Gozukara E M, Fagan J, Pastewka J V, Guengerich F P, Gelboin H V
Arch Biochem Biophys. 1984 Aug 1;232(2):660-9. doi: 10.1016/0003-9861(84)90586-1.
Cytochrome P-450 mRNAs were quantitated by in vitro translation of liver RNA followed by immunoprecipitation with antibodies specific for cytochromes P-450. The kinetics of cytochrome P-450 mRNA induction by 3-methylcholanthrene and by phenobarbital were examined, and differences in the types of cytochrome P-450 mRNAs induced by 3-methylcholanthrene (MC), beta-naphthoflavone (BNF), and phenobarbital (PB) in male and female rats were determined. Phenobarbital strongly induced the mRNA encoding a single peptide antigenically related to the phenobarbital-induced cytochrome P-450PB-B. Male rat liver contained 62% more translatable mRNA for this peptide than did female. 3-Methylcholanthrene and beta-naphthoflavone, but not phenobarbital, induced mRNAs encoding three peptides that were immunologically related to cytochrome P-450BNF/MC-B, which is induced by 3-methylcholanthrene. The levels of translatable mRNA coding for these peptides were twice as high in females as in males. Striking sex differences were observed in the levels of translatable mRNAs for peptides related to cytochrome P-450PB/PCN-E, which is induced by phenobarbital and by pregnenolone-16-alpha-carbonitrile. In females, only RNA preparations from the livers of phenobarbital-treated rats had significant levels of mRNAs encoding these peptides. In contrast, significant levels of these RNAs were observed even in untreated males, and the levels of these mRNAs increased markedly following phenobarbital treatment. All cytochrome P-450 inducers examined caused a 50 to 70% decrease in translatable albumin mRNA. This effect was specific for albumin mRNA, since levels of total translatable mRNA were not generally altered by these inducers. The kinetics of induction of cytochrome P-450 mRNA differed from those of induction of aryl hydrocarbon hydroxylase (AHH) activity. Translatable cytochrome P-450 mRNA was increased as early as 4 h after phenobarbital treatment, peaked between 24 and 36 h, and dropped back to control levels by 120 h. The induction of AHH lagged behind the increase in translatable mRNA, remaining at control levels well after levels of translatable mRNA began to increase but then decreasing roughly in parallel with translatable mRNA. These findings suggest that transcription was not rate limiting for regulation of PB-inducible cytochrome P-450 activity. 3-Methylcholanthrene caused parallel increases in AHH activity and translatable cytochrome P-450 mRNA, but when translatable mRNA began to decrease after about 24 h, AHH activity remained high, suggesting that this P-450 mRNA was less stable than the enzyme for which it coded.
通过对肝脏RNA进行体外翻译，然后用细胞色素P - 450特异性抗体进行免疫沉淀，对细胞色素P - 450 mRNA进行定量分析。研究了3 - 甲基胆蒽和苯巴比妥诱导细胞色素P - 450 mRNA的动力学，并确定了雄性和雌性大鼠中由3 - 甲基胆蒽（MC）、β - 萘黄酮（BNF）和苯巴比妥（PB）诱导的细胞色素P - 450 mRNA类型的差异。苯巴比妥强烈诱导编码一种与苯巴比妥诱导的细胞色素P - 450PB - B抗原相关的单一肽的mRNA。雄性大鼠肝脏中该肽的可翻译mRNA比雌性多62%。3 - 甲基胆蒽和β - 萘黄酮而非苯巴比妥诱导编码三种与细胞色素P - 450BNF/MC - B免疫相关的肽的mRNA，细胞色素P - 450BNF/MC - B由3 - 甲基胆蒽诱导。这些肽的可翻译mRNA水平在雌性中是雄性的两倍。在与苯巴比妥和孕烯醇酮 - 16 - α - 腈诱导的细胞色素P - 450PB/PCN - E相关的肽的可翻译mRNA水平上观察到显著的性别差异。在雌性中，仅苯巴比妥处理大鼠肝脏的RNA制剂具有编码这些肽的显著水平的mRNA。相反，即使在未处理的雄性中也观察到这些RNA的显著水平，并且在苯巴比妥处理后这些mRNA的水平显著增加。所有检测的细胞色素P - 450诱导剂均导致可翻译的白蛋白mRNA降低50%至70%。这种效应是白蛋白mRNA特有的，因为这些诱导剂通常不会改变总可翻译mRNA的水平。细胞色素P - 450 mRNA的诱导动力学与芳烃羟化酶（AHH）活性的诱导动力学不同。可翻译的细胞色素P - 450 mRNA早在苯巴比妥处理后4小时就增加，在24至36小时达到峰值，并在120小时回落至对照水平。AHH的诱导滞后于可翻译mRNA的增加，在可翻译mRNA水平开始增加后很长时间保持在对照水平，但随后大致与可翻译mRNA平行下降。这些发现表明转录不是PB诱导的细胞色素P - 450活性调节的限速步骤。3 - 甲基胆蒽导致AHH活性和可翻译的细胞色素P - 450 mRNA平行增加，但当可翻译mRNA在约24小时后开始下降时，AHH活性仍然很高，这表明这种P - 450 mRNA比其编码的酶更不稳定。
