title: Rab11-FIP2 interaction with MYO5B regulates movement of Rab11a-containing recycling vesicles.
abstract: A tripartite association of Rab11a with both Rab11-FIP2 and MYO5B regulates recycling endosome trafficking. We sought to define the intermolecular interactions required between Rab11-FIP2 and MYO5B. Using a random mutagenesis strategy, we identified point mutations at S229P or G233E in Rab11-FIP2 that caused loss of interaction with MYO5B in yeast two-hybrid assays as well as loss of interaction of Rab11-FIP2(129-356) with MYO5B tail when expressed in HeLa cells. Single mutations or the double S229P/G233E mutation failed to alter the association of full-length Rab11-FIP2 with MYO5B tail in HeLa cells. While EGFP-Rab11-FIP2 wild type colocalized with endogenous MYO5B staining in MDCK cells, EGFP-Rab11-FIP2(S229P/G233E) showed a significant decrease in localization with endogenous MYO5B. Analysis of Rab11a-containing vesicle movement in live HeLa cells demonstrated that when the MYO5B/Rab11-FIP2 association is perturbed by mutation or by Rab11-FIP2 knockdown, vesicle movement is increased in both speed and track length, consistent with an impairment of MYO5B tethering at the cytoskeleton. These results support a critical role for the interaction of MYO5B with Rab11-FIP2 in stabilizing the functional complex with Rab11a, which regulates dynamic movements of membrane recycling vesicles.
authors.affiliation: Section of Surgical Sciences and the Epithelial Biology Center, Vanderbilt University School of Medicine, Nashville, TN, USA.
authors.forename: Jenny C
authors.initials: JC
authors.lastname: Schafer
authors.forename: Nicholas W
authors.initials: NW
authors.lastname: Baetz
authors.forename: Lynne A
authors.initials: LA
authors.lastname: Lapierre
authors.forename: Rebecca E
authors.initials: RE
authors.lastname: McRae
authors.forename: Joseph T
authors.initials: JT
authors.lastname: Roland
authors.forename: James R
authors.initials: JR
authors.lastname: Goldenring
mesh_terms: D000818:Animals; D001665:Binding Sites; D002352:Carrier Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D004285:Dogs; D011992:Endosomes / Q000378:metabolism*; D006367:HeLa Cells; D006801:Humans; D061985:Madin Darby Canine Kidney Cells; D008565:Membrane Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D018995:Myosin Heavy Chains / Q000235:genetics / Q000378:metabolism*; D024701:Myosin Type V / Q000235:genetics / Q000378:metabolism*; D017354:Point Mutation; D011485:Protein Binding; D021381:Protein Transport; D020691:rab GTP-Binding Proteins / Q000378:metabolism*; D000099193:rab11 GTP-Binding Proteins
chemical_list: D002352:Carrier Proteins; D008565:Membrane Proteins; D018995:Myosin Heavy Chains; D024701:Myosin Type V; D020691:rab GTP-Binding Proteins; D000099193:rab11 GTP-Binding Proteins; C505832:MYO5B protein, human; C463963:RAB11FIP2 protein, human
keywords: MYO5B; Rab11; Rab11-FIP2; membrane recycling