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Phospholipase d promotes lipid microdomain-associated signaling events in mast cells

Felipe A Lisboa1, Ze Peng, Christian A Combs

  • 1Laboratory of Molecular Immunology, National Heart, Lung, and Blood Institute, National Institutes of Health, Bethesda, MD 20892-1760, USA.

Journal of Immunology (Baltimore, Md. : 1950)
|October 2, 2009
PubMed

Insights

Phospholipase D (PLD) activity is crucial for IgE-dependent mast cell signaling. PLD promotes the accumulation of key signaling proteins within lipid microdomains, impacting allergic responses.

Area of Science:

  • Immunology
  • Cell Biology
  • Biochemistry

Background:

  • Immunoglobulin E (IgE)-dependent signaling initiates mast cell activation.
  • These initial signaling events occur within detergent-resistant membrane microdomains.
  • Fc(epsilon)RI receptor aggregation within these domains is critical for downstream signaling.

Purpose of the Study:

  • To investigate the role of phospholipase D (PLD) in IgE-dependent signaling within mast cell lipid microdomains.
  • To determine if PLD activity influences the localization of Fc(epsilon)RI and associated signaling molecules.
  • To assess the impact of PLD on mast cell degranulation.

Main Methods:

  • Utilized RBL-2H3 mast cells stimulated with antigen (Ag).
  • Employed 1-butanol to inhibit phosphatidic acid production and assess its effect on Fc(epsilon)RI localization.
  • Used small inhibitory RNAs (siRNAs) to knockdown PLD1 and PLD2 expression.
  • Monitored Fc(epsilon)RI and Lyn kinase accumulation in detergent-resistant membrane fractions.
  • Assessed phosphorylation of signaling proteins and mast cell degranulation.
  • Visualized lipid microdomain changes using fluorescent-tagged cholera toxin B subunit and confocal microscopy.

Main Results:

  • 1-butanol treatment inhibited Fc(epsilon)RI association with detergent-resistant membranes and reduced phosphatidic acid production.
  • Knockdown of PLD2, and to a lesser extent PLD1, suppressed Fc(epsilon)RI and Lyn accumulation in lipid microdomains.
  • PLD inhibition/knockdown reduced phosphorylation of Fc(epsilon)RI, Lyn, and linker for activation of T cells, and inhibited degranulation.
  • Changes in the distribution of the lipid microdomain component ganglioside GM1 were observed.

Conclusions:

  • PLD activity is essential for the proper localization of Fc(epsilon)RI and associated signaling molecules within lipid microdomains during IgE-dependent signaling.
  • PLD plays a significant role in regulating mast cell activation and degranulation.
  • Targeting PLD may offer a therapeutic strategy for allergic inflammatory diseases.

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