DESCRIPTION (from Applicant's Abstract): This proposal focuses on an investigation of the effect of nitric oxide on protein tyrosine phosphorylation that is associated with signal transduction. The studies originate from our observation that nitric oxide can stimulate protein tyrosine phosphorylation of protein bands 126, 56 and 43 kD as found by SDS PAGE in HER 14 cells(a fibroblast NIH3T3 cell line transfected with epidermal growth actor receptor). In our first specific aim, we will continue our investigation on the effect of the exogenous generation of nitric oxide on protein tyrosine phosphorylation while also investigating the endogenous production of nitric oxide in HER 14 cells, with the intent of determining whether or not endogenously generated nitric oxide can also modulate protein tyrosine phosphorylation by epidermal growth factor receptor, or by integrin, a membrane associated adhesion molecule that also stimulates protein tyrosine phosphorylation. Growth and survival studies will be used to assess physiologic outcome. In our second specific aim, we will measure the activities of selected proteins in the growth factor signal transduction pathway (ex. ras) with the intent of observing whether or not these activities are activated by nitric oxide. The experiments will also indicate if the signalling pathway must be fully intact for observing the effect of nitric oxide on protein tyrosine phosphorylation. To further substantiate this idea, in our third specific aim, HER 14 cells will be infected with inactive dominant negative as. These transfected cells will be assessed for changes in protein tyrosine phosphorylation and selected protein activities involved in signalling associated with protein tyrosine phosphorylation. In our fourth specific aim, we will determine the identity of the protein bands that undergo protein tyrosine phosphorylation on exposure of HER 14 cells to nitric oxide. Specific antibodies will be used to identify these proteins using immuno-blotting and immunoprecipitation techniques. Our approach could define the possible relationship between exogenously or endogenously generated nitric oxide and signalling pathways associated with protein tyrosine phosphorylation . This may have implications for cellular proliferation and cellular toxicity, such as occurs in the proliferation of vascular cells, where nitric oxide produced by endothelial cells interacts with circulating growth factors to modulate cell proliferation in the arterial wall, in macrophage induced cytotoxicity associated with host defense mechanisms and in environmental exposure (pulmonary) to nitric oxide that is found in cigarette smoke and as a product of combustion found in smog.