Regulation of cytochrome P450 expression in Drosophila: Genomic insights

Abstract

Genomic tools such as the availability of the Drosophila genome sequence, the relative ease of stable transformation, and DNA microarrays have made the fruit fly a powerful model in insecticide toxicology research. We have used transgenic promoter-GFP constructs to document the detailed pattern of induced Cyp6a2 gene expression in larval and adult Drosophila tissues. We also compared various insecticides and xenobiotics for their ability to induce this cytochrome P450 gene, and show that the pattern of Cyp6a2 inducibility is comparable to that of vertebrate CYP2B genes, and different from that of vertebrate CYP1A genes, suggesting a degree of evolutionary conservation for the "phenobarbital-type" induction mechanism. Our results are compared to the increasingly diverse reports on P450 induction that can be gleaned from whole genome or from "detox" microarray experiments in Drosophila. These suggest that only a third of the genomic repertoire of CYP genes is inducible by xenobiotics, and that there are distinct subsets of inducers / induced genes, suggesting multiple xenobiotic transduction mechanisms. A relationship between induction and resistance is not supported by expression data from the literature. The relative abundance of expression data now available is in contrast to the paucity of studies on functional expression of P450 enzymes, and this remains a challenge for our understanding of the toxicokinetic aspects of insecticide action.

Fig. 1

Phenobarbital-induced GFP in third instar larva and pupa. A: Early third instar larva. B: Brain-ganglion complex and anterior part of the digestive system. and C: Abdominal region showing body wall and internal organs. D and E: anterior and posterior ends of the larva. F: Anterior part of the salivary gland. G: Heart and pericardial cells. H: Anterior spiracle and spiracular glands. I: Body wall showing muscles. J and K: Anterior and posterior ends of control larva (without Phenobarbital). L: Newly formed pupa, treated with Phenobarbital at larval stage. M: Control pupa. Br, brain; Ed, epidermis; Es, esophagus; Fb, fat body; Gn, ganglion; He, heart; Hg, hindgut; Mg, midgut; Ms, muscles; Pv, proventriculus; Sbr, spiracular branches; So, sense organ; Sp, spiracle; Spg, spiracular glands; Tr, trachea.

Fig. 2

Phenobarbital (Pheno)- or PCB-treated adults. A: Female fly (Pheno). B and C: Apex of proboscis (frontal view), surface and interior (PCB). D: Brain, thoracic ganglion and anterior region of the digestive system (Pheno). E: Part of midgut and hindgut and proximal region of Malpighian tubules (PCB). F: Wing (PCB). G: Abdominal wall showing tergosternal muscles and epidermis (PCB). H: Hindgut and rectum (Pheno) I: Abdominal fat body, heart and pericardial cells (Pheno). J: Part of the salivary gland (Pheno). K: Part of male reproductive system (PCB). L–N Control female, and parts of the digestive system. AcGl, Accessory gland; Br, brain; Cr, crop; Ejd, ejaculatory duct; Fb, fatbody; He, heart; Hg, hindgut; Mg, midgut; Mt, malpighian tubule; Mu, muscles; Pc, pericardial cells; Pr, proventriculus; Pt, pseudotrachea; Re, rectum; Rv, rectal valve; SeCe, sensory cells; SeNe, sensory nerve; Sg, salivary gland Te, testis; Vde, vas deferens. Scale bar:D, 40 µm, E,F, H, K, M and N, 100 µm; B, C, G and I, 50µm; J, 10µm.

Fig. 3

Venn diagram summarizing the inducibility of Drosophila CYP genes by various chemicals. 1: ethanol; 2: pyrethrum; 3: phenobarbital; 4: caffeine; 5: piperonyl butoxide ; 6: Piper nigrum extract ; 7: atrazine; 8: tunicamycin ; 9: paraquat. The limited overlap between the CYP genes induced by paraquat and tunicamycin and those induced by the other chemicals led us to draw an arbitrary line to separate the genes that we designate as representing the response to stress from those representing the response to chemical challenge.

Fig. 4

Venn diagram showing the overlap between CYP genes inducible by chemicals and CYP genes associated to resistance in field or laboratory studies. The overlapping genes are identified, and of those, the three genes that have been causally linked to resistance in field-derived strains or experimental studies are in bold.