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ItemThe regulation and function of the Drosophila melanogaster Cytochrome P450 gene, Cyp12d1Boey, Hui Kuang Adrian ( 2011)Cytochrome P450s are an important family of monooxygenase enzymes implicated in numerous xenobiotic detoxification events as well as in essential endogenous functions. The vinegar fly Drosophila melanogaster has 85 P450 genes; however, the large majority of them remain uncharacterised in terms of their function and regulation. Cyp12d1 is arguably the most xenobiotic inducible P450 gene in the D. melanogaster genome. It has been suggested that Cyp12d1 is an excellent candidate gene to study Drosophila xenobiotic induction pathways as it responds to a wide range of chemical inducers, indicating that it contains most if not all of the cis-regulatory elements needed for xenobiotic induction in Drosophila. Hence, the transcriptional regulation of Cyp12d1 was investigated to identify novel P450 induction pathways in D. melanogaster. Cyp12d1 basal transcriptional regulators were found in Cyp12d1 upstream and downstream regulatory regions, while enhancers for Phenobarbital and caffeine induction were located upstream. Site-directed mutagenesis experiments identified GATA family transcription factors as important Cyp12d1 midgut expression regulatory proteins. However, their role in xenobiotic induction remains unclear. Biochemical sequencing of electomobility-shift assay protein bands, and genetic RNAi screens of genes encoding other candidate transcription factors, failed to identify any other potential xenobiotic regulatory proteins. Cyp12d1 function was also investigated in this study. Cyp12d1 overexpression has been shown to confer resistance to the insecticides DDT and dicyclanil, but other functions have not been identified prior to this study. Adult Cyp12d1 functions were investigated through Cyp12d1 RNAi and overexpression studies. Cyp12d1 was found to be involved in adult longevity and oxidative stress resistance, suggesting other potential functions in addition to known detoxification functions. Cyp12d1 has been tandemly duplicated in D. melanogaster, and this duplication exists as a polymorphism in field populations. The geographical distribution of the Cyp12d1 duplication was examined in flies collected along the eastern coastline of Australia. The frequency of the duplicated Cyp12d1 gene was found to vary spatially, with flies in lower latitudes being more likely to possess the Cyp12d1 duplication and flies in higher latitudes being less likely. Cyp12d1 tissue-specific embryonic expression and mRNA transcript length was different in Cyp12d1-duplicated lines when compared to non-Cyp12d1 duplicated lines. These results indicate the Cyp12d1 duplication confers changes in Cyp12d1 expression patterns and suggest that Cyp12d1 may be involved in local adaptation to the microenvironment.