Cysteine oxidation regulates the RNA-binding activity of iron regulatory protein 2.
Iron regulatory protein 2 (IRP2) is an RNA-binding protein that regulates the posttranscriptional expression of proteins required for iron homeostasis such as ferritin and transferrin receptor 1. IRP2 RNA-binding activity is primarily regulated by iron-mediated proteasomal degradation, but studies have suggested that IRP2 RNA binding is also regulated by thiol oxidation. We generated a model of IRP2 bound to RNA and found that two cysteines (C512 and C516) are predicted to lie in the RNA-binding cleft. Site-directed mutagenesis and thiol modification show that, while IRP2 C512 and C516 do not directly interact with RNA, both cysteines are located within the RNA-binding cleft and must be unmodified/reduced for IRP2-RNA interactions. Oxidative stress induced by cellular glucose deprivation reduces the RNA-binding activity of IRP2 but not IRP2-C512S or IRP2-C516S, consistent with the formation of a disulfide bond between IRP2 C512 and C516 during oxidative stress. Decreased IRP2 RNA binding is correlated with reduced transferrin receptor 1 mRNA abundance. These studies provide insight into the structural basis for IRP2-RNA interactions and reveal an iron-independent mechanism for regulating iron homeostasis through the redox regulation of IRP2 cysteines.
Zumbrennen
Kimberly B
KB
Department of Oncological Sciences, University of Utah, Salt Lake City, UT 84112, USA.
Wallander
Michelle L
ML
Romney
S Joshua
SJ
Leibold
Elizabeth A
EA
D000818:Animals; D015703:Antigens, CD / Q000235:genetics*; D001665:Binding Sites; D003545:Cysteine / Q000378:metabolism*; D006706:Homeostasis; D006801:Humans; D035942:Iron Regulatory Protein 2 / Q000378:metabolism*; D051379:Mice; D010084:Oxidation-Reduction; D018384:Oxidative Stress; D012333:RNA, Messenger / Q000032:analysis; D016601:RNA-Binding Proteins / Q000378:metabolism*; D011990:Receptors, Transferrin / Q000235:genetics*
D015703:Antigens, CD; C068126:CD71 antigen; D012333:RNA, Messenger; D016601:RNA-Binding Proteins; D011990:Receptors, Transferrin; D035942:Iron Regulatory Protein 2; D003545:Cysteine