Involvement of SH2-SH2-SH3 domain of phospholipase C gamma 1 in NF-kappa Bsignaling

To directly define the role of phospholipase C gamma 1 (PLC gamma 1) in NF- kappa B activation, NF-kappa B promoted luciferase reporter gene plasmid (p NF-kappa B-Luc) was transfected into rat-3Y1 fibroblasts that overexpress w hole PLC gamma 1 (PLC gamma 1-3Y1), src homology domains SH2-SH2-SH3 of PLC gamma 1 (SH223-3Y1) and v-src (Src-3Y1), Transient transfection with pNF-k appa B-Luc remarkably increased the luciferase activity in all three transf ormants compared with normal rat-3Y1 cells. Pretreatment with inhibitors of protein tyrosine kinase reduced this increase in luciferase activity, but U73122 (a PLC inhibitor) did not. While PD98059, an inhibitor of mitogen ac tivated protein kinase (MAPK), significantly reduced the luciferase activit y, there was no effect by wortmannin and Ro-31-8220, inhibitors of phosphat idylinositol 3-kinase (PI3K) and protein kinase C (PKC), respectively. This study shows a direct evidence that the SH2-SH2-SH3 region of PLC gamma 1 c ontributes to the NF-kappa B signaling and that MAPK, but not PI3K and PKC, is involved in SH2-SH2-SH3 mediated NF-kappa B activation in these cells. (C) 2000 Federation of European Biochemical Societies.