TRANSCRIPTIONAL REGULATION OF CYTOCHROME P450 GENES

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Cytochrome P450s are regulated through inducer-dependent and tissue- specific pathways. Most P450s involved in metabolism of foreign chemicals including chemical carcinogens are expressed in liver, although some P450s are found in extrahepatic tissues such as lung, kidney and the gastrointestinal tract. Tissue-specific gene expression is governed through tissue-enriched transcription factors. The rat is being used as a model system to study liver-specific expression of P450 genes. Cotransfection trans-activation assays are being used to determine which liver-enriched transcription factors are involved in control of the CYP2D5 P450 gene promoter. These studies revealed that the liver- enriched factor C/EBP-beta and the general transcription factor Sp1 cooperate to control expression of CYP2D5. DNA binding studies suggest that Sp1 interacts with C/EBP-beta to increase its ability to bind to a segment of DNA upstream of the CYP2D5 gene. In the absence of Sp1, C/EBP-beta can not directly bind to this DNA element. Deletion and mutagenesis analysis are being used to determine which regions of C/EBP- beta and Sp1 are required for their cooperative interactions. To further understand the role of liver-enriched transcription factors in P450 gene expression, conditional knockout mice are being generated. Genomic clones to several liver-enriched transcription factors were isolated from an 129/SV gene library and used to prepare conditional knockout constructs that contain the Lox recombination signals flanking exonic regions of the genes. The recombinase gene Cre is being fused to the CYP2C6 promoter to generate a mouse that will express this protein in the liver at puberty. Thus, upon breeding the knockout and CYP2C6-Cre mice, the transcription factor genes will be abolished specifically in adult mice and the expression of P450 genes will be analyzed.