Expression of the phosphotyrosine phosphatase CD45 is essential for B
cell Ag receptor (BCR)-mediated p21(ras) activation and calcium mobili
zation. To examine the molecular basis of this requirement, we analyze
d signaling events following BCR ligation in CD45-deficient (CD45(-))
and CD45-reconstituted (CD45(+)) variants of J558L mu m3 cells. Ag sti
mulation resulted in tyrosine phosphorylation of cellular proteins in
both cells. However, the spectrum of proteins phosphorylated in the CD
45(+) cells was qualitatively and/or quantitatively distinct from that
in the CD45(-) cells. Among the protein tyrosine kinases examined, th
e Src family kinases Fyn and Blk were inducibly tyrosine phosphorylate
d and activated by receptor ligation only in CD45(+) cells. While Ag-i
nduced Btk tyrosine phosphorylation occurred in both cells, its activa
tion 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 pho
sphorylation and recruitment of Grb2 depended on CD45 expression. Thus
, Shc tyrosine phosphorylation may be the primary CD45-dependent mecha
nism by which Ag receptors are coupled to the p21(ras) pathway in J558
L mu m3. In addition, phospholipase C gamma 1 (PLC gamma 1) and PLC ga
mma 2 were tyrosine phosphorylated upon Ag stimulation in CD45(-) cell
s, despite much reduced inositol trisphosphate production and lack of
calcium mobilization. These findings suggest that CD45 may modulate ev
ents other than PLC gamma phosphorylation, which regulate phosphoinosi
tide hydrolysis and the calcium mobilization response following BCR li
gation.