Ribosomal pausing and scanning arrest as mechanisms of translational regulation from cap-distal iron-responsive elements.
Iron regulatory protein 1 (IRP-1) binding to an iron-responsive element (IRE) located close to the cap structure of mRNAs represses translation by precluding the recruitment of the small ribosomal subunit to these mRNAs. This mechanism is position dependent; reporter mRNAs bearing IREs located further downstream exhibit diminished translational control in transfected mammalian cells. To investigate the underlying mechanism, we have recapitulated this position effect in a rabbit reticulocyte cell-free translation system. We show that the recruitment of the 43S preinitiation complex to the mRNA is unaffected when IRP-1 is bound to a cap-distal IRE. Following 43S complex recruitment, the translation initiation apparatus appears to stall, before linearly progressing to the initiation codon. The slow passive dissociation rate of IRP-1 from the cap-distal IRE suggests that the mammalian translation apparatus plays an active role in overcoming the cap-distal IRE-IRP-1 complex. In contrast, cap-distal IRE-IRP-1 complexes efficiently repress translation in wheat germ and yeast translation extracts. Since inhibition occurs subsequent to 43S complex recruitment, an efficient arrest of productive scanning may represent a second mechanism by which RNA-protein interactions within the 5' untranslated region of an mRNA can regulate translation. In contrast to initiating ribosomes, elongating ribosomes from mammal, plant, and yeast cells are unaffected by IRE-IRP-1 complexes positioned within the open reading frame. These data shed light on a characteristic aspect of the IRE-IRP regulatory system and uncover properties of the initiation and elongation translation apparatus of eukaryotic cells.
Paraskeva
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E
European Molecular Biology Laboratory, D-69117 Heidelberg, Germany.
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Schläger
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Wehr
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Hentze
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D000818:Animals; D002457:Cell Extracts; D005786:Gene Expression Regulation; D035941:Iron Regulatory Protein 1; D035925:Iron-Regulatory Proteins; D007506:Iron-Sulfur Proteins / Q000378:metabolism*; D016366:Open Reading Frames; D010442:Peptide Chain Initiation, Translational; D014176:Protein Biosynthesis; D012315:RNA Caps; D016601:RNA-Binding Proteins / Q000378:metabolism*; D011817:Rabbits; D020218:Response Elements; D012156:Reticulocytes; D012270:Ribosomes
D002457:Cell Extracts; D035925:Iron-Regulatory Proteins; D007506:Iron-Sulfur Proteins; D012315:RNA Caps; D016601:RNA-Binding Proteins; D035941:Iron Regulatory Protein 1