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 (IR E) located close to the cap structure of mRNAs represses translation by pre cluding the recruitment of the small ribosomal subunit to these mRNAs, This mechanism is position dependent; reporter mRNAs bearing IREs located furth er downstream exhibit diminished translational control in transfected mamma lian 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 comple x recruitment, the translation initiation apparatus appears to stall, befor e linearly progressing to the initiation codon. The slow passive dissociati on rate of IRP-1 from the cap-distal IRE suggests that the mammalian transl ation apparatus plays an active role in overcoming the cap-distal IRE-IRP-1 complex. In contrast, cap-distal IRE IRP-1 complexes efficiently repress t ranslation in wheat germ and yeast translation extracts. Since inhibition o ccurs subsequent to 43S complex recruitment, an efficient arrest of product ive scanning may represent a second mechanism by which RNA-protein interact ions within the 5' untranslated region of an mRNA can regulate translation. In contrast to initiating ribosomes, elongating ribosomes from mammal, pla nt, and yeast cells are unaffected by IRE-IRP-1 complexes positioned within the open reading frame. These data shed light on a characteristic aspect o f the IRE-IRP regulatory system and uncover properties of the initiation an d elongation translation apparatus of eukaryotic cells.