title: Phosphorylation of Rab11-FIP2 regulates polarity in MDCK cells.
abstract: The Rab11 effector Rab11-family interacting protein 2 (Rab11-FIP2) regulates transcytosis through its interactions with Rab11a and myosin Vb. Previous studies implicated Rab11-FIP2 in the establishment of polarity in Madin-Darby canine kidney (MDCK) cells through phosphorylation of Ser-227 by MARK2. Here we examine the dynamic role of Rab11-FIP2 phosphorylation on MDCK cell polarity. Endogenous Rab11-FIP2 phosphorylated on Ser-227 coalesces on vesicular plaques during the reestablishment of polarity after either monolayer wounding or calcium switch. Whereas expression of the nonphosphorylatable Rab11-FIP2(S227A) elicits a loss in lumen formation in MDCK cell cysts grown in Matrigel, the putative pseudophosphorylated Rab11-FIP2(S227E) mutant induces the formation of cysts with multiple lumens. On permeable filters, Rab11-FIP2(S227E)-expressing cells exhibit alterations in the composition of both the adherens and tight junctions. At the adherens junction, p120 catenin and K-cadherin are retained, whereas the majority of the E-cadherin is lost. Although ZO-1 is retained at the tight junction, occludin is lost and the claudin composition is altered. Of interest, the effects of Rab11-FIP2 on cellular polarity did not involve myosin Vb or Rab11a. These results indicate that Ser-227 phosphorylation of Rab11-FIP2 regulates the composition of both adherens and tight junctions and is intimately involved in the regulation of polarity in epithelial cells.
authors.affiliation: Section of Surgical Sciences and Epithelial Biology Center, Vanderbilt University Medical Center, Nashville, TN 37232, USA.
authors.forename: Lynne A
authors.initials: LA
authors.lastname: Lapierre
authors.forename: Kenya M
authors.initials: KM
authors.lastname: Avant
authors.forename: Cathy M
authors.initials: CM
authors.lastname: Caldwell
authors.forename: Asli
authors.initials: A
authors.lastname: Oztan
authors.forename: Gerard
authors.initials: G
authors.lastname: Apodaca
authors.forename: Byron C
authors.initials: BC
authors.lastname: Knowles
authors.forename: Joseph T
authors.initials: JT
authors.lastname: Roland
authors.forename: Nicole A
authors.initials: NA
authors.lastname: Ducharme
authors.forename: James R
authors.initials: JR
authors.lastname: Goldenring
mesh_terms: D022005:Adherens Junctions / Q000378:metabolism; D000818:Animals; D015153:Blotting, Western; D015820:Cadherins / Q000235:genetics / Q000378:metabolism; D051177:Catenins / Q000235:genetics / Q000378:metabolism; D002460:Cell Line; D016764:Cell Polarity; D057167:Claudins / Q000235:genetics / Q000378:metabolism; D004285:Dogs; D004847:Epithelial Cells / Q000378:metabolism*; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D057809:HEK293 Cells; D006801:Humans; D007668:Kidney / Q000166:cytology / Q000378:metabolism; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism; D018613:Microscopy, Confocal; D009154:Mutation; D062793:Occludin; D010766:Phosphorylation; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012694:Serine / Q000235:genetics / Q000378:metabolism; D019108:Tight Junctions / Q000378:metabolism; D033921:Vesicular Transport Proteins / Q000235:genetics / Q000378:metabolism*; D000095002:Delta Catenin
chemical_list: D015820:Cadherins; D051177:Catenins; D057167:Claudins; D008565:Membrane Proteins; C568985:OCLN protein, human; D062793:Occludin; D033921:Vesicular Transport Proteins; D049452:Green Fluorescent Proteins; D012694:Serine; C087310:K cadherin; D000095002:Delta Catenin