Molecular cloning and sequence analysis of cDNAs coding for 3-methylcholanthrene-inducible cytochromes P450 in Xenopus laevis liver

Liver microsomes of Xenopus laevis were investigated for specific cytochrom e P450s (CYPs) that would be inducible in response to the administration of either 3-methylcholanthrene (3MC) or beta-naphthoflavone (BNF), potent ind ucers for mammalian CYP1A. When probed with antibodies raised against rat C YP1A1, a 54-kDa protein was detected after administration of polycyclic aro matic hydrocarbons. However, there was no immunoreactive protein in microso mes from untreated frogs. In order to obtain structural information about t his CYP1A-like protein, a liver cDNA library of 3MC-treated frog was constr ucted and screened using a fragment of rat CYP1A2 cDNA under Pow stringency conditions. We have isolated two cDNA clones (MC1 and MC2) with inherent f eatures of the CYP1A subfamily. The sequence determination revealed that bo th of them coded for polypeptides composed of 526 amino acid residues, whic h differed from each other by 30 amino acids. A comparison with other mamma lian CYP enzymes demonstrated that both of the sequences share 55 to 63% id entity with the sequences of CYP1A family members. Northern blot analysis a nd RT-PCR results further demonstrated that two discrete transcripts corres ponding to clones MC1 and MC2 are indeed inducible in the frog liver by tre atment with 3MC or BNF. The names CYP1A6 and CYP1A7 were given to clones MC 1 and MC2, respectively. (C) 1999 Academic Press.