Agonist-induced PIP(2) hydrolysis inhibits cortical actin dynamics: regulation at a global but not at a micrometer scale

Mol Biol Cell. 2002 Sep;13(9):3257-67. doi: 10.1091/mbc.e02-04-0231.


Phosphatidylinositol 4, 5-bisphosphate (PIP(2)) at the inner leaflet of the plasma membrane has been proposed to locally regulate the actin cytoskeleton. Indeed, recent studies that use GFP-tagged pleckstrin homology domains (GFP-PH) as fluorescent PIP(2) sensors suggest that this lipid is enriched in membrane microdomains. Here we report that this concept needs revision. Using three distinct fluorescent GFP-tagged pleckstrin homology domains, we show that highly mobile GFP-PH patches colocalize perfectly with various lipophilic membrane dyes and, hence, represent increased lipid content rather than PIP(2)-enriched microdomains. We show that bright patches are caused by submicroscopical folds and ruffles in the membrane that can be directly visualized at approximately 15 nm axial resolution with a novel numerically enhanced imaging method. F-actin motility is inhibited significantly by agonist-induced PIP(2) breakdown, and it resumes as soon as PIP(2) levels are back to normal. Thus, our data support a role for PIP(2) in the regulation of cortical actin, but they challenge a model in which spatial differences in PIP(2) regulation of the cytoskeleton exist at a micrometer scale.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • 3T3 Cells
  • Actins / metabolism*
  • Animals
  • Cell Membrane / metabolism
  • Cell Movement
  • Cytoskeleton / metabolism
  • Green Fluorescent Proteins
  • Humans
  • Hydrolysis
  • Luminescent Proteins / metabolism
  • Mice
  • Microscopy, Confocal
  • Microscopy, Fluorescence
  • Models, Statistical
  • Phosphatidylinositol 4,5-Diphosphate / agonists
  • Phosphatidylinositol 4,5-Diphosphate / metabolism*
  • Photobleaching
  • Time Factors
  • Transfection
  • Tumor Cells, Cultured


  • Actins
  • Luminescent Proteins
  • Phosphatidylinositol 4,5-Diphosphate
  • Green Fluorescent Proteins