Phosphoinositide 3-Kinase Facilitates Antigen-stimulated Ca²⁺ Influx in RBL-2H3 Mast Cells via a Phosphatidylinositol 3,4,5-Trisphosphate-sensitive Ca²⁺ Entry Mechanism

This study presents evidence that phosphoinositide 3-kinase (PI3K) plays a concerted role with phospholipase Cγ in initiating antigen-mediated Ca²⁺ signaling in mast cells via a phosphatidylinositol 3,4,5-trisphosphate (PI(3,4,5)P₃)-sensitive Ca²⁺ entry pathway. Exogenous PI(3,4,5)P₃ at concentrations close to its physiological level induces instantaneous Ca²⁺ influx into RBL-2H3 cells. This PI(3,4,5)P₃-induced intracellular Ca²⁺ increase is independent of phospholipase C activity or the depletion of internal stores. Moreover, inhibition of PI3K by LY294002 or by overexpression of the dominant negative inhibitor Δp85 suppresses the Ca²⁺ response to the cross-linking of the high affinity receptor for IgE (FcεRI). Concomitant treatment of RBL-2H3 cells with LY294002 or Δp85 and 2-aminoethyl diphenylborate, a cell-permeant antagonist of D-myo-inositol 1,4,5-trisphosphate receptors, abrogates antigen-induced Ca ²⁺ signals, whereas either treatment alone gives rise to partial inhibition. Conceivably, PI(3,4,5)P₃-sensitive Ca²⁺ entry and capacitative Ca²⁺ entry represent major Ca²⁺ influx pathways that sustain elevated [Ca²⁺]_i to achieve optimal physiological responses. This study also refutes the second messenger role of D-myo-inositol 1,3,4,5-tetrakisphosphate in regulating FcεRI-mediated Ca²⁺ response. Considering the underlying mechanism, our data suggest that PI(3,4,5)P₃ directly stimulates a Ca²⁺ transport system in plasma membranes. Together, these data provide a molecular basis to account for the role of PI3K in the regulation of FcεRI-mediated degranulation in mast cells.