Polyamines Regulate the Expression of Ornithine Decarboxylase Antizyme in vitro by Inducing Ribosomal Frame-Shifting

• Published in: Proceedings of the National Academy of Sciences - PNAS Vol. 91; no. 9; pp. 3959 - 3963
• Main Authors: Rom, Eran, Kahana, Chaim
• Format: Journal Article
• Language: English
• Published: United States National Academy of Sciences of the United States of America 26.04.1994; National Acad Sciences; National Academy of Sciences
• Subjects: Amino Acid Sequence; Animals; Base Sequence; Biochemistry; Codons; Complementary DNA; Gene expression regulation; Gene Expression Regulation, Enzymologic; In Vitro Techniques; Mammals; Messenger RNA; Molecular Sequence Data; Mutagenesis, Site-Directed; Open reading frames; Ornithine Decarboxylase - genetics; Peptide Chain Initiation, Translational; Peptide Chain Termination, Translational; Polyamines; Polyamines - pharmacology; Protein Biosynthesis - drug effects; Proteins - genetics; Rats; Reading frames; Reticulocytes; Ribonucleic acid; Ribosomes - metabolism; RNA; RNA, Messenger - genetics; Stop codon; Structure-Activity Relationship
• ISSN: 0027-8424; 1091-6490
• DOI: 10.1073/pnas.91.9.3959

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 ≈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.