Iron regulates cytoplasmic levels of a novel iron-responsive element-binding protein without aconitase activity

Iron-responsive element-binding proteins (IRE-BPs) are cytosolic proteins that bind to a conserved RNA stem-loop, termed the iron-responsive element (IRE), that is located in the 5'- or 3'-untranslated regions of mRNAs involved in iron metabolism. Binding of the IRE-BP to 5'-IREs represses translation, whereas binding to 3'-IREs stabilizes the mRNA. The previously identified IRE-BP (BP1) contains a 4Fe-4S cluster and has sequence homology to mitochondrial aconitase. The 4Fe-4S cluster is important for iron-dependent regulation: BP1 containing iron has low affinity for the IRE and contains aconitase activity, whereas BP1 lacking iron has high affinity for the IRE, but lacks aconitase activity. A second IRE-BP (BP2) has been identified in rat tissues and cells and exhibits many of the hallmarks of an IRE-BP, including binding to the IRE and functioning as a translational repressor of IRE-containing RNAs. BP1 and BP2 RNA binding activities are decreased in extracts from cells treated with iron, indicating that BP1 and BP2 are negatively regulated by iron. Although BP1 and BP2 share similar characteristics, they differ in two significant ways. Unlike BP1 levels, which do not change when RNA binding activity decreases in response to iron, BP2 decreases to undetectable levels in extracts from cells treated with iron; and unlike BP1, BP2 does not have aconitase activity. These data indicate that BP1 and BP2 are distinct proteins that have similar specificity for IRE binding and that function similarly in translation, but are regulated by iron via different mechanisms.