Phenobarbital- and 3-methylcholanthrene-induced synthesis of two different molecular species of microsomal cytochrome P-450 in rat liver.

The syntheses of two different molecular species of cytochrome P-450, P-450(PB), and P-450(MC), were examined using normal and drug-treated rats, and the rate of synthesis was correlated with the drug-induced increase of the amounts in the liver microsomes of treated animals. Phenobarbital (PB) and 3-methylcholanthrene (MC) were used as inducers. The syntheses of cytochrome b5 and NADPH-cytochrome P-450 reductase (fpT) were also examined. In vivo incorporation of L-[3H]leucine indicated a large increase of P-450(PB) synthesis in the livers of PB-treated animals, which reached a plateau value about 18-fold higher than the control level at 12 h. The synthesis of fpT was also stimulated by PB showing a peak value of 3 times the control at 12 h after PB administration, but it returned to the control level afterwards. On the other hand, the syntheses of P-450(MC) and cytochrome b5 did not change at all. Similarly, MC administration selectively induced the synthesis of P-450(MC), which was about 24 times the control at 6 h, whereas those of P-450(PB), cytochrome b5, and fpT were not affected by MC. Analysis of nascent peptides and in vitro translation of polysomes and total liver RNA prepared from control and drug-treated animals were also carried out, and the results of these in vitro experiments confirmed the observations in in vivo incorporation studies. It was found that both free and membrane-bound ribosomes participate in the syntheses of P-450(PB) and P-450(MC) in the case of control animals. PB and MC induced the syntheses of P-450(PB) and P-450(MC) by bound ribosomes but not by free ribosomes, and the contribution of bound ribosomes to the syntheses of these two species of cytochrome P-450 was predominant in the case of drug-treated animals. The molecular sizes of in vitro synthesized P-450(PB) and P-450(MC) were the same as those of authentic samples prepared from liver microsomes.