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Fluorescence-Based Measurements of Phosphatidylserine/Phosphatidylinositol 4-Phosphate Exchange Between Membranes
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Distinct mechanisms of phospholipase D activation and attenuation utilized by different mitogens in NIH-3T3

P Ben-Av1, Y Eli, U S Schmidt

  • 1Department of Hormone Research, Weizmann Institute of Science, Rehovot, Israel.

European Journal of Biochemistry
|July 15, 1993
PubMed

Insights

Platelet-derived growth factor (PDGF), prostaglandin F2 alpha, and TPA rapidly activate phospholipase D (PLD) in fibroblasts. Protein kinase C mediates PDGF and TPA effects, but not prostaglandin F2 alpha signaling.

Area of Science:

  • Cellular signaling pathways
  • Enzymology
  • Signal transduction

Background:

  • Phospholipase D (PLD) is a key enzyme in cellular signaling.
  • Platelet-derived growth factor (PDGF), prostaglandin F2 alpha, and 12-O-tetradecanoylphorbol 13-acetate (TPA) are known mitogens that can activate cellular responses.

Purpose of the Study:

  • To investigate the activation mechanisms of phospholipase D (PLD) by different mitogens in NIH-3T3 fibroblasts.
  • To elucidate the role of protein kinase C (PKC) in PLD activation by PDGF, prostaglandin F2 alpha, and TPA.

Main Methods:

  • Metabolic labeling of NIH-3T3 fibroblasts with [3H]oleic acid.
  • Quantification of [3H]phosphatidic acid and [3H]phosphatidylpropanol to measure PLD activity.
  • Treatment with mitogens (PDGF, prostaglandin F2 alpha, TPA) and protein kinase inhibitors (staurosporine, GF109203X).
  • PKC depletion experiments.

Main Results:

  • All tested mitogens rapidly activated PLD.
  • PLD activation kinetics varied: transient for prostaglandin F2 alpha and PDGF, sustained for TPA.
  • Protein kinase C inhibition differentially affected PLD activation by the mitogens.
  • PKC depletion abolished PLD activation by all mitogens, indicating its crucial role.

Conclusions:

  • Agonist-specific mechanisms regulate PLD activation and inactivation.
  • Protein kinase C is involved in PDGF and TPA-induced PLD activation.
  • Prostaglandin F2 alpha activates PLD via a pathway independent of protein kinase C.

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