BLNK is associated with the CD72/SHP-1/Grb2 complex in the WEHI231 cell line after membrane IgM cross-linking

Tyrosine phosphorylation of CD72 strongly correlates with B cell antigen receptor signals leading to apoptosis. We have previously shown that CD72 carrying two immunoreceptor tyrosine-based inhibition motifs (ITIM) is an in vivo substrate of SHP-1. CD72 forms a complex with SHP-1 and Grb2 via its tyrosine-phosphorylated ITIM when the WEHI231 cell line, which is representative of immature B cells, undergoes apoptosis. The CD72 complex formation was also demonstrated in normal primary B cells, suggesting that the complex formation in apoptotic B cells is a universal mechanism. In this study, we further investigated the molecular components of the CD72 complex in WEHI231 cells in order to understand the molecular mechanism involved in the signaling pathway mediated through the complex. Our experiments demonstrate that BLNK, a recently identified adaptor molecule predominantly expressed in B cells, is associated with the CD72 complex via the Src homology 3 domain(s) of Grb2 in the cell line after membrane IgM (mIgM) engagement. The results suggest that the mIgM-mediated signal strongly correlates with the formation of the CD72 / SHP-1 / Grb2 / BLNK complex.