title: Cellular regulation of the iron-responsive element binding protein: disassembly of the cubane iron-sulfur cluster results in high-affinity RNA binding.
abstract: The translation of ferritin mRNA and degradation of transferrin receptor mRNA are regulated by the interaction of an RNA-binding protein, the iron-responsive element binding protein (IRE-BP), with RNA stem-loop structures known as iron-responsive elements (IREs) contained within these transcripts. IRE-BP produced in iron-replete cells has aconitase (EC 4.2.1.3) activity. The protein shows extensive sequence homology with mitochondrial aconitase, and sequences of peptides prepared from cytosolic aconitase are identical with peptides of IRE-BP. As an active aconitase, IRE-BP is expected to have an Fe-S cluster, in analogy to other aconitases. This Fe-S cluster has been implicated as the region of the protein that senses intracellular iron levels and accordingly modifies the ability of the IRE-BP to interact with IREs. Expression of the IRE-BP in cultured cells has revealed that the IRE-BP functions either as an active aconitase, when the cells are iron-replete, or as an active RNA-binding protein, when the cells are iron-depleted. We compare properties of purified authentic cytosolic aconitase from beef liver with those of IRE-BP from tissue culture cells and establish that characteristics of the physiologically relevant form of the protein from iron-depleted cells resemble those of cytosolic aconitase apoprotein. We demonstrate that loss of the labile fourth iron atom of the Fe-S cluster results in loss of aconitase activity, but that more extensive cluster alteration is required before the IRE-BP acquires the capacity to bind RNA with the affinity seen in vivo. These results are consistent with a model in which the cubane Fe-S cluster is disassembled when intracellular iron is depleted.
mesh_terms: D000154:Aconitate Hydratase / Q000378:metabolism*; D000818:Animals; D001059:Apoproteins / Q000378:metabolism; D003676:Deferoxamine / Q000737:chemistry; D005292:Ferricyanides / Q000737:chemistry; D005293:Ferritins / Q000235:genetics; D006427:Hemin / Q000737:chemistry; D006801:Humans; D066298:In Vitro Techniques; D007501:Iron / Q000378:metabolism*; D035925:Iron-Regulatory Proteins; D007506:Iron-Sulfur Proteins / Q000378:metabolism*; D051379:Mice; D010084:Oxidation-Reduction; D012313:RNA / Q000378:metabolism*; D016601:RNA-Binding Proteins / Q000378:metabolism*; D013329:Structure-Activity Relationship; D014407:Tumor Cells, Cultured
chemical_list: D001059:Apoproteins; D005292:Ferricyanides; D035925:Iron-Regulatory Proteins; D007506:Iron-Sulfur Proteins; D016601:RNA-Binding Proteins; C007931:hexacyanoferrate III; D012313:RNA; D006427:Hemin; D005293:Ferritins; D007501:Iron; D000154:Aconitate Hydratase; D003676:Deferoxamine