Disease relevance of Cyp6a2

• To determine the impact of each mutation on the function of CYP6A2, the wild-type enzyme (CYP6A2wt) of Cyp6a2 was expressed in Escherichia coli as well as three variants carrying a single mutation, the double mutant CYP6A2vSV and the triple mutant CYP6A2vSVL [1].
• Heterologous expression of the CYP6A2 protein in lepidopteran cells infected with a Cyp6a2-recombinant baculovirus was observed by Western blotting of cell lysates and by spectral characterization of the reduced-CO complex of the P450 [2].

High impact information on Cyp6a2

• The cytochrome P450 Cyp6a2 and the diazepam-binding inhibitor gene (Dbi) were over transcribed in the two DDT-resistant genotypes when compared to the wild-type Drosophila, and this difference was significant at the most stringent statistical level, a Bonferroni correction [3].
• The P450-B1 gene in 91-C contained a solitary long terminal repeat of transposable element 17.6 in its 3' untranslated region [4].
• P450-A is present among all strains tested, whereas expression of P450-B is associated with resistance to insecticides [4].
• On the basis of features of the long terminal repeat and its location in the gene of the susceptible fly, we propose that a posttranscriptional mechanism involving mRNA stability could be involved in regulating P450-B1 gene expression [4].
• Four of these exons (6a1, 6a2, 6b1, and 6b2) are mutually exclusive and very similar in sequence [5].

Biological context of Cyp6a2

• Taken together, these results suggest that the second chromosome-linked Cyp6a2 and Cypa8 genes are regulated by loci present on the third chromosome, and the wild-type function of these loci is to repress these two Cyp genes [6].
• This suggested that the underproducer strains carry loci which downregulate Cyp6a2 and Cyp6a8 gene expression [6].
• The polypeptides encoded by these two genes, Cyp6a8 and Cyp6a9, show 77 and 75% amino acid sequence similarity, and 60 and 55% identity with Cyp6a2 of D. melanogaster, respectively [7].
• DNA from the Cyp6a2 promoter region was functional when linked to a luciferase reporter gene and transfected into D. melanogaster Schneider cells [2].
• Promoter activity of the -11/-761 DNA of the overproducer 91-R strain was found to be 4-fold lower in the genome of underproducer ry(506) strain, which is wild type for the putative repressor gene, than in the genome of F1 hybrids of 91-R and ry(506) strains [8].

Anatomical context of Cyp6a2

• The encoded protein had been isolated as an ecdysone-binding cytochrome P450 from microsomes of the same tissue in previous work [9].
• Here, we report the cloning and sequencing of a full-length cDNA of a new cytochrome P450, classified as CYP6H1, from malpighian tubules of the locust, Locusta migratoria [9].
• Baculovirus-mediated expression of the CYP6B4v1 protein in lepidopteran cell lines demonstrates that this P450 isozyme metabolizes isopimpinellin, imperatorin, and bergapten at high rates, xanthotoxin and psoralen at intermediate rates and angelicin, sphondin, and trioxsalen only at very low rates [10].

Associations of Cyp6a2 with chemical compounds

• The Cyp6a2 gene appears to be a suitable model for a genetic analysis of the phenobarbital induction process [2].
• Endogenous Cyp6a8 and Cyp6a2 genes in these flies also showed caffeine-induced expression [11].
• Partial reconstitution of 7-ethoxycoumarin de-ethylase activity was achieved using rabbit NADPH: cytochrome P450 reductase and purified preparations containing P450-B [12].
• Significant purification of the ubiquitous cytochrome P-450-A and the strain-specific P-450-B from Drosophila melanogaster has been achieved by sequential chromatography on octylamino-agarose, DEAE-cellulose and hydroxyapatite [12].
• The level of expression of the Cyp6a2 gene is much higher in the DDT-resistant 91-R strain than in the susceptible 91-C strain of Drosophila melanogaster (Waters et al. (1992b) Proc. Natl. Acad. Sci. USA, 89, 4855-4859) [7].

Regulatory relationships of Cyp6a2

• Analysis of Cyp6a8 upstream DNA of the overproducer 91-R strain showed that DNA sequences between -199 and -761 bp are required for the highest constitutive and barbital-induced expression of Cyp6a8 [8].

Other interactions of Cyp6a2

• To examine whether caffeine, the most widely used xenobiotic compound, would induce insect cytochrome P450 or CYP gene expression, upstream DNA fragments of Cyp6a2 (0.12, 0.26, 0.52 and 0.98-kb) and Cyp6a8 (0.06, 0.1, 0.2, 0.5 and 0.8-kb) genes of Drosophila melanogaster were individually fused to the firefly luciferase (luc) reporter gene [11].

Analytical, diagnostic and therapeutic context of Cyp6a2

• Induction of two cytochrome P450 genes, Cyp6a2 and Cyp6a8, of Drosophila melanogaster by caffeine in adult flies and in cell culture [11].
• Preparations enriched for P-450-B (specific contents of 4-7 nmol/mg) contained significant amounts of P-450-A but were essentially free of other proteins as judged by SDS/PAGE [12].
• Using northern and western blot analysis we observed the expression pattern of these two genes in two DDT susceptible (Canton-S and 91-C) and three DDT resistant strains (Wisconsin, 91-R and Hikone-R) [13].
• Sequence alignments with CYP102A1 provided clues to the positions of the amino acids mutated in CYP6A2 [1].
• Degenerate primers corresponding to conserved amino acid sequences in three insect P450s, Musca domestica (CYP6A1), Drosophila melanogaster (CYP6A2) and Papilio polyxenes (CYP6B1), were used to clone xanthotoxin-induced P450 transcripts from P. glaucus larvae by a reverse transcription-polymerase chain reaction (RT-PCR) strategy [10].

References