title: Identification of Rab11 as a small GTPase binding protein for the Evi5 oncogene.
abstract: The Evi5 oncogene has recently been shown to regulate the stability and accumulation of critical G(1) cell cycle factors including Emi1, an inhibitor of the anaphase-promoting complex/cyclosome, and cyclin A. Sequence analysis of the amino terminus of Evi5 reveals a Tre-2, Bub2, Cdc16 domain, which has been shown to be a binding partner and GTPase-activating protein domain for the Rab family of small Ras-like GTPases. Here we describe the identification of Evi5 as a candidate binding protein for Rab11, a GTPase that regulates intracellular transport and has specific roles in endosome recycling and cytokinesis. By yeast two-hybrid analysis, immunoprecipitation, and Biacore analysis, we demonstrate that Evi5 binds Rab11a and Rab11b in a GTP-dependent manner. However, Evi5 displays no activation of Rab11 GTPase activity in vitro. Evi5 colocalizes with Rab11 in vivo, and overexpression of Rab11 perturbs the localization of Evi5, redistributing it into Rab11-positive recycling endosomes. Interestingly, in vitro binding studies show that Rab11 effector proteins including FIP3 compete with Evi5 for binding to Rab11, suggesting a partitioning between Rab11-Evi5 and Rab11 effector complexes. Indeed, ablation of Evi5 by RNA interference causes a mislocalization of FIP3 at the abscission site during cytokinesis. These data demonstrate that Evi5 is a Rab11 binding protein and that Evi5 may cooperate with Rab11 to coordinate vesicular trafficking, cytokinesis, and cell cycle control independent of GTPase-activating protein function.
authors.affiliation: Genentech, Inc., 1 DNA Way, South San Francisco, CA 94080, USA.
authors.forename: Christopher J
authors.initials: CJ
authors.lastname: Westlake
authors.forename: Jagath R
authors.initials: JR
authors.lastname: Junutula
authors.forename: Glenn C
authors.initials: GC
authors.lastname: Simon
authors.forename: Manohar
authors.initials: M
authors.lastname: Pilli
authors.forename: Rytis
authors.initials: R
authors.lastname: Prekeris
authors.forename: Richard H
authors.initials: RH
authors.lastname: Scheller
authors.forename: Peter K
authors.initials: PK
authors.lastname: Jackson
authors.forename: Adam G
authors.initials: AG
authors.lastname: Eldridge
mesh_terms: D002453:Cell Cycle; D018797:Cell Cycle Proteins; D002460:Cell Line; D020558:GTP Phosphohydrolases / Q000378:metabolism*; D020690:GTPase-Activating Proteins; D006801:Humans; D051550:I-kappa B Kinase / Q000378:metabolism; D009687:Nuclear Proteins / Q000235:genetics / Q000378:metabolism*; D009857:Oncogenes; D011485:Protein Binding; D020349:Surface Plasmon Resonance; D020691:rab GTP-Binding Proteins / Q000378:metabolism*; D000099193:rab11 GTP-Binding Proteins
chemical_list: D018797:Cell Cycle Proteins; D020558:GTP Phosphohydrolases; D020690:GTPase-Activating Proteins; D051550:I-kappa B Kinase; D009687:Nuclear Proteins; D020691:rab GTP-Binding Proteins; D000099193:rab11 GTP-Binding Proteins; C506399:EVI5 protein, human; C491731:IKBKG protein, human