S-nitrosylation is a ubiquitous redox-related modification of cysteine thiols by nitric oxide (NO), which mediates many aspects of NO action. Nitrosylation has thus been shown to modulate the activity of a wide variety of proteins, thereby influencing many aspects of cell physiology, including endothelial cells. In particular, recent work has raised the idea that protein nitrosylation is a reversible and highly ordered process, both spatially and temporally, features that enable it to regulate cell signaling. However, the in vivo mechanisms that control nitrosylation/denitrosylation of proteins are poorly understood. Here, we explore cellular mechanisms that control the nitrosylation/denitrosylation cycle of caspase-3, a key regulator of apoptosis. We employ novel biochemical and genetic screens, specifically designed to identify NO dependent protein-protein interactions, to isolate enzymes that regulate nitrosylation of caspases. We extend the findings of these studies to endothelial cell dysfunction and vascular disease. Specifically, we will address the possible role of protein nitrosylation in these pathologic conditions using in-vitro systems of endothelial cell apoptosis and in animal models atherosclerosis.
