The iron-responsive element binding protein. Purification, cloning, and regulation in rat liver.
The iron-responsive element binding protein (IRE-BP) is a cytosolic protein that binds a highly conserved sequence in the untranslated regions of mRNAs involved in iron metabolism including ferritin, transferrin receptor, and erythroid 5-aminolevulinate acid synthase. This conserved sequence is termed the iron-responsive element and is necessary for the post-transcriptional regulation of these mRNAs by iron. The rat liver IRE-BP was purified to homogeneity by chromatographic methods and partial amino acid sequence was obtained. A cDNA was isolated from a rat liver cDNA library and sequenced. The amino acid sequence deduced from the cDNA sequence corresponds to a protein of 889 amino acids with a predicted molecular weight of 97.946. The NH2-terminal sequence obtained by Edman degradation matched the deduced amino acid sequence obtained from the cDNA, confirming the translational start site. Rat liver IRE-BP shares 95% identity with human IRE-BP and 98% identity with mouse IRE-BP indicating that the IRE-BPs have remained highly conserved during evolution. The 5'-untranslated region is at least 236 nucleotides and contains interesting structural features including two direct repeats, an inverted repeat, and three small open reading frames. The rat IRE-BP mRNA is approximately 3600 nucleotides and is expressed in a variety of rat tissues including liver, spleen, and gut. Over the course of 16 h following an intraperitoneal injection of iron in rats. IRE-BP RNA binding activity decreases to 50% of control levels. The decrease in IRE-BP RNA binding activity in extracts from iron-treated rats is reversible by pretreatment of the extracts with reducing agents. The steady-state levels of IRE-BP mRNA remain constant during iron treatment. These data suggest that the decrease in IRE-BP RNA binding activity by iron in rat liver is due to post-translational changes in the RNA binding affinity of the IRE-BP and not due a decrease in the transcription of the IRE-BP gene or to the destabilization of the IRE-BP mRNA.
Yu
Y
Y
Program in Human Molecular Biology and Genetics, University of Utah, Salt Lake City 84112.
Radisky
E
E
Leibold
E A
EA
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D015152:Blotting, Northern; D003001:Cloning, Molecular; D004591:Electrophoresis, Polyacrylamide Gel; D035925:Iron-Regulatory Proteins; D008099:Liver / Q000378:metabolism*; D008297:Male; D008969:Molecular Sequence Data; D012323:RNA Processing, Post-Transcriptional; D012333:RNA, Messenger / Q000378:metabolism; D016601:RNA-Binding Proteins / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism; D051381:Rats; D011919:Rats, Inbred Strains; D012689:Sequence Homology, Nucleic Acid
D035925:Iron-Regulatory Proteins; D012333:RNA, Messenger; D016601:RNA-Binding Proteins