Stimulation of the B cell Ag receptor (BCR), a multimeric complex containing heterodimers of Ig-alpha and Ig-beta, initiates a cascade of
tyrosine phosphorylation that results in cellular activation. One of the earliest substrates phosphorylated is Ig-alphabeta, and it appears that
kinase activation emanates from this structure with the most proximal
kinases themselves, and some of their immediate substrates, associating with the heterodimer. To identify other molecules that may be involved in proximal BCR signaling, we examined the substrates that were
tyrosine phosphorylated following stimulation with either
anti-IgG Abs or
pervanadate in the murine
B cell lymphoma A20 IIA1.6 and in resting splenic B cells. Immunoblotting with anti-
phosphotyrosine Abs revealed that a doublet of 40 and 42 kDa was phosphorylated within 1 min of stimulation with either agonist. The phosphorylation of p40/42 in A20 cells induced by
anti-IgG was rapid and transient, peaking at 2 min after stimulation and becoming almost undetectable after 10 min. Furthermore, at least 25% of phosphorylated p40/42 co-immunoprecipitated with Ig-alphabeta, but none precipitated with MHC II, CD40,
Fc(gamma)RII, Fyn, HS-1, or Syk, suggesting that this
protein complex specifically associates with the Ig-alphabeta heterodimer. p40/42 did not react with Abs to Ig-alpha, Ig-beta,
mitogen-activated protein kinase, or Lnk. Furthermore, and in contrast to Ig-alphabeta, p40/42 was highly acidic and not part of a
disulfide-linked complex. Finally, p40/42 was demonstrated to be a glycosylated
surface protein that was constitutively associated with Ig-alphabeta. These results suggest that p40/42 is a novel constituent of the resting B cell Ag receptor complex.