Molecular targets of CD45 in B cell antigen receptor signal transduction

Expression of the phosphotyrosine phosphatase CD45 is essential for B cell Ag receptor (BCR)-mediated p21(ras) activation and calcium mobilization. To examine the molecular basis of this requirement, we analyzed signaling events following BCR ligation in CD45-deficient (CD45(-)) and CD45-reconstituted (CD45(+)) variants of J558L mu m3 cells. Ag stimulation resulted in tyrosine phosphorylation of cellular proteins in both cells. However, the spectrum of proteins phosphorylated in the CD45(+) cells was qualitatively and/or quantitatively distinct from that in the CD45(-) cells. Among the protein tyrosine kinases examined, the Src family kinases Fyn and Blk were inducibly tyrosine phosphorylated and activated by receptor ligation only in CD45(+) cells. While Ag-induced Btk tyrosine phosphorylation occurred in both cells, its activation was greatly diminished in the CD45(-) cells, Analysis of specific effector molecules revealed that tyrosine phosphorylation of Shc, but not rasGAP or Vav, correlated with the unique ability of BCR ligation to trigger p21(ras) activation in CD45(+) cells. BCR-mediated She phosphorylation and recruitment of Grb2 depended on CD45 expression. Thus, Shc tyrosine phosphorylation may be the primary CD45-dependent mechanism by which Ag receptors are coupled to the p21(ras) pathway in J558L mu m3. In addition, phospholipase C gamma 1 (PLC gamma 1) and PLC gamma 2 were tyrosine phosphorylated upon Ag stimulation in CD45(-) cells, despite much reduced inositol trisphosphate production and lack of calcium mobilization. These findings suggest that CD45 may modulate events other than PLC gamma phosphorylation, which regulate phosphoinositide hydrolysis and the calcium mobilization response following BCR ligation.