Mutant RBL mast cells defective in Fc epsilon RI signaling and lipid raft biosynthesis are reconstituted by activated Rho-family GTPases

Characterization of defects in a variant subline of RBL mast cells has reve aled a biochemical event proximal to IgE receptor (Fc epsilon RI)-stimulate d tyrosine phosphorylation that is required for multiple functional respons es. This cell line, designated B6A4C1, is deficient in both Fc epsilon RI-m ediated degranulation and biosynthesis of several lipid raft components. Ag ents that by-pass receptor-mediated Ca2+ influx stimulate strong degranulat ion responses in these variant cells. Cross linking of IgE-Fc epsilon RI on these cells stimulates robust tyrosine phosphorylation but fails to mobili ze a sustained Ca2+ response. Fc epsilon RI-mediated inositol phosphate pro duction is not detectable in these cells, and failure of adenosine receptor s to mobilize Ca2+ suggests a general deficiency in stimulated phospholipas e C activity. Antigen stimulation of phospholipases A(2) and D is also defe ctive. Infection of B6A4C1 cells with vaccinia virus constructs expressing constitutively active Rho family members Cdc42 and Rac restores antigen-sti mulated degranulation, and active Cdc42 (but not active Rac) restores gangl ioside and GPI expression, The results support the hypothesis that activati on of Cdc42 and/or Rac is critical for Fc epsilon RI-mediated signaling tha t leads to Ca2+ mobilization and degranulation. Furthermore, they suggest t hat Cdc42 plays an important role in the biosynthesis and expression of cer tain components of lipid rafts.