Organization and differential expression of two highly similar genes in the rabbit alcohol-inducible cytochrome P-450 subfamily.

The exon-intron organization of two rabbit genes that hybridize with cytochrome P-450 3a (P-450ALC) cDNA has been determined by restriction mapping and sequence analysis. Gene 1 encodes cytochrome P-450 3a as judged by the complete identity of its coding nucleotide sequence with P-450 3a cDNA. Gene 2 encodes a previously uncharacterized cytochrome P-450 that is 97% identical in primary structure to P-450 3a, with 16 amino acid differences scattered throughout the protein. Genes 1 and 2, which are 10 and 9 kilobases in length, respectively, are comprised of 9 exons with exon-intron junctions occurring at identical positions along the mRNA sequences. Each gene contains two transcription start sites approximately 27 and 33 nucleotides upstream from the translation initiation codon, as determined by primer extension and S1 nuclease protection experiments. The predicted lengths of gene 1 and 2 transcripts from the first transcription start site to the poly(A) attachment site are 1999 and 1660 nucleotides, respectively. This difference in size is primarily the result of a 338-base pair deletion in the 3' nontranslated portion of the gene 2 transcript relative to that of gene 1. The two genes show considerable similarity in their 5' flanking regions, including a "TATAA" transcriptional promoter element at position -28. However, a 32-base pair element that is repeated in gene 1 is present only as a single inexact copy in gene 2. By use of synthetic oligonucleotides as hybridization probes, gene 2 transcripts were shown to be present in poly(A)+ RNA from untreated rabbit liver at approximately 50% of P-450 3a mRNA levels. In kidney, however, no gene 2 mRNA was detected although 3a mRNA was present at approximately 10% of the level in liver.