Constitutive and Inducible Cytochromes P450 in Rat Lung Mitochondria: Xenobiotic Induction, Relative Abundance, and Catalytic Properties

Abstract

The presence of xenobiotic-inducible CYP1A1, 2B1/2, and 3A1/2 in rat lung mitochondria was investigated using mitochondrial preparations of defined purity. The mitochondrial P450 content in uninduced lung was 1.5-fold higher compared to microsomes. Administration of BNF induced the P450 contents by twofold in both mitochondrial and microsomal membrane fractions. BNF treatment induced EROD activity to about 40-fold in the microsomal fraction and 25-fold in the mitochondrial fraction. The microsomal induction was observed at 4 days of BNF treatment, while the mitochondrial induction required 10 days of treatment. Consistent with the activity profile, Western blot analysis showed the presence of CYP1A1 antibody reactive protein only in lung mitochondria from BNF-treated rats. BNF administration also caused a 50 to 80% reduction in the CYP2B1/2-associated PROD and BROD activities and CYP3A1/2-associated ERND activity in both mitochondria and microsomes. There was also a parallel reduction in the antibody reactive CYP2B1/2 and 3A1/2 proteins in both of these membrane fractions. Administration of DEX for 4 days induced mitochondrial and microsomal ERND activity by 1.7- and 2.5-fold, respectively. Mitochondrial EROD activity was inhibited by antibodies to P450MT2, as well as Adx, but not by antibody against P450 reductase, indicating the mitochondrial localization of CYP1A1. Protease protection and alkaline extraction experiments indicated that CYP1A1 associated with lung mitochondria is localized inside the inner membrane and exists as a membrane extrinsic protein. In summary, this is probably the first report of inducible P450s in rat lung mitochondria, and our results suggest a possible functional role for these mitochondrial enzymes in xenobiotic metabolism.