Atypical protein kinase C (iota) activates ezrin in the apical domain of intestinal epithelial cells

Flavia A. Wald, Andrea S. Oriolo, Anastasia Mashukova, Nevis L. Fregien, Amber H. Langshaw, Pedro J.I. Salas

Research output: Contribution to journalArticle

52 Scopus citations

Abstract

Atypical protein kinase iota (PKCι) is a key organizer of the apical domain in epithelial cells. Ezrin is a cytosolic protein that, upon activation by phosphorylation of T567, is localized under the apical membrane where it connects actin filaments to membrane proteins and recruits protein kinase A (PKA). To identify the kinase that phosphorylates ezrin T567 in simple epithelia, we analyzed the expression of active PKC and the appearance of T567-P during enterocyte differentiation in vivo. PKCι phosphorylated ezrin on T567 in vitro, and in Sf9 cells that do not activate human ezrin. In CACO-2 human intestinal cells in culture, PKCι co-immunoprecipitated with ezrin and was knocked down by ARNA expression. The resulting phenotype showed a modest decrease in total ezrin, but a steep decrease in T567 phosphorylation. The PKCι-depleted cells showed fewer and shorter microvilli and redistribution of the PKA regulatory subunit. Expression of a dominant-negative form of PKCι also decreased T567-P signal, and expression of a constitutively active PKCι mutant showed depolarized distribution of T567-P. We conclude that, although other molecular mechanisms contribute to ezrin activation, apically localized phosphorylation by PKCι is essential for the activation and normal distribution of ezrin at the early stages of intestinal epithelial cell differentiation.

Original languageEnglish (US)
Pages (from-to)644-654
Number of pages11
JournalJournal of Cell Science
Volume121
Issue number5
DOIs
StatePublished - Mar 1 2008

Keywords

  • Apical domain
  • Brush border
  • Epithelial polarity
  • ERM proteins

ASJC Scopus subject areas

  • Cell Biology

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