Nitric Oxide Inhibition of ERK1/2 Activity in Cells Expressing Neuronal Nitric-oxide Synthase.

Neuronal nitric-oxide synthase (nNOS) is a constitutively expressed enzyme responsible for the production of nitric oxide (NO•) from L-arginine and O2. Nitric oxide is an intra- and intercellular messenger that mediates a diversity of signaling pathways in target cells. In the absence of L-arginine, nNOS has been shown to generate superoxide (O2...). Superoxide, either directly or through its self-dismutation to H2O2, is likewise believed to be a cell-signaling agent. Because nNOS can generate NO• and O2..., we examined the activation of cellular signal transduction pathways in nNOS-transfected cells grown in the presence or absence of L-arginine. Spin trapping/ EPR spectroscopy confirmed that stimulated nNOS-transfected cells grown in an L-arginine environment secreted NO• into the surrounding milieu. Production of NO• blocked Ca2+ ionophore-induced activation of the ERK½ through a mechanism involving inhibition of the Ras G-protein and Raf-1 kinase. In contrast, ERK actiration was largely unaffected in nNOS-transfected cells grown in L-arginine-free media. Inhibition of nNOS-generated NO• with the competitive NOS inhibitor, NG-nitro-L-arginine methyl ester, in cells grown in L-arginine restored ERK½ activation to levels similar to that found when nNOS was activated in L-arginine-free media. These findings indicate that nNOS can differentially regulate the ERK signal transduction pathway in a manner dependent on the presence of L-arginine and the production of NO•. [ABSTRACT FROM AUTHOR]