RESUMEN
Nitric oxide (â¢NO) is a biologically important short-lived free radical signaling molecule. Both the enzymatic synthesis and the predominant forms of cellular metabolism of â¢NO are oxygen-dependent. For these reasons, changes in local oxygen concentrations can have a profound influence on steady-state â¢NO concentrations. Many proteins are regulated by â¢NO in a concentration-dependent manner, but their responses are elicited at different thresholds. Using soluble guanylyl cyclase (sGC) and p53 as model â¢NO-sensitive proteins, we demonstrate that their concentration-dependent responses to â¢NO are a function of the O2 concentration. p53 requires relatively high steady-state â¢NO concentrations (>600 nM) to induce its phosphorylation (P-ser-15), whereas sGC responds to low â¢NO concentrations (<100 nM). At a constant rate of â¢NO production (liberation from â¢NO-donors), decreasing the O2 concentration (1%) lowers the rate of â¢NO metabolism. This raises steady-state â¢NO concentrations and allows p53 activation at lower doses of the â¢NO donor. Enzymatic â¢NO production, however, requires O2 as a substrate such that decreasing the O2 concentration below the K m for O2 for nitric oxide synthase (NOS) will decrease the production of â¢NO. We demonstrate that the amount of â¢NO produced by RAW 264.7 macrophages is a function of the O2 concentration. Differences in rates of â¢NO production and â¢NO metabolism result in differential sGC activation that is not linear with respect to O2. There is an optimal O2 concentration (≈5-8%) where a balance between the synthesis and metabolism of â¢NO is established such that both the â¢NO concentration and sGC activation are maximal.