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  • The EMBO Journal (2001) 20, 2723 - 2741
  • doi:10.1093/emboj/20.11.2723

Ezrin is a downstream effector of trafficking PKC–integrin complexes involved in the control of cell motility

Tony Ng1,2,8, Maddy Parsons1,8, William E. Hughes3, James Monypenny4, Daniel Zicha4, Alexis Gautreau5, Monique Arpin5, Steve Gschmeissner6, Peter J. Verveer7, Philippe I.H. Bastiaens7 and Peter J. Parker3

  1. Richard Dimbleby Department of Cancer Research, St Thomas' Hospital, Lambeth Palace Road, London SE1 7EH, UK
  2. Cell Biophysics Laboratory, Imperial Cancer Research Fund, 44 Lincoln's Inn Fields, London WC2A 3PX, UK
  3. Protein Phosphorylation Laboratory, Imperial Cancer Research Fund, 44 Lincoln's Inn Fields, London WC2A 3PX, UK
  4. Light Microscopy Laboratory, Imperial Cancer Research Fund, 44 Lincoln's Inn Fields, London WC2A 3PX, UK
  5. Laboratoire de Morphogenese et Signalisation Cellulaires, UMR 144 CNRS/Institut Curie, 75248 Paris Cedex 05, France
  6. Electron Microscopy Unit, Imperial Cancer Research Fund, 44 Lincoln's Inn Fields, London WC2A 3PX, UK
  7. Cell Biology and Cell Biophysics Programme, EMBL, Meyerhofstrasse 1, D-69117 Heidelberg, Germany
  8. T.Ng and M.Parsons contributed equally to this work

Correspondence to:

Tony Ng, E-mail: T.Ng@icrf.icnet.uk

Received 30 November 2000; Accepted 5 April 2001; Revised 5 April 2001

Protein kinase C (PKC) alpha has been implicated in beta1 integrin-mediated cell migration. Stable expression of PKCalpha is shown here to enhance wound closure. This PKC-driven migratory response directly correlates with increased C-terminal threonine phosphorylation of ezrin/moesin/radixin (ERM) at the wound edge. Both the wound migratory response and ERM phosphorylation are dependent upon the catalytic function of PKC and are susceptible to inhibition by phosphatidylinositol 3-kinase blockade. Upon phorbol 12,13-dibutyrate stimulation, green fluorescent protein–PKCalpha and beta1 integrins co-sediment with ERM proteins in low-density sucrose gradient fractions that are enriched in transferrin receptors. Using fluorescence lifetime imaging microscopy, PKCalpha is shown to form a molecular complex with ezrin, and the PKC-co-precipitated endogenous ERM is hyperphosphorylated at the C-terminal threonine residue, i.e. activated. Electron microscopy showed an enrichment of both proteins in plasma membrane protrusions. Finally, overexpression of the C-terminal threonine phosphorylation site mutant of ezrin has a dominant inhibitory effect on PKCalpha-induced cell migration. We provide the first evidence that PKCalpha or a PKCalpha-associated serine/threonine kinase can phosphorylate the ERM C-terminal threonine residue within a kinase–ezrin molecular complex in vivo.

  • Keywords:

    • ERM,
    • FLIM,
    • migration,
    • PKC,
    • wound