Biosynthesis of nitric oxide activates iron regulatory factor in macrophages

Biosynthesis of nitric oxide (NO) from L‐arginine modulates activity of iron‐dependent enzymes, including mitochondrial acontiase, an [Fe‐S] protein. We examined the effect of NO on the activity of iron regulatory factor (IRF), a cytoplasmic protein which modulates both ferritin mRNA translation and transferrin receptor mRNA stability by binding to specific mRNA sequences called iron responsive elements (IREs). Murine macrophages were activated with interferon‐gamma and lipopolysaccharide to induce NO synthase activity and cultured in the presence or absence of NG‐substituted analogues of L‐arginine which served as selective inhibitors of NO synthesis. Measurement of the nitrite concentration in the culture medium was taken as an index of NO production. Mitochondria‐free cytosols were then prepared and aconitase activity as well as IRE binding activity and induction of IRE binding activity were correlated and depended on NO synthesis after IFN‐gamma and/or LPS stimulation. Authentic NO gas as well as the NO‐generating compound 3‐morpholinosydnonimine (SIN‐1) also conversely modulated aconitase and IRE binding activities of purified recombinant IRF. These results provide evidence that endogenously produced NO may modulate the post‐transcriptional regulation of genes involved in iron homeostasis and support the hypothesis that the [Fe‐S] cluster of IRF mediates iron‐dependent regulation.