Nitric oxide (NO) is a potent cell signaling and effector molecule that participates in numerous physiological and pathophysiological events in a variety of cell types and tissues. NO derived from all major isoforms of NO synthase can S-nitrosylate cysteine thiols in target proteins, potentially altering their functional activities in a redox-dependent, cGMP-independent manner. Formation of protein S-nitrosocysteine adducts appears to occur through multiple pathways. Emerging evidence suggests that S-nitrosylation is a specific, reversible and regulated covalent post-translational modifi-cation that modulates diverse biological and physiological functions. In addition to altering protein activity, localization and stability, S-nitrosylation participates in the control of cellular metabolism, apoptosis, protein – protein interactions, transcription factor function, ion channel activity and cellular redox balance. Increasingly sophisticated proteomic ap-proaches used in various cell types and tissues have identified S-nitrosylation of proteins of virtually all major classes, in-cluding cytoskeletal proteins, chaperones, proteins of the translational and transcriptional machinery, vesicular transport and signaling. S-nitrosylation has also been shown to regulate the NO synthase isoforms themselves, reversibly inhibiting endothelial NO synthase activity and feedback inhibiting PARP-1, a transactivator of inducible NO synthase. Imbalances in NO metabolism and dysregulated S-nitrosylation have been implicated in a growing list of human diseases, such as neurodegenerative disorders, endotoxemic shock and insulin resistance. Here we review the key discoveries and directions in this field, including the role of S-nitrosylation as a potential therapeutic target in specific human diseases.