Activated Cdc42/Rac reconstitutes Fc epsilon RI-mediated Ca2+ mobilizationand degranulation in mutant RBL mast cells

Antigen stimulation of mast cells via Fc epsilon RI, the high-affinity rece ptor for IgE, triggers a signaling cascade that requires Ca2+ mobilization for exocytosis of secretory granules during an allergic response. This stud y investigates critical signaling components by using mutant RBL mast cells that are defective in antigen-stimulated phospholipase C gamma (PLC gamma) activation, as well as other signaling activities downstream of stimulated tyrosine phosphorylation. We show that the expression of activated version s of the Cdc42 or Rac1 GTPase restores antigen-stimulated Ca2+ mobilization necessary for degranulation in these mutant cells. Wild-type Cdc42 and Rac 1, as well as activated Cdc42 containing effector domain mutations, all fai l to restore antigen-stimulated signaling leading to exocytosis, Expression of oncogenic Dbl, a guanine nucleotide exchange factor for Cdc42 and Rac1, partially restores sustained Ca2+ mobilization and degranulation, suggesti ng that activation of endogenous Cdc42 and/or Rac1 is impaired in the mutan t cells. Overexpression of PLC gamma1 with either activated Cdc42 or Rad sy nergistically stimulates degranulation, consistent with a critical defect i n PLC gamma activation in these cells. Thus, our results point to activatio n of Cdc42 and/or Rad playing an essential role in antigen stimulation of e arly events that culminate in mast cell degranulation.