Regulation of B cell antigen receptor-mediated signaling by SHP-1

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By using mouse immature B cell line, WEHI-231, we have investigated how SHP-1 and SHP-1-associated molecules, cytoplasmic proteintyrosine phosphatase (PTP) containing two SH2 domains, regulate B cell antigen receptor (BCR)-mediated signaling event, especially MAP kinase (MAPK) pathways which play important roles in cell proliferation, differentiation and apoptotic cell death. The results obtained are follows ;(1) WEHI-231 cells were transfected with cDNA encoding wild-type SHP-1 (SHP-1-wt) or mutant SHP-1 (SHP-1-CS) which lacks PTP activity by substituting Cys to Ser in PTP domain. Then, the activity of each member of MAPK family was measured by Western blotting using antibodies against phosphorylated forms of MAPKs. ERK activity in SHP-1-wt or SHP-1-CS transfectant after the stimulation with anti-IgM Ab was almost identical to that in untransfected control cells, whereas anti-IgM-induced JNK and p38 activities in both transfectants were enhanced when compared to those in control cells … More . It should be noted that enhancement of JNK activity after anti-IgM stimulation was most prominent. In addition, SHP-1-wt transfectant showed enhanced anti-IgM-induced apoptotic cell death.(2) We have previously showen that SHP-1 is constitutively associated with SLP-76, leukocyte specific protein containing a SH2 domain at its C terminus. Next, WEHI-231 cells were transfected with cDNA encoding wild-type or mutant forms of SLP-76 and anti-IgM-induced MAPK activation was analyzed in these transfectants. There are little differences in the degree of activities of three MAPK members between untransfected and transfected cells. In contrast, anti-IgM-induced cell death was significantly suppressed in the cells transfected with mutant SLP-76 whose SH2 domain fails to bind phosphorylated Tyr residues.(3) When cells were transfected with SHP-1-CS-encoding cDNA, tyrosine phosphorylation of several proteins were enhanced after anti-IgM stimulation. We focused on one protein whose phosphorylation was most enhanced as a candidate for the SHP-1 substrate and characterized its nature. Less