Iron-dependent oxidation, ubiquitination, and degradation of iron regulatory protein 2: Implications for degradation of oxidized proteins

Iron-dependent oxidation, ubiquitination, and degradation of iron regulatory protein 2: Implications for degradation of oxidized proteins

^*Cell Biology and Metabolism Branch, National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, MD 20892-5430; ^‡Laboratory of Biochemistry, National Heart, Lung and Blood Institute, National Institutes of Health, Bethesda, MD 20892-0320; ^§Laboratory of Immune Cell Biology, Division of Basic Sciences, National Cancer Institute, Bethesda, MD 20892; and ^†Department of Immunology and Cell Biology, Graduate School of Medicine, Kyoto University, Sakyo-ku, Kyoto 606, Japan 606-8501

Contributed by Richard D. Klausner

Abstract

The ability of iron to catalyze formation of reactive oxygen species significantly contributes to its toxicity in cells and animals. Iron uptake and distribution is regulated tightly in mammalian cells, in part by iron regulatory protein 2 (IRP2), a protein that is degraded efficiently by the proteasome in iron-replete cells. Here, we demonstrate that IRP2 is oxidized and ubiquitinated in cells before degradation. Moreover, iron-dependent oxidation converts IRP2 into a substrate for ubiquitination in vitro. A regulatory pathway is described in which excess iron is sensed by its ability to catalyze site-specific oxidations in IRP2, oxidized IRP2 is ubiquitinated, and ubiquitinated IRP2 subsequently is degraded by the proteasome. Selective targeting and removal of oxidatively modified proteins may contribute to the turnover of many proteins that are degraded by the proteasome.

Footnotes

↵ ¶ To whom reprint requests should be addressed at: Cell Biology and Metabolism Branch, National Institute of Child Health and Human Development, Building 18T, Room 101, National Institutes of Health, Bethesda, MD 20892-5430. e-mail: trou{at}helix.nih.gov.
ABBREVIATIONS:

IRP,

iron regulatory protein;

Df,

desferrioxamine;

FAC,

ferric amminium citrate;

DNP,

2,4-dinitrophenylhydrazine