title: Rab14 specifies the apical membrane through Arf6-mediated regulation of lipid domains and Cdc42.
abstract: The generation of cell polarity is essential for the development of multi-cellular organisms as well as for the function of epithelial organs in the mature animal. Small GTPases regulate the establishment and maintenance of polarity through effects on cytoskeleton, membrane trafficking, and signaling. Using short-term 3-dimensional culture of MDCK cells, we find that the small GTPase Rab14 is required for apical membrane specification. Rab14 knockdown results in disruption of polarized lipid domains and failure of the Par/aPKC/Cdc42 polarity complex to localize to the apical membrane. These effects are mediated through tight control of lipid localization, as overexpression of the phosphatidylinositol 4-phosphate 5-kinase α [PtdIns(4)P5K] activator Arf6 or PtdIns(4)P5K alone, or treatment with the phosphatidylinositol 3-kinase (PtdInsI3K) inhibitor wortmannin, rescued the multiple-apical domain phenotype observed after Rab14 knockdown. Rab14 also co-immunoprecipitates and colocalizes with the small GTPase Cdc42, and Rab14 knockdown results in increased Cdc42 activity. Furthermore, Rab14 regulates trafficking of vesicles to the apical domain, mitotic spindle orientation, and midbody position, consistent with Rab14's reported localization to the midbody as well as its effects upon Cdc42. These results position Rab14 at the top of a molecular cascade that regulates the establishment of cell polarity.
authors.affiliation: Department of Cellular and Molecular Medicine, University of Arizona, Tucson, AZ 85724, USA.
authors.forename: Ruifeng
authors.initials: R
authors.lastname: Lu
authors.affiliation: Department of Cellular and Molecular Medicine, University of Arizona, Tucson, AZ 85724, USA.
authors.forename: Jean M
authors.initials: JM
authors.lastname: Wilson
mesh_terms: D000089982:ADP-Ribosylation Factor 6; D020727:ADP-Ribosylation Factors / Q000235:genetics / Q000378:metabolism*; D000818:Animals; D001692:Biological Transport / Q000235:genetics; D002462:Cell Membrane / Q000378:metabolism*; D016764:Cell Polarity / Q000235:genetics / Q000502:physiology*; D004285:Dogs; D015151:Immunoblotting; D061985:Madin Darby Canine Kidney Cells; D021962:Membrane Microdomains / Q000378:metabolism; D008856:Microscopy, Fluorescence; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000378:metabolism; D034622:RNA Interference; D020764:cdc42 GTP-Binding Protein / Q000235:genetics / Q000378:metabolism*; D020691:rab GTP-Binding Proteins / Q000235:genetics / Q000378:metabolism*
chemical_list: D000089982:ADP-Ribosylation Factor 6; D017853:Phosphotransferases (Alcohol Group Acceptor); C020077:1-phosphatidylinositol-4-phosphate 5-kinase; D020727:ADP-Ribosylation Factors; D020764:cdc42 GTP-Binding Protein; D020691:rab GTP-Binding Proteins