Src homology region 2 (SH2) domain-containing phosphatase-1 dephosphorylates B cell linker protein/SH2 domain leukocyte protein of 65 kDa and selectively regulates c-Jun NH2-terminal kinase activation in B cells

Src homology region 2 (SH2) domain-containing phosphatase-1 (SXIP-1) is a c ytosolic protein tyrosine phosphatase containing two SH2 domains in its NH2 terminus. That immunological abnormalities of the motheaten and viable mot heaten mice are caused by mutations in the gene encoding SHP-1 indicates th at SHP-1 plays important roles in lymphocyte differentiation, proliferation , and activation. To elucidate molecular mechanisms by which SHP-1 regulate s BCR-mediated signal transduction, we determined SHP-1 substrates in B cel ls using the substrate-trapping approach. When the phosphatase activity-def icient form of SHP-1, in which the catalytic center cysteine (C453) was rep laced with serine (SHP-1-C/S), was introduced in WEHI-231 cells, tyrosine p hosphorylation of a protein of about 70 kDa was strongly enhanced. Immunopr ecipitation and Western blot analyses revealed that this protein is the B c ell linker protein (BLNK), also named SH2 domain leukocyte protein of 65 kD a, and that upon tyrosine phosphorylation BLNK binds to SHP-1-C/S in vitro. In vitro kinase assays demonstrated that hyperphosphorylation of BLNK in S HP-1-C/S-expressing cells was not due to enhanced activity of Lyn or Syk, F urthermore, BCR-induced activation of c-Jun NH2-terminal kinase was shown t o be significantly enhanced in SHP-1-C/S transfectants. Taken collectively, our results suggest that BLNK is a physiological substrate of SHP-1 in B c ells and that SHP-1 selectively regulates c-Jun NH2-terminal kinase activat ion.