Distinct phosphoinositide 3-kinases mediate mast cell degranulation in response to G-protein-coupled versus FcεRI receptors

David A. Windmiller, Jonathan M. Backer

Research output: Contribution to journalArticle

40 Citations (Scopus)

Abstract

Phosphoinositide (PI) 3-kinases are critical regulators of mast cell degranulation. The Class IA PI 3-kinases p85/p110β and p85/p110δ but not p85/p110α are required for antigen-mediated calcium flux in RBL-2H3 cells (Smith, A. J., Surviladze, Z., Gaudet, E. A., Backer, J. M., Mitchell, C. A., and Wilson, B. S. et al., (2001) J. Biol. Chem. 276, 17213-17220). We now examine the role of Class IA PI 3-kinases isoforms in degranulation itself, using a single-cell degranulation assay that measures the binding of fluorescently tagged annexin V to phosphatidylserine in the outer leaflet of the plasma membrane of degranulated mast cells. Consistent with previous data, antibodies against p110δ and p110β blocked FcεR1-mediated degranulation in response to FcεRI ligation. However, antigen-stimulated degranulation was also inhibited by antibodies against p110α, despite the fact that these antibodies have no effect on antigen-induced calcium flux. These data suggest that p110α mediates a calcium-independent signal during degranulation. In contrast, only p110β was required for enhancement of antigen-stimulated degranulation by adenosine, which augments mast cell-mediated airway inflammation in asthma. Finally, we examined carbachol-stimulated degranulation in RBL2H3 cells stably expressing the M1 muscarinic receptor (RBL-2H3-M1 cells). Surprisingly, carbachol-stimulated degranulation was blocked by antibody-mediated inhibition of the Class III PI 3-kinase hVPS34 or by titration of its product with FYVE domains. Antibodies against Class IA PI 3-kinases had no effect. These data demonstrate: (a) a calcium-independent role for p110α in antigen-stimulated degranulation; (b) a requirement for p110β in adenosine receptor signaling; and (c) a requirement for hVPS34 during M1 muscarinic receptor signaling. Elucidation of the intersections between these distinct pathways will lead to new insights into mast cell degranulation.

Original languageEnglish (US)
Pages (from-to)11874-11878
Number of pages5
JournalJournal of Biological Chemistry
Volume278
Issue number14
DOIs
StatePublished - Apr 4 2003

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Cell Degranulation
1-Phosphatidylinositol 4-Kinase
Phosphatidylinositols
GTP-Binding Proteins
Mast Cells
Phosphotransferases
Antigens
Antibodies
Muscarinic M1 Receptors
Calcium
Carbachol
Fluxes
Purinergic P1 Receptors
Immunoglobulin Isotypes
Annexin A5
Phosphatidylserines
Cell membranes
Titration
Adenosine
Ligation

ASJC Scopus subject areas

  • Biochemistry

Cite this

Distinct phosphoinositide 3-kinases mediate mast cell degranulation in response to G-protein-coupled versus FcεRI receptors. / Windmiller, David A.; Backer, Jonathan M.

In: Journal of Biological Chemistry, Vol. 278, No. 14, 04.04.2003, p. 11874-11878.

Research output: Contribution to journalArticle

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