MOL PHARM #051888 Knock-In Mouse Lines Expressing either Mitochondrial or Microsomal CYP1A1: Differing Responses to Dietary Benzo[a]pyrene as Proof-of-Principle

Historically CYP1A1 protein is known to be located in the endoplasmic reticulum (ER; microsomes). More recently, CYP1A1 was shown to be targeted to the inner mitochondrial membrane; mitochondrial import is dependent on NH2-terminal processing that exposes a cryptic targeting signal. Intriguingly, microsomal and mitochondrial CYP1A1 enzymes exhibit different substrate specificities, electron donors, and inducer properties. To understand the physiological functions of microsomal versus mitochondrial CYP1A1, we have generated three knock-in lines by altering the CYP1A1 NH2-terminus. Cyp1a1(mtt/mtt) mice encode an NH2-terminal 31-amino acidtruncated protein, deleting the ER-targeting signal and exposing the cryptic mitochondrial-targeting signal. Cyp1a1(mtp/mtp) mice encode a protein carrying Leu7Asn and Leu17Asn mutations; this mutant lacks the signal recognition particle (SRP)-binding site and subsequent ER-targeting, but requires proteolysis by a cytosolic peptidase for mitochondrial import. Cyp1a1(mc/mc) mice encode a microsomal protein having Arg34Asp and Lys39Ile mutations which abolish the mitochondrial targeting signal. Following dioxin or β-naphthoflavone treatment of these mouse lines, the CYP1A1 protein was shown to be located in mitochondria of the Cyp1a1(mtp/mtp) and Cyp1a1(mtt/mtt) lines and in microsomes of the Cyp1a1(mc/mc) line. To test for differences in function, we compared the response to dietary benzo[a]pyrene (BaP). After 18 days of daily oral BaP, wild-type and Cyp1a1(mc/mc) mice were completely protected, whereas Cyp1a1(-/-) and Cyp1a1(mtp/mtp) mice showed striking toxicity and compensatory up-regulation of CYP1A2 and CYP1B1 mRNA in several tissues. Our data support the likelihood that it is the microsomal rather than mitochondrial CYP1A1 enzyme that protects against oral BaP toxicity. This article has not been copyedited and formatted. The final version may differ from this version. Molecular Pharmacology Fast Forward. Published on December 1, 2008 as DOI: 10.1124/mol.108.051888 at A PE T Jornals on A ril 8, 2017 m oharm .aspeurnals.org D ow nladed from