NITRIC-OXIDE ACTION ON GROWTH FACTOR-ELICITED SIGNALS - PHOSPHOINOSITIDE HYDROLYSIS AND [CA2+](I) RESPONSES ARE NEGATIVELY MODULATED VIA A CGMP-DEPENDENT PROTEIN-KINASE-I PATHWAY

The role of nitric oxide (NO) in the phosphatidylinositol 4,5-bisphosphate (PIP2) hydrolysis and intracellular Ca2+ release responses induced by epidermal, platelet derived, and fibroblast growth factors was investigated in three cell Lines, a clone of NIH-3T3 fibroblasts overexpressing epidermal growth factor receptors and the tumoral epithelial cells A431 and KB. In all three cell types, pretreatment with NO donors decreased growth factor-induced PIP2 and Ca2+ responses, whereas pretreatment with NO synthase inhibitors increased them. The Ca2+-dependent PIP, hydroysis induced by micromolar concentrations-of the Ca2+ ionophore, ionomycin, was also modulated negatively and positively by NO donors and synthase inhibitors, respectively. In contrast, the Ca2+ content of the intracellular stores was unaffected by the various pretreatments employed. NO donors and synthase inhibitors induced an increase and decrease, respectively, of the intracellular cGMP formation in all three cell lines investigated, All of the effects of the NO donors were mimicked by 8-bromo-cGMP administration and abolished by pretreatment with the specific blocker of the cGMP-dependent protein kinase I, KT5823, which by itself mimicked the effects of the synthase inhibitors. Together with previous observations on G protein-coupled receptors, the present results demonstrate that PIP, hydrolysis and Ca2+ release occur under the feedback control of NO, independently of the phospholipase C (beta, gamma, or delta type) involved and of the mechanism of activation. Such a control, which appears to be effected by the cGMP-dependent protein kinase I acting at the level of the phospholipases C themselves, might ultimately contribute to the inhibitory role of NO on growth previously observed with various cell types.