Immunostaining evidence for PI(4,5)P2 localization at the leading edge of chemoattractant-stimulated HL-60 cells

Ved P. Sharma, Vera DesMarais, Colin Sumners, Gerry Shaw, Atul Narang

Research output: Contribution to journalArticle

18 Scopus citations

Abstract

It is well known that in fMLP-stimulated neutrophils, phosphatidyl inositol 3,4,5-trisphosphate [PI(3,4,5)P3] localizes at the leading edge of the cells. However, no effort has been made to study the PI 4,5-bisphosphate [PI(4,5)P2] distribution in these cells. In fact, it has been suggested that PI(4,5)P2 is unlikely to localize, as its basal level is orders of magnitude higher than that of PI(3,4,5)P3. We developed an optimized immunostaining protocol for studying the endogenous distribution of PI(4,5)P2 in neutrophil-like HL-60 cells. We show that PI(4,5)P 2 localizes sharply at the leading edge with an intensity gradient similar to that for PI(3,4,5)P3. The enzymes for the production of PI(4,5)P2, namely, PI5KIα and PI5KIγ, were also found to localize at the leading edge, further supporting our finding that PI(4,5)P 2 localizes at the leading edge. These results imply that complementary regulation of PI3K and phosphate and tensin homolog (PTEN) is not the sole or dominant mechanism of PI(3,4,5)P3 polarization in HL-60 cells.

Original languageEnglish (US)
Pages (from-to)440-447
Number of pages8
JournalJournal of Leukocyte Biology
Volume84
Issue number2
DOIs
StatePublished - Aug 1 2008

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Keywords

  • Chemotaxis
  • Gradient sensing
  • Phosphoinositides

ASJC Scopus subject areas

  • Immunology and Allergy
  • Immunology
  • Cell Biology

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