Membrane ruffling requires coordination between type Ialpha phosphatidylinositol phosphate kinase and Rac signaling

J Biol Chem. 2003 Jun 20;278(25):23036-45. doi: 10.1074/jbc.M211397200. Epub 2003 Apr 6.

Abstract

Membrane ruffle formation requires remodeling of cortical actin filaments, a process dependent upon the small G-protein Rac. Growth factors stimulate actin remodeling and membrane ruffling by integration of signaling pathways that regulate actin-binding proteins. Phosphatidylinositol 4,5-bisphosphate (PIP2) regulates the activity of many actin-binding proteins and is produced by the type I phosphatidylinositol phosphate kinases (PIPKIs). Here we show in MG-63 cells that only the PIPKIalpha isoform is localized to platelet-derived growth factor (PDGF)-induced membrane ruffles. Further, expression of kinase dead PIPKIalpha, which acts as a dominant negative mutant, blocked membrane ruffling, suggesting that PIPKIalpha and PIP2 participate in ruffling. To explore this, PIPKIalpha was overexpressed in serum-starved cells and stimulated with PDGF. In serum-starved cells, PIPKIalpha expression did not stimulate actin remodeling, but when these cells were stimulated with PDGF, actin rapidly reorganized into foci but not membrane ruffles. PIPKIalpha-mediated formation of actin foci was independent of both Rac1 and phosphatidylinositol 3-kinase activities. Significantly, coexpression of dominant active Rac1 with PIPKIalpha in PDGF-stimulated cells resulted in membrane ruffling. The PDGF- and Rac1-stimulated ruffling was inhibited by expression of kinase-dead PIPKIalpha. Combined, these data support a model where the localized production of PIP2 by PIPKIalpha is necessary for actin remodeling, whereas formation of membrane ruffles required Rac signaling.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Amino Acid Substitution
  • Base Sequence
  • Cell Line
  • Cell Membrane / drug effects
  • Cell Membrane / ultrastructure*
  • Chromones / pharmacology
  • Cloning, Molecular
  • DNA Primers
  • Enzyme Inhibitors / pharmacology
  • Fibroblasts / cytology
  • Fibroblasts / enzymology
  • Humans
  • Microscopy, Confocal
  • Minor Histocompatibility Antigens
  • Morpholines / pharmacology
  • Mutagenesis, Site-Directed
  • Phosphotransferases (Alcohol Group Acceptor) / metabolism*
  • Platelet-Derived Growth Factor / pharmacology
  • Recombinant Proteins / chemistry
  • Recombinant Proteins / metabolism
  • Signal Transduction
  • rac GTP-Binding Proteins / chemistry
  • rac GTP-Binding Proteins / metabolism*

Substances

  • Chromones
  • DNA Primers
  • Enzyme Inhibitors
  • Minor Histocompatibility Antigens
  • Morpholines
  • Platelet-Derived Growth Factor
  • Recombinant Proteins
  • 2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one
  • Phosphotransferases (Alcohol Group Acceptor)
  • phosphatidylinositol phosphate 4-kinase
  • rac GTP-Binding Proteins