The Journal of biological chemistry
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Iron regulatory factor (IRF) is a cytoplasmic mRNA-binding protein with specificity for iron-responsive element (IRE) RNA stem-loops. IRF post-transcriptionally regulates intracellular iron levels via binding to IREs in the untranslated regions of ferritin, transferrin receptor, and erythroid 5-aminolevulinic-acid synthase mRNAs. Specific IRE nucleotides are phylogenetically conserved: those of the 6-base loop (5'-CAGUGN-3') and an unpaired "bulge" cytosine. ⋯ This novel finding predicts base pairing within the IRE loop between positions 1 and 5, thus facilitating the formation of a specific loop structure in which nucleotides at positions 2-4 are made accessible for protein interaction. Nucleotide substitution at these loop positions, or at the position of the bulge cytosine, decreased binding by 36-99%. In addition, we demonstrate a preferred IRE bulge structure and report a striking difference in the RNA binding specificity of rat IRF compared with that of the related IRE-binding protein, IRFB.