Migration and retraction of endothelial and epithelial cells require PHI-1, a specific protein-phosphatase-1 inhibitor protein

J Cell Sci. 2004 Nov 15;117(Pt 24):5905-12. doi: 10.1242/jcs.01506. Epub 2004 Nov 2.

Abstract

Cell migration and retraction are interrelated activities that are crucial for a range of physiological processes such as wound healing and vascular permeability. Immunostaining of brain sections for the specific inhibitor of type-1 protein Ser/Thr phosphatase called PHI-1 showed high expression levels in smooth muscle and especially in vascular endothelial cells. During migration of cultured human lung microvascular endothelial cells, endogenous PHI-1 was concentrated to the trailing edge of the cells. Knockdown of PHI-1 using small interfering RNAs reduced by 45% the rate of HeLa cell migration in a wound-healing assay. These cells exhibited an extremely elongated phenotype relative to controls and time-lapse movies revealed a defect in retraction of the trailing edge. Both HeLa and human vascular endothelial cells depleted of PHI-1 showed increased surface areas relative to controls during cell spreading in a replating assay. Analysis of sequential microscopic images demonstrated this was due to a significant decrease in the number of retraction events, whereas protrusive action was unaffected. The Ser/Thr phosphorylation of several signaling, cytoskeletal and focal-adhesion proteins was unchanged in PHI-1-depleted cells, so the target of PHI-1 inhibited protein-phosphatase 1 remains unidentified. Nonetheless, the results show that PHI-1 participates in regulatory events at the trailing edge of migrating cells and modulates retraction of endothelial and epithelial cells.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Animals
  • Brain / metabolism
  • Cell Movement
  • Cells, Cultured
  • Cytoskeleton / metabolism
  • Endothelium, Vascular / cytology*
  • Epithelial Cells / cytology*
  • Gene Silencing
  • HeLa Cells
  • Humans
  • Immunohistochemistry
  • Lung / blood supply
  • Lymphocytes / cytology
  • Microcirculation
  • Microscopy, Fluorescence
  • Phosphoprotein Phosphatases / metabolism
  • Phosphorylation
  • Proteins / metabolism*
  • RNA Interference
  • RNA, Small Interfering / metabolism
  • Rats
  • Serine / chemistry
  • Threonine / chemistry
  • Time Factors
  • Transfection
  • Wound Healing

Substances

  • Proteins
  • RNA, Small Interfering
  • phosphoprotein phosphatase inhibitor 1
  • Threonine
  • Serine
  • Phosphoprotein Phosphatases