CLONING AND CHARACTERIZATION OF A RAT GENE ENCODING ORNITHINE DECARBOXYLASE ANTIZYME

We cloned an ornithine decarboxylase antizyme-encoding gene (Oaz) from a rat liver genomic library. The entire gene was located on a 4367-bp EcoRI fragment, which corresponded to one of two fragments hybridizable with the antizyme-encoding cDNA, Z1, on Southern blot analysis. Sequence analysis of the cloned gene showed that it consisted of five exons which were identical with the cDNA. The transcription start points of the Oaz mRNA were located 75 and 76 nucleotides upstream from the first ATG codon, as determined by S1 nuclease protection and primer extension analyses. The 5'-flanking region of the gene contained typical promoter motifs, such as a TATA box and Spl-binding sites. Introduction of a chimeric gene consisting of the 5'-flanking region and the bacterial cat gene into Chinese hamster ovary cells revealed a promoter activity in the region, which was comparable in strength to that of the simian virus 40 promoter. In addition, we isolated a 12-kb EcoRI fragment, the other sequence hybridizable to the cDNA. Sequence analysis showed that it represented a processed Oaz pseudogene and was not able to encode any active protein.