Regulation of interaction of the iron-responsive element binding protein with iron-responsive RNA elements.
The 5' untranslated region of the ferritin heavy-chain mRNA contains a stem-loop structure called an iron-responsive element (IRE), that is solely responsible for the iron-mediated control of ferritin translation. A 90-kilodalton protein, called the IRE binding protein (IRE-BP), binds to the IRE and acts as a translational repressor. IREs also explain the iron-dependent control of the degradation of the mRNA encoding the transferrin receptor. Scatchard analysis reveals that the IRE-BP exists in two states, each of which is able to specifically interact with the IRE. The higher-affinity state has a Kd of 10 to 30 pM, and the lower affinity state has a Kd of 2 to 5 nM. The reversible oxidation or reduction of a sulfhydryl is critical to this switching, and the reduced form is of the higher affinity while the oxidized form is of lower affinity. The in vivo rate of ferritin synthesis is correlated with the abundance of the high-affinity form of the IRE-BP. In lysates of cells treated with iron chelators, which decrease ferritin biosynthesis, a four- to fivefold increase in the binding activity is seen and this increase is entirely caused by an increase in high-affinity binding sites. In desferrioxamine-treated cells, the high-affinity form makes up about 50% of the total IRE-BP, whereas in hemin-treated cells, the high-affinity form makes up less than 1%. The total amount of IRE-BP in the cytosol of cells is the same regardless of the prior iron treatment of the cell. Furthermore, a mutated IRE is not able to interact with the IRE-BP in a high-affinity form but only at a single lower affinity Kd of 0.7 nM. Its interaction with the IRE-BP is insensitive to the sulfhydryl status of the protein.
Haile
D J
DJ
Cell Biology and Metabolism Branch, National Institute of Child Health and Human Development, Bethesda, Maryland 20892.
Hentze
M W
MW
Rouault
T A
TA
Harford
J B
JB
Klausner
R D
RD
D001483:Base Sequence; D002352:Carrier Proteins / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel; D005293:Ferritins / Q000235:genetics*; D006801:Humans; D007501:Iron / Q000494:pharmacology*; D007502:Iron Chelating Agents / Q000494:pharmacology; D008969:Molecular Sequence Data; D009154:Mutation; D010084:Oxidation-Reduction; D014176:Protein Biosynthesis; D012333:RNA, Messenger / Q000378:metabolism*; D012097:Repressor Proteins / Q000378:metabolism; D013438:Sulfhydryl Compounds / Q000378:metabolism; D014407:Tumor Cells, Cultured
D002352:Carrier Proteins; D007502:Iron Chelating Agents; D012333:RNA, Messenger; D012097:Repressor Proteins; D013438:Sulfhydryl Compounds; D005293:Ferritins; D007501:Iron