Basal and stimulated protein S-nitrosylation in multiple cell types and tissues

There is substantial evidence that protein S-nitrosylation provides a significant route through which nitric oxide (NO)-derived bioactivity is conveyed. However, most examples of S-nitrosylation have been characterized on the basis of analysis in vitro, and relatively little progress has been made in assessing the participant forms of nitric-oxide synthase (NOS) or the dynamics of protein S-nitrosylation in situ. Here we utilize antibodies specific for the nitrosothiol (SNO) moiety to provide an immunohistochemical demonstration that protein S-nitrosylation is coupled to the activity of each of the major forms of NOS. In cultured endothelial cells, SNOprotein immunoreactivity increases in response to Ca2+- stimulated endothelial NOS (eNOS) activity, and in aortic rings, endothelium-derived and eNOS-mediated relaxation (EDRF) is coupled to increased protein S-nitrosylation in both endothelial and associated smooth muscle cells. In cultured macrophages, SNO-protein levels increase upon cytokine induction of induced NOS (iNOS), and in PC12 cells, increased protein S-nitrosylation is linked to nerve growth factor induction of neuronal NOS (nNOS). In addition, we describe developmental and pathophysiological. increases in SNOprotein immunoreactivity within human lung. These results, which demonstrate Ca2+, neurohumoral, growth factor, cytokine, and developmental regulation of protein S-nitrosylation that is coupled to NOS expression and activity, provide unique evidence for the proposition that this ubiquitous NO-derived post-translational protein modification serves as a major effector of NO-related bioactivity.