Haeusler, L.C., Moolenaar, W.H., and Kranenburg, O. (2001). Characterization of p190RhoGEF, a RhoA-specific guanine nucleotide exchange factor that interacts with microtubules. J. Biol. Chem.276, 4948–4956.10.1074/jbc.M003839200Search in Google Scholar

Vetter, I.R. and Wittinghofer, A. (2001). Signal transduction-the guanine nucleotide-binding switch in three dimensions. Science294, 1299–1304.10.1126/science.1062023Search in Google Scholar

Whitehead, I.P., Campbell, S., Rossman, K.L., and Der, C.J. (1997). Dbl family proteins. Biochim. Biophys. Acta1332, F1–F23.10.1016/S0304-419X(96)00040-6Search in Google Scholar

Whitehead, L.P., Lambert, Q.T., Glaven, J.A., Abe, K., Rossman, K.L., Mahon, G.M., Trzaskos, J.M., Kay, R., Campbell, S.L., and Der, C.J. (1999). Dependence of Dbl and Dbs transformation on MEK and NF-κB activation. Mol. Cell. Biol.19, 7759–7770.10.1128/MCB.19.11.7759Search in Google Scholar

Zheng, Y. (2001). Dbl family guanine nucleotide exchange factors. Trends Biochem. Sci.26, 724–732.10.1016/S0968-0004(01)01973-9Search in Google Scholar

Published Online: 2007-01-10
Published in Print: 2007-01-01

©2007 by Walter de Gruyter Berlin New York

Downloaded on 29.11.2023 from
Scroll to top button