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J. Biol. Chem., Vol. 259, Issue 16, 10145-10149, Aug, 1984

Titration of mRNAs for cytochrome P-450c and P-450d under drug- inductive conditions in rat livers by their specific probes of cloned DNAs

K Kawajiri, O Gotoh, Y Tagashira, K Sogawa and Y Fujii-Kuriyama

By sequence analysis of the cloned cDNA or genomic DNA, we have recently deduced the complete primary structures of two forms of 3- methylcholanthrene-inducible cytochromes P-450 (P-450c and P-450d). Comparing these sequences, we identified two highly conserved regions, amino acid numbers from 35 to 200 and from 340 to 470. The nucleotide sequences corresponding to these homologous regions are also well conserved, whereas other regions have undergone considerable sequence divergence. In RNA blot analysis with unfractionated mRNA isolated from 3-methylcholanthrene-treated rat livers, Probe A (specific to P-450c sequence) hybridized with mRNA around 23 S, while Probe B (specific to P-450d sequence) hybridized with mRNA around 18 S. When common sequence between P-450c and P-450d was used as the probes (Probe C or D), two bands were clearly observed around 23 and 18 S mRNAs. With the common DNA sequence between P-450c and P-450d as a probe (93.7% homology), we studied the induction of specific mRNA for P-450c and P-450d by a single dose of several chemical compounds to rats. 3-Methylcholanthrene increased both P-450c and P-450d mRNA levels by 50 and 10 times above the control at 17 h after the administration, respectively. Despite the lower induction rates, the P-450d mRNA level was constantly higher than or at least similar to that of P-450c mRNA. beta-Naphthoflavone and Kaneclor KC 500 showed similar induction ability to 3- methylcholanthrene. On the other hand, isosafrole induced P-450d mRNA to a much greater extent than P-450c mRNA.
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