Endogenous endothelial cell nitric-oxide synthase modulates apoptosis in cultured breast cancer cells and is transcriptionally regulated by p53

Nitric oxide can both stimulate and suppress apoptosis. By reverse transcri ptase-polymerase chain reaction and sequencing we show that human breast ca ncer (MCF-7) cells express endothelial cell nitric-oxide synthase (ecNOS), but not other nitric-oxide synthase isoforms. Inhibition of ecNOS activity in MCF-7 cells increased tumor cell apoptosis, and this effect was also see n following treatment with an NO scavenger. In addition, low concentrations of the NO donor sodium nitroprusside inhibited, whereas high concentration s stimulated MCF-7 cell apoptosis. The ecNOS promoter was found to contain a specific binding site for the apoptosis-regulating protein p53. In co-tra nsfection studies wild-type, but not mutant, p53 down-regulated transcripti on of an ecNOS promoter-luciferase reporter gene construct. In addition, NO donors up-regulated p53 protein levels in MCF-7 cells. These data point to a previously unrecognized p53-dependent regulation of ecNOS expression tha t may be important both for regulating apoptosis and for avoiding the gener ation of genotoxic quantities of NO.