PROTEIN-TYROSINE-PHOSPHATASE 1B IS A NEGATIVE REGULATOR OF INSULIN-GROWTH-FACTOR-I-STIMULATED AND INSULIN-LIKE GROWTH-FACTOR-I-STIMULATED SIGNALING

To understand the physiological role of protein-tyrosine phosphatase 1 B (PTPase 1B) in insulin and insulinlike growth factor-I (IGF-I) signa ling we established clonal cell lines overexpressing wild type or inac tive mutant (C215S) PTPase 1B in cells overexpressing insulin (Hirc) o r IGF-I (CIGFR) receptors. PTPase 1B overexpression in transfected cel ls was verified by immunoblot analysis with a molloclonal PTPase 1B an tibody. Subfractionation of parental cells demonstrated that greater t han 90% of PTPase activity was localized in the Triton X-100-soluble p articulate (P1) cell fraction. PTPase activity in the P1 fraction of c ells overexpressing wild type PTPase 1B was 3-6-fold higher than paren tal cells but was unaltered in all fractions from C215S PTPase 1B-cont aining cells. The overexpression of wild type and C215S PTPase 1B had no effects on intrinsic receptor kinase activity growth rate, or gener al. cell morphology. The effects of PTPase IB overexpression on cellul ar protein tyrosine phosphorylation were examined by anti-phosphotyros ine immunoblot analysis. No differences were apparent under basal cond itions, but hormone-stimulated receptor autophosphorylation and/or ins ulin receptor substrate tyrosine phosphorylation were inhibited in cel ls overexpressing wild type PTPase 1B and increased in cells expressin g mutant PTPase 1B, in comparison with parental cells. Metalbolic sign aling, assessed by ligand-stimulated [C-14]glucose incorporation into glycogen, was also inhibited in cells overexpressing active PTPase 1B and enhanced in cells containing C215S PTPase 1B. These data strongly suggest that PTPase 1B acts as a negative regulator of insulin and IGF -I signaling.