POLYAMINES REGULATE THE EXPRESSION OF ORNITHINE DECARBOXYLASE ANTIZYME IN-VITRO BY INDUCING RIBOSOMAL FRAME-SHIFTING

We provide here an example of a mammalian cellular gene expressed by frame-shifting. Conventional reading of the sequence of ornithine decarboxylase-antizyme mRNA (a protein that modulates the rate of ornithine decarboxylase degradation) results in premature termination at an in-frame termination codon (stop-1), located shortly after the initiation codon. By translating, in vitro in reticulocyte lysate, antizyme mRNA with a full coding capacity and various mutants derived from it, we demonstrate that antizyme expression requires that ribosomes shift from the first open reading frame (termed ORF(0)) to a second +1 open reading frame (ORF(1)). Our studies show that this frame-shifting, which occurs at maximal efficiency of approximate to 20%, is stimulated by polyamines and requires the functional integrity of the stop codon (stop-1) of ORF(0). By introducing in-frame deletions, we have shown that an 87-nt segment surrounding stop-1 enhances frame-shifting efficiency, whereas the 6 nt located just upstream to stop-1 are absolutely essential for this process. Because this segment does not contain sequences that were previously characterized as shifty segments, our results suggest that another mechanism of frame-shifting is involved in mediating antizyme expression.