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Nitric oxide and peroxynitrite activate the iron regulatory protein-1 of J774A.1 macrophages by direct disassembly of the Fe-S cluster of cytoplasmic aconitase.
Biochemistry. 2002 Jun 11; 41(23):7435-42.B

Abstract

Posttranscriptional regulation of iron homeostasis involves, among other factors, a reversible conversion of the Fe-S enzyme cytoplasmic aconitase to a mRNA-binding iron regulatory protein (IRP-1) that lacks an Fe-S cluster. Previous studies have shown that aconitase/IRP-1 may be a target of *NO or peroxynitrite (ONOO(-)), formed after reaction of *NO with superoxide anion (O(2)(*-)); however, the mechanisms and consequences of such interactions have remained uncertain. In this study, recombinant aconitase/IRP-1 was exposed to SIN-1, whose thermal decomposition releases *NO and O(2)(*-). Results showed that SIN-1 was able to induce concomitant inactivation of aconitase and activation of IRP-1, attributable to cluster disassembly induced by ONOO(-). SIN-1 was used also in lysates of J774A.1 mouse macrophages grown under control conditions, or subjected to iron loading or starvation by treatment with hemin or desferrioxamine, respectively. Three lines of evidence confirmed that ONOO(-) activated IRP-1 by removing iron from the Fe-S cluster of cytoplasmic aconitase. First, IRP-1 activation was accompanied by iron release and loss of aconitase activity. Second, aconitase activity was recovered by reassembling Fe-S clusters with cysteine and ferrous ammonium sulfate. Third, iron release and IRP-1 activation were observed in lysates from control or iron-loaded macrophages, containing increasing levels of Fe-S clusters, but not in lysates from iron-starved macrophages, in which aconitase had already undergone cluster disassembly and switched to IRP-1. *NO was less efficient than ONOO(-) in attacking the Fe-S cluster of cytoplasmic aconitase; in fact, SIN-1-dependent iron release and IRP-1 activation were diminished by superoxide dismutase, which scavenged O(2)(*-) before it reacted with *NO to form ONOO(-). Under comparable conditions, however, both *NO and ONOO(-) inactivated an IRP-2 unable to assemble an Fe-S cluster. These results indicate that *NO and ONOO(-) may activate IRP-1 by attacking the Fe-S cluster of cytoplasmic aconitase, while also inactivating the cluster-deficient IRP-2. Such divergent actions offer clues to explain links between iron homeostasis and reactive nitrogen species in macrophages involved in inflammation or other pathophysiologic conditions.

Authors+Show Affiliations

Institute of General Pathology, University of Milan, via Mangiagalli 31, 20133 Milan, Italy. gaetano.cairo@unimi.itNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info available

Pub Type(s)

Journal Article
Research Support, Non-U.S. Gov't

Language

eng

PubMed ID

12044177

Citation

Cairo, Gaetano, et al. "Nitric Oxide and Peroxynitrite Activate the Iron Regulatory Protein-1 of J774A.1 Macrophages By Direct Disassembly of the Fe-S Cluster of Cytoplasmic Aconitase." Biochemistry, vol. 41, no. 23, 2002, pp. 7435-42.
Cairo G, Ronchi R, Recalcati S, et al. Nitric oxide and peroxynitrite activate the iron regulatory protein-1 of J774A.1 macrophages by direct disassembly of the Fe-S cluster of cytoplasmic aconitase. Biochemistry. 2002;41(23):7435-42.
Cairo, G., Ronchi, R., Recalcati, S., Campanella, A., & Minotti, G. (2002). Nitric oxide and peroxynitrite activate the iron regulatory protein-1 of J774A.1 macrophages by direct disassembly of the Fe-S cluster of cytoplasmic aconitase. Biochemistry, 41(23), 7435-42.
Cairo G, et al. Nitric Oxide and Peroxynitrite Activate the Iron Regulatory Protein-1 of J774A.1 Macrophages By Direct Disassembly of the Fe-S Cluster of Cytoplasmic Aconitase. Biochemistry. 2002 Jun 11;41(23):7435-42. PubMed PMID: 12044177.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Nitric oxide and peroxynitrite activate the iron regulatory protein-1 of J774A.1 macrophages by direct disassembly of the Fe-S cluster of cytoplasmic aconitase. AU - Cairo,Gaetano, AU - Ronchi,Raffaella, AU - Recalcati,Stefania, AU - Campanella,Alessandro, AU - Minotti,Giorgio, PY - 2002/6/5/pubmed PY - 2002/7/20/medline PY - 2002/6/5/entrez SP - 7435 EP - 42 JF - Biochemistry JO - Biochemistry VL - 41 IS - 23 N2 - Posttranscriptional regulation of iron homeostasis involves, among other factors, a reversible conversion of the Fe-S enzyme cytoplasmic aconitase to a mRNA-binding iron regulatory protein (IRP-1) that lacks an Fe-S cluster. Previous studies have shown that aconitase/IRP-1 may be a target of *NO or peroxynitrite (ONOO(-)), formed after reaction of *NO with superoxide anion (O(2)(*-)); however, the mechanisms and consequences of such interactions have remained uncertain. In this study, recombinant aconitase/IRP-1 was exposed to SIN-1, whose thermal decomposition releases *NO and O(2)(*-). Results showed that SIN-1 was able to induce concomitant inactivation of aconitase and activation of IRP-1, attributable to cluster disassembly induced by ONOO(-). SIN-1 was used also in lysates of J774A.1 mouse macrophages grown under control conditions, or subjected to iron loading or starvation by treatment with hemin or desferrioxamine, respectively. Three lines of evidence confirmed that ONOO(-) activated IRP-1 by removing iron from the Fe-S cluster of cytoplasmic aconitase. First, IRP-1 activation was accompanied by iron release and loss of aconitase activity. Second, aconitase activity was recovered by reassembling Fe-S clusters with cysteine and ferrous ammonium sulfate. Third, iron release and IRP-1 activation were observed in lysates from control or iron-loaded macrophages, containing increasing levels of Fe-S clusters, but not in lysates from iron-starved macrophages, in which aconitase had already undergone cluster disassembly and switched to IRP-1. *NO was less efficient than ONOO(-) in attacking the Fe-S cluster of cytoplasmic aconitase; in fact, SIN-1-dependent iron release and IRP-1 activation were diminished by superoxide dismutase, which scavenged O(2)(*-) before it reacted with *NO to form ONOO(-). Under comparable conditions, however, both *NO and ONOO(-) inactivated an IRP-2 unable to assemble an Fe-S cluster. These results indicate that *NO and ONOO(-) may activate IRP-1 by attacking the Fe-S cluster of cytoplasmic aconitase, while also inactivating the cluster-deficient IRP-2. Such divergent actions offer clues to explain links between iron homeostasis and reactive nitrogen species in macrophages involved in inflammation or other pathophysiologic conditions. SN - 0006-2960 UR - https://www.unboundmedicine.com/medline/citation/12044177/Nitric_oxide_and_peroxynitrite_activate_the_iron_regulatory_protein_1_of_J774A_1_macrophages_by_direct_disassembly_of_the_Fe_S_cluster_of_cytoplasmic_aconitase_ L2 - https://doi.org/10.1021/bi025756k DB - PRIME DP - Unbound Medicine ER -