Iron-regulatory protein 2 (IRP2), a posttranscriptional regulator of
iron metabolism, undergoes proteasomal degradation in
iron-replete cells, while it is stabilized in
iron deficiency or
hypoxia. IRP2 also responds to
nitric oxide (NO), as shown in various cell types exposed to pharmacological NO donors and in
gamma interferon/
lipopolysaccharide-stimulated macrophages. However, the diverse experimental systems have yielded conflicting results on whether NO activates or inhibits IRP2. We show here that a treatment of mouse B6 fibroblasts or human H1299
lung cancer cells with the NO-releasing
drug S-nitroso-N-acetyl-
penicillamine (SNAP) activates IRP2 expression. Moreover, the exposure of H1299 cells to SNAP leads to stabilization of
hemagglutinin (HA)-tagged IRP2, with kinetics analogous to those elicited by the
iron chelator desferrioxamine. Similar results were obtained with IRP2(Delta)(73), a mutant lacking a conserved, IRP2-specific
proline- and
cysteine-rich domain. Importantly, SNAP fails to stabilize HA-tagged p53, suggesting that under the above experimental conditions, NO does not impair the capacity of the
proteasome for protein degradation. Finally, by employing a coculture system of B6 and H1299 cells expressing
NO synthase II or IRP2-HA cDNAs, respectively, we demonstrate that NO generated in B6 cells stabilizes IRP2-HA in target H1299 cells by passive diffusion. Thus, biologically synthesized NO promotes IRP2 stabilization without compromising the overall proteasomal activity. These results are consistent with the idea that NO may negatively affect the labile
iron pool and thereby trigger responses to
iron deficiency.