Nitric oxide inhibits the degradation of IRP2.
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.
Wang
Jian
J
Lady Davis Institute for Medical Research, Sir Mortimer B. Davis Jewish General Hospital, 3755 Cote-Ste-Catherine Rd., Montreal, Quebec H3T 1E2, Canada.
Chen
Guohua
G
Pantopoulos
Kostas
K
D000818:Animals; D003676:Deferoxamine / Q000494:pharmacology; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects / Q000502:physiology*; D006801:Humans; D007501:Iron / Q000378:metabolism; D035942:Iron Regulatory Protein 2 / Q000378:metabolism*; D051379:Mice; D009569:Nitric Oxide / Q000378:metabolism*; D020030:Nitric Oxide Donors / Q000494:pharmacology; D019001:Nitric Oxide Synthase / Q000378:metabolism*; D052247:Nitric Oxide Synthase Type II; D010396:Penicillamine / Q000031:analogs & derivatives* / Q000494:pharmacology; D046988:Proteasome Endopeptidase Complex / Q000378:metabolism*; D011489:Protein Denaturation / Q000187:drug effects / Q000502:physiology; D014407:Tumor Cells, Cultured; D016159:Tumor Suppressor Protein p53 / Q000378:metabolism
D020030:Nitric Oxide Donors; C110881:S-nitro-N-acetylpenicillamine; D016159:Tumor Suppressor Protein p53; D009569:Nitric Oxide; D007501:Iron; D019001:Nitric Oxide Synthase; D052247:Nitric Oxide Synthase Type II; D046988:Proteasome Endopeptidase Complex; D035942:Iron Regulatory Protein 2; D010396:Penicillamine; D003676:Deferoxamine