Biosynthesis of nitric oxide (NO) from L-arginine modulates activity o
f iron-dependent enzymes, including mitochondrial aconitase, an [Fe-S]
protein. We examined the effect of NO on the activity of iron regulat
ory 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). Murin
e macrophages were activated with interferon-gamma and lipopolysacchar
ide to induce NO synthase activity and cultured in the presence or abs
ence of N(G)-substituted analogues Of L-arginine which served as selec
tive inhibitors of NO synthesis. Measurement of the nitrite concentrat
ion in the culture medium was taken as an index of NO production. Mito
chondria-free cytosols were then prepared and aconitase activity as we
ll as IRE binding activity assessed in parallel. Inhibition of enzymat
ic activity and induction of IRE binding activity were correlated and
depended on NO synthesis after IFN-gamma and/or LPS stimulation. Authe
ntic NO gas as well as the NO-generating compound 3-morpholinosydnonim
ine (SIN-1) also conversely modulated aconitase and IRE binding activi
ties of purified recombinant IRF. These results provide evidence that
endogenously produced NO may modulate the post-transcriptional regulat
ion of genes involved in iron homeostasis and support the hypothesis t
hat the [Fe-S] cluster of IRF mediates iron-dependent regulation.