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 ORF0) to a second +1 open reading frame (ORF1). Our studies show that this frame-shifting, which occurs at maximal efficiency of approximately 20%, is stimulated by polyamines and requires the functional integrity of the stop codon (stop-1) of ORF0. 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.
Rom
E
E
Department of Molecular Genetics and Virology, Weizmann Institute of Science, Rehovot, Israel.
Kahana
C
C
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D015971:Gene Expression Regulation, Enzymologic; D066298:In Vitro Techniques; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009955:Ornithine Decarboxylase / Q000235:genetics*; D010442:Peptide Chain Initiation, Translational; D010443:Peptide Chain Termination, Translational; D011073:Polyamines / Q000494:pharmacology*; D014176:Protein Biosynthesis / Q000187:drug effects*; D011506:Proteins / Q000235:genetics*; D012333:RNA, Messenger / Q000235:genetics; D051381:Rats; D012270:Ribosomes / Q000378:metabolism; D013329:Structure-Activity Relationship
D011073:Polyamines; D011506:Proteins; D012333:RNA, Messenger; C031017:ornithine decarboxylase antizyme; D009955:Ornithine Decarboxylase