Modulation of the basal activity of N-acetylglucosaminyltransferase V by phosphatidylinositol-3-kinase/protein kinase B signaling pathway in human hepatocarcinoma cells

The modulation of GnT-V activity by signaling molecules in PI-3-K/PKB pathw ay in human hepatocarcinoma cell line 7721 was studied. GnT-V activity was determined after the transfection of sense or antisense cDNA of PKB into th e cells, as well as the addition of activators, specific inhibitors, and th e antibodies to the enzyme assay system or culture medium. It was found tha t the basal activity of GnT-V was up regulated by the sense and down regula ted by the antisense cDNA of PKB transfected into 7721 cells. GnT-V was act ivated by PIP2, PIP3 or GTP gamma [S] added to the assay system, and the ac tivation of PIP2 or GTP gamma [S] was abolished by LY2940002, a specific in hibitor of PI-3-K, but the activation of PIP3 was not attenuated by LY29400 02. In addition, GnT-V activity in cultured parental or H-ras transfected c ells was inhibited by the antibody against PKB or PI-3-K. These findings de monstrated the involvement of PI-3-K/PKB signaling pathway in the regulatio n of GnT-V. Moreover, ET18-OCH3, an inhibitor of Raf translocation and PI-P LC enzyme, which produces the activator of PKC, as well as the antibodies a gainst Raf-1 or MEK also inhibited GnT-V activity in the parental and H-ras transfected cells. The inhibitory rates, however, were less in the transfe cted cells than those in the parental cells. These results reveal that in p arental and H-ras transfected 7721 cells, the basal activity of GnT-V is al so regulated by the Ras/Raf-1/MEK/MAPK cascade in addition to PI-3-K/PKB si gnaling pathway. The significance of these two pathways in the regulation o f GnT-V and their relations to the activation of PKC previously reported by our laboratory (Ju TZ et al., 1995 Glyconjugate J 12, 767-772) was discuss ed.