IRON REGULATES THE INTRACELLULAR DEGRADATION OF IRON REGULATORY PROTEIN-2 BY THE PROTEASOME

Iron regulatory proteins (IRP1 and IRP2) are RNA-binding proteins that bind to specific structures, termed iron responsive elements (IREs), that are located in the 5'- or 3'-untranslated regions of mRNAs that e ncode proteins involved in iron homeostasis. IRP1 and IRP2 RNA binding activities are regulated by iron; IRP1 and IRP2 bind IREs with high a ffinity in iron-depleted cells and with low affinity in iron-repleted cells. The decrease in IRP1 RNA binding activity occurs by a switch be tween apoprotein and 4Fe-4S forms, without changes in IRP1 levels, whe reas the decrease in IRP2 RNA binding activity reflects a reduction in IRP2 levels. To determine the mechanism by which iron decreases IRP2 levels, we studied IRP2 regulation by iron in rat hepatoma and human H eLa cells. The iron-dependent decrease in IRP2 levels was not due to a decrease in the amount of IRP2 mRNA or to a decrease in the rate of I RP2 synthesis. Pulse chase experiments demonstrated that iron resulted in a 3-fold increase in the degradation rate of IRP2. IRP2 degradatio n depends on protein synthesis, but not transcription, suggesting a re quirement for a labile protein. IRP2 degradation is not prevented by l ysosomal inhibitors or calpain II inhibitors, but is prevented by inhi bitors that block proteasome function. These data suggest the involvem ent of the proteasome in iron-mediated IRP2 proteolysis.