Cyp12a4 confers lufenuron resistance in a natural population of Drosophila melanogaster

Abstract

Lufenuron is an insect growth regulator insecticide mainly used for the control of the cat flea. To understand mechanisms of resistance to lufenuron, we have characterized lufenuron resistance in a natural population of Drosophila melanogaster. In this study we have used precise genetic mapping to identify a mechanism of lufenuron resistance: the overexpression of the cytochrome P450 gene Cyp12a4. Cyp12a4 is predicted to encode a mitochondrial cytochrome P450 enzyme. Expression of Cyp12a4 in D. melanogaster third-instar larvae was detected in the midgut and Malpighian tubules of both lufenuron-resistant and wild-type strains. The level of Cyp12a4 expression in the midgut is higher in the lufenuron-resistant strain than in wild-type strains. Driving the expression of Cyp12a4 in the midgut and Malpighian tubules by using the GAL4͞UAS gene expression system results in lufenuron resistance, but it does not result in resistance to three other insecticide classes. Transgenic expression of Cyp12a4 in a ubiquitous expression pattern results in late embryonic lethality, suggesting that high-level ectopic expression of Cyp12a4 is detrimental to development. cytochrome P450 ͉ insecticide resistance ͉ insect growth regulator ͉ genetic mopping I nsecticide resistance is an important example of natural selection (1). Resistance is predominantly mediated either by changes in the metabolism of the insecticide or by changes in the sensitivity of insecticide targets (2). Insecticide resistance mediated by target site modification is well documented for most commonly used insecticides (3-7). Molecular mechanisms of resistance due to increased insecticide metabolism by cytochrome P450s, esterases, and glutathione S-transferases are less well understood. Resistance is commonly found to be the result of up-regulation of single or multiple members of these enzyme families (8-13); however, resistance due to structural changes in enzymes has also been documented (14-16).Lufenuron is an insect growth regulator insecticide, mainly used in the control of the cat flea, Ctenocephalides felis. It is active against larval developmental stages, causing cuticular lesions resulting from the disruption of chitin synthesis (17). Lufenuron is also active against Diptera, including Drosophila melanogaster (18). To understand possible resistance mechanisms to lufenuron, we have used D. melanogaster as a model. Lufenuron resistance in D. melanogaster was originally reported in populations from two widely separated locations in the United States (19). As it was not expected that D. melanogaster had been in contact with lufenuron, it was postulated that this resistance resulted from cross-resistance that had evolved to an earlier, widely used insecticide (19). Subsequently, it was found that resistance to lufenuron was caused by the up-regulation of the cytochrome P450 gene Cyp6g1, which confers resistance to numerous insecticide classes, and that Cyp6g1-mediated resistance is widespread (8,20).Here we describe the discovery of a mechanism of lufenuron r...

Xenobiotic response in Drosophila melanogaster: Sex dependence of P450 and GST gene induction

“…Several of the genes shown to be transcriptionally regulated by phenobarbital or atrazine have been reported before to be associated with metabolic resistance to insecticides, particularly Cyp6a2 (Waters et al, 1992;Brun et al, 1996;Amichot et al, 2004) and Cyp6g1 (Daborn et al, 2002) but also Cyp6a8, Cyp12d1, Cyp4e2 and Cyp12a4 (Amichot et al, 1994;Maitra et al, 2000;Le Goff et al, 2003;Pedra et al, 2004;Bogwitz et al, 2005). Some of these genes are also inducible by other chemicals, such as DDT (Brandt et al, 2002).…”

Resistance to lambda‐cyhalothrin in Spanish field populations of Ceratitis capitata and metabolic resistance mediated by P450 in a resistant strain

“…P450 genes linked to pyrethroid resistance mostly belong to the CYP4, CYP6 and CYP9 families (Yang et al, 2006;Komagata et al, 2010;Brun-Barale et al, 2010), though other CYP genes such as CYP325A3 in Anopheles gambiae may also be involved in resistance (David et al, 2005). Besides, the up-regulation of genes of the CYP12 family has been shown to confer resistance to DDT (Brandt et al, 2002) and lufenuron (Bogwitz et al, 2005). We have then analyzed by qPCR fifty three P450 genes belonging to the CYP4, CYP6, CYP9 and CYP12 families in C. capitata.…”

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

“…Cyp4e2 (Amichot et al, 1994), Cyp6a2 (Maitra et al, 1996(Maitra et al, , 2000Pedra et al, 2004), Cyp6a8 (Maitra et al, 2000), Cyp6a9 (Maitra et al, 1996), Cyp6g1 (Catania et al, 2004;Daborn et al, 2002;Schlenke and Begun, 2004), Cyp6w1 (Pedra et al, 2004), Cyp12a4 (Bogwitz et al, 2005), Cyp12d1 (Brandt et al, 2002) and Cyp308a1 (Gene Ontology Consortium, 2001). Six of these genes have been associated with high levels of gene transcription in resistant insects (Amichot et al, 2004;Bogwitz et al, 2005;Daborn et al, 2002Daborn et al, , 2007Helvig et al, 2004;Le Goff et al, 2003).…”

“…Six of these genes have been associated with high levels of gene transcription in resistant insects (Amichot et al, 2004;Bogwitz et al, 2005;Daborn et al, 2002Daborn et al, , 2007Helvig et al, 2004;Le Goff et al, 2003).…”

“…In addition, Marsano et al (2005) and Abbreviations: CDS, coding sequence; Cyp, cytochrome P450 monooxygenases; DINE-1, Drosophila interspersed element 1; LTR, long terminal repeat; nt, nucleotide; MITE, miniature inverted-repeat transposable element; SINE, short interspersed nuclear element; TEs, transposable elements; TFBS, transcription factor binding site; TIR, terminal inverted repeat. Bogwitz et al (2005) have suggested that the presence of the Bari-1 transposon at the 3′ end of Cyp12a4 in D. melanogaster increases gene expression. These reports associate TE insertion with gene overexpression and insecticide resistance.…”