Inducible nitric-oxide-synthase mRNA is transiently expressed and destroyed by a cycloheximide-sensitive process

Abstract

Nitric oxide is a mediator of a wide range of physiological processes. It is produced by an enzyme family, the nitric-oxide synthases, one form of which is induced in many cells following stimulation with cytokines and lipopolysaccharide. The aim of the experiments reported in this study was to investigate the regulation of mRNA expression for this inducible nitric-oxide synthase in smooth muscle cells and macrophages. Stimulation of these cells with cytokines and lipopolysaccharide results in a marked elevation of nitric-oxide-synthase mRNA levels, which however do not remain elevated, but reach a maximum at 3-6 h after stimulation before returning to baseline levels over the next 20 h. Enzyme activity, however, remained virtually constant for 48 h following stimulation. Inspection of the 3' untranslated segment of both murine and human inducible nitric-oxide-synthase mRNAs showed the presence of a conserved AU-rich octanucleotide sequence, previously identified in cytokine and oncogene mRNAs and shown to mediate mRNA instability. A particular feature of the breakdown of mRNAs bearing this sequence is that degradation is prevented by protein-synthesis inhibition. We show in this study that the half-life of inducible nitric-oxide-synthase mRNA is 6 h and that in the presence of an inhibitor of protein synthesis this breakdown is prevented. Thus, the mRNA for inducible nitric-oxide synthase shares some features in common with cytokines such as the transient expression and decay of its mRNA which can be prevented by protein-synthesis inhibition.