Cytoglobin regulates cell respiration and nitrosative stress through NO dioxygenation and co‐localizes with inducible nitric oxide synthase during vascular injury.

Abstract

Disposition of the second messenger nitric oxide (NO) in mammalian tissues occurs through multiple pathways including reaction with erythrocyte hemoglobin, red muscle myoglobin, and metabolism by NO dioxygenase activities in non‐striated tissues. Although the newly discovered globin cytoglobin binds molecular oxygen and dioxygenates NO in vitro, the lack of an associated reductase activity has raised doubts about the ability of cytoglobin to dioxygenate NO in vivo. We hypothesize that non‐striated cells can dioxygenate NO and that cytoglobin contributes to this activity. For this purpose, we stably expressed short hairpin RNA targeting cytoglobin in mouse fibroblasts. This resulted in an 80 percent reduction in cytoglobin protein expression. Wild type cells demonstrated intracellular nitrate generation upon addition of exogenous NO that was oxygen‐dependent and cyanide‐inhibitable. The sustained dioxygenation of NO observed over a 30 min exposure to NO was diminished in cells with low cytoglobin expression. Protein nitrosation and heme‐nitrosylation were increased in the absence of cytoglobin. These cells were also more sensitive to NO‐induced inhibition of cell respiration. Normal response to NO could be re‐established through expression of human cytoglobin in the knock down cells. In summary, our study reveals a pivotal role for cytoglobin in cell‐mediated NO dioxygenation to regulate nitrosative stress and cell respiration during conditions of chronic NO exposure. This may impact regulation of NO bioavailability in the vasculature.