|Abstract:||Cytochromes P-450 are a superfamily of monooxygenases which metabolize a myriad of endogenous and exogenous substances. Cytochrome P-450 genes are differentially expressed in various tissues and differentially induced by many chemical agents. The rabbit cytochrome P-450IIC subfamily contains both phenobarbital inducible and constitutive genes. Comparative studies of the gene structures of this subfamily may shed light on the mechanism of differential induction of cytochrome P-450 genes. The purpose of this project is to characterize the cytochrome P-450IIC3 gene, which is a constitutive gene with respect to phenobarbital, and the 5$\sp\prime$-flanking region of the P-450IIC1 gene which is induced by phenobarbital. Two rabbit liver DNA genomic libraries were screened for the presence of the cytochrome P-450IIC3 and IIC1 genes. DNA from genomic clones C3.1, C3.2, C3.3, and C1.1 were isolated, and subcloned into phage M13 mp11 and phagemid pTZ18R for sequencing by the chain termination procedure. Exons 7, 8, and 9 of the P-450IIC3 gene were contained within genomic clone C3.1, exons 2, 3, 4, 5, and 6 were in clone C3.2, and exons 1, and 2 were in clone C3.3. Only two differences between the gene and the cDNA nucleotide sequences were observed, one of which resulted an amino acid change. The total length of the cytochrome P-450IIC3 gene is more than 25 kilobases. All the intron/exon junctions obey the GT/AG rule. The derived amino acid sequence has nine differences from the direct protein sequence of rabbit cytochrome P-450 3b of Ozols et al. All the intron positions of the rabbit P-450IIC3 and rat P-450e gene (a member of P-450IIB subfamily) are identical. This provides further evidence for divergent evolution by gene duplication within the cytochrome P-450 gene superfamily. The 5$\sp\prime$-flanking region and exon 1 of the P-450IIC1 gene were in clone C1.1. The intron 1 position and intron/exon junction were the same as in the P-450IIC3 gene. The 5$\sp\prime$-flanking regions of both P-450IIC1 and IIC3 contained TATA boxes which were about 25 bp upstream from the RNA start sites as determined by primer extension. The potential regulatory sequence modules included a CCAAT sequence and binding sites similar to the liver-specific factor HNF-1 and the general transcription factors, AP-1 and OCT.