Cis-Regulatory Elements in the Accord Retrotransposon Result in Tissue-Specific Expression of the Drosophila melanogaster Insecticide Resistance Gene Cyp6g1

Abstract

Transposable elements are a major mutation source and powerful agents of adaptive change. Some transposable element insertions in genomes increase to a high frequency because of the selective advantage the mutant phenotype provides. Cyp6g1-mediated insecticide resistance in Drosophila melanogaster is due to the upregulation of the cytochrome P450 gene Cyp6g1, leading to the resistance to a variety of insecticide classes. The upregulation of Cyp6g1 is correlated with the presence of the long terminal repeat (LTR) of an Accord retrotransposon inserted 291bp upstream of the Cyp6g1 transcription start site. This resistant allele (DDT-R) is currently at a high frequency in D. melanogaster populations around the world. Here, we characterize the spatial expression of Cyp6g1 in insecticide-resistant and -susceptible strains. We show that the Accord LTR insertion is indeed the resistance-associated mutation and demonstrate that the Accord LTR carries regulatory sequences that increase the expression of Cyp6g1 in tissues important for detoxification, the midgut, Malpighian tubules, and the fat body. This study provides a significant example of how changes in tissue-specific gene expression caused by transposableelement insertions can contribute to adaptation.

Characterization of Drosophila melanogaster cytochrome P450 genes

“…These include the P450s involved in the synthesis of ecdysone, Cyp302a1, Cyp306a1, Cyp307a2, Cyp314a1, and Cyp315a1. Cyp6g1 is detected in the midgut and Malpighian tubules in both feeding and wandering third instar larvae, with additional expression detected in the fat body of wandering larvae (49). Expression for a subset of these P450s was confirmed by semiquantitative PCR (SI Methods; Fig.…”

“…S1). Nine P450s are detected in the fat body, including the previously characterized Cyp6g1, Cyp12d1, and Cyp314a1 (22,40,49). Cyp4p2 and Cyp4s3 are the only P450s detected exclusively in the fat body.…”

“…The P450s detected only in embryos are mostly expressed in structures such as the yolk cells, or are maternal transcripts, whereas the P450s detected in larvae only are mostly expressed in tissues involved in detoxification, such as the midgut, Malpighian tubules, and the fat body. Cyp6g1 and Cyp12a4, which confer insecticide resistance when over-expressed (36,38,49), are expressed in the midgut and Malpighian tubules in feeding third instar larvae, but show no detectable expression in embryos. RNAi of these 2 genes showed no mortality under laboratory conditions.…”

Genomic regions harboring insecticide resistance-associated Cyp genes are enriched by transposable element fragments carrying putative transcription factor binding sites in two sibling Drosophila species

“…Our data show that these insertions in Cyp genes carry putative TFBSs, which reinforces the hypothesis that they might play important roles in the adaptation of Drosophila species. These particular characteristics may explain why TEs interfere in the transcription of some Cyp genes by elevating their expression levels (Catania et al, 2004;Chung et al, 2007;Daborn et al, 2002). Empirical studies involving transfection and expression experiments for these putative regulatory sequences can corroborate the hypothesis that DNAREP1 sequences have been domesticated to contribute to new, alternative transcription start sites in Drosophila genomes.…”

“…Daborn et al (2002) reported that the insecticide resistance mapping to the DDT-R locus of D. melanogaster is due to the over-expression of one gene, Cyp6g1, and that a single Cyp6g1 allele linked to this resistance is distributed worldwide. Unlike other insecticide resistance genes in which overexpression is associated with point mutations, over-expression of Cyp6g1is characterized by the insertion of a fragment of the Accord retrotransposon upstream of the gene (Catania et al, 2004;Chung et al, 2007;Daborn et al, 2002). Schlenke and Begun (2004) have also reported an association between a TE insertion and insecticide resistance in Drosophila simulans.…”