The NOSs are a family of complex enzymes that catalyze the five-electron oxidation of L-arginine to form NO and L-citrulline. They are best characterized as cytochrome P-450-like hemeproteins that depend on molecular oxygen, NADPH, flavins, and tetrahydrobiopterin. The three human NOS isoforms identified to date, ecNOS, nNOS, and iNOS, are found on human chromosomes 7, 12, and 17, respectively. Regulation of NO synthesis and release occurs at the levels of enzyme activity and mRNA synthesis. The nNOS mRNA is structurally diverse as a consequence of alternative promoters and alternate splicing. The iNOS gene is predominantly regulated at the level of transcription by synergistic combinations of proinflammatory cytokines and bacterial wall products. Changes in mRNA levels of the ecNOS following endothelium activation are mediated by altered rates of transcription as well as by the intriguing process of changes in mRNA stability. Given the essential role of the NO pathway in a wide variety of physiological and pathophysiological process, it is possible that the three isoforms of NOS contribute to polygenic genetic diversity in neurological, immune, and cardiovascular biology. Further studies are needed to determine the mechanisms of gene regulation of NOS in health and disease.