INHIBITION OF LYN FUNCTION IN MAST-CELL ACTIVATION BY SH3 DOMAIN BINDING PEPTIDES

While Lyn tyrosine kinase has been shown to be necessary for IgE-recep tor (Fc epsilon RI)-mediated mast cell activation, the mechanism of Ly n activation is not yet understood, Using a micro-electroporation tech nique to quantitatively introduce peptides into the cytosol of tumor m ast cells, we show that proline-rich peptides that preferentially bind Src family SH3 domains block receptor-induced repetitive calcium spik es in a concentration dependent manner. The Src family member Lyn was the Likely target, since a series of phage displaying derived peptides with increased Lyn SH3 domain binding specificity inhibited Fc epsilo n RI-mediated calcium signaling at concentrations consistent with bind ing to Lyn rather than other Src-type kinases. Furthermore, SH3 bindin g peptides prevented the plasma membrane translocation of a fluorescen tly labeled Syk tandem SH2 domain, which binds to phosphorylated Fc ep silon RI, suggesting that the peptides specifically block the Lyn-medi ated step by which Fc epsilon RI cross-linking leads to receptor phosp horylation. Our study suggests that the binding of proline-rich peptid es, or corresponding cellular interaction partners, to Lyn SH3 domain suppresses the Lyn-mediated phosphorylatation of Fc epsilon RI and cal cium signaling.