| Humans are exposed to a variety of inhaled and systemic xenobiotic compounds that cause respiratory toxicity. Cytochrome P450 enzymes, expressed specifically in lung{09} tissues, are known to bioactivate these xenobiotics to reactive pneumotoxic and carcinogenic intermediates. Thus, differential, tissue-specific expression of cytochrome P450 genes may lead to interindividual susceptibilities to pneumotoxicants. Therefore, characterization of lung P450 enzymes is an important goal. The CYP2F1 gene is transcribed specifically in lung tissues, and its gene product bioactivates several prototypical pneumotoxicants, but the mechanisms regulating the pulmonary-specific transcription of CYP2F1 are not known. Therefore, the hypothesis of this dissertation was that transcription of CYP2F1 in pulmonary tissues is controlled by cis-regulatory elements that respond to constitutive or tissue-specific trans-activating factors. The following objectives were performed: (1) identified human genomic clones that contain the regulatory region of CYP2F1 , characterized the intron/exon boundaries, and determined the transcriptional start site; (2) identified potential transcription factor-binding sites in the promoter region of the CYP2F1 gene; and (3) utilized deletion reporter constructs with lung and liver cells to identify functional regulatory regions of the CYP2F1 gene and characterized potential cis-regulatory elements using DNA-binding assays. The structure of the CYP2F1 gene, including the promoter region, which was specifically active in human lung epithelial cells, was elucidated. Furthermore, the consensus sequence of a lung-specific factor(s)-binding site in the promoter of the CYP2F1 gene was characterized, and the contributions of the ubiquitous nuclear factors, Sp1 and Sp3, to basal promoter activity in lung epithelial cells were presented. Additional studies documented the exacerbation of cell death when CYP2F1 was over-expressed in human lung cells, and identified the unique 3-methylindole methyl oxidation pathway by another lung-specific P450, CYP4B2. In summary, this dissertation provided novel information concerning the function and transcriptional regulation of P450 enzymes in the lung and established a molecular basis for the biochemical mechanisms regulating the pulmonary-specific expression of the CYP2F1 gene. |
