Identification of protein-tyrosine phosphatase 1B as the major tyrosine phosphatase activity capable of dephosphorylating and activating c-Src in several human breast cancer cell lines

c-Src tyrosine kinase activity is elevated in several types of human cancer , and this has been attributed to elevated c-Src expression levels, increas ed c-Src specific activity, and activating mutations in c-Src, We have foun d a number of human breast cancer cell lines with elevated c-Src specific a ctivity that also possess elevated phosphatase activity directed against th e carboxyl-terminal negative regulatory domain of Src family kinases, To id entify this phosphatase, cell extracts from MDA-MB-435S cells were chromato graphed and the fractions were assayed for phosphatase activity. Four peaks of phosphatase activity directed against the nonspecific substrate poly(Gl u/Tyr) were detected. One peak also dephosphorylated a peptide modeled agai nst the c-Src carboxyl-terminal negative regulatory domain and intact human c-Src. Immunoblotting and immunodepletion experiments identified the phosp hatase as protein-tyrosine phosphatase 1B (PTP1B), Examination of several h uman breast cancer cell lines with increased c-Src activity showed elevated levels of PTP1B protein relative to normal control breast cells. In vitro c-Src reactivation experiments confirmed the ability of PTP1B to dephosphor ylate and activate c-Src, In vivo overexpression of PTP1B in 293 cells caus ed a 2-fold increase of endogenous c-Src kinase activity. Our findings indi cate that PTP1B is the primary protein-tyrosine phosphatase capable of deph osphorylating c-Src in several human breast cancer cell lines and suggests a regulatory role for PTP1B in the control of c-Src kinase activity.