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Co-up-regulation of three P450 genes in response to permethrin exposure in permethrin resistant house flies, Musca domestica.

Zhu F, Li T, Zhang L, Liu N - BMC Physiol. (2008)

Bottom Line: Comparison of the deduced protein sequences of these three P450s from resistant ALHF and susceptible aabys and CS house flies revealed identical protein sequences.Genetic linkage analysis located CYP4D4v2 and CYP6A38 on autosome 5, corresponding to the linkage of P450-mediated resistance in ALHF, whereas CYP4G2 was located on autosome 3, where the major insecticide resistance factor(s) for ALHF had been mapped but no P450 genes reported prior to this study.Our study provides the first direct evidence that multiple P450 genes are co-up-regulated in permethrin resistant house flies through the induction mechanism, which increases overall expression levels of P450 genes in resistant house flies.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Entomology and Plant Pathology, Auburn University, Auburn, AL 36849, USA. fangzhudy@uky.edu

ABSTRACT

Background: Insects may use various biochemical pathways to enable them to tolerate the lethal action of insecticides. For example, increased cytochrome P450 detoxification is known to play an important role in many insect species. Both constitutively increased expression (overexpression) and induction of P450s are thought to be responsible for increased levels of detoxification of insecticides. However, unlike constitutively overexpressed P450 genes, whose expression association with insecticide resistance has been extensively studied, the induction of P450s is less well characterized in insecticide resistance. The current study focuses on the characterization of individual P450 genes that are induced in response to permethrin treatment in permethrin resistant house flies.

Results: The expression of 3 P450 genes, CYP4D4v2, CYP4G2, and CYP6A38, was co-up-regulated by permethrin treatment in permethrin resistant ALHF house flies in a time and dose-dependent manner. Comparison of the deduced protein sequences of these three P450s from resistant ALHF and susceptible aabys and CS house flies revealed identical protein sequences. Genetic linkage analysis located CYP4D4v2 and CYP6A38 on autosome 5, corresponding to the linkage of P450-mediated resistance in ALHF, whereas CYP4G2 was located on autosome 3, where the major insecticide resistance factor(s) for ALHF had been mapped but no P450 genes reported prior to this study.

Conclusion: Our study provides the first direct evidence that multiple P450 genes are co-up-regulated in permethrin resistant house flies through the induction mechanism, which increases overall expression levels of P450 genes in resistant house flies. Taken together with the significant induction of CYP4D4v2, CYP4G2, and CYP6A38 expression by permethrin only in permethrin resistant house flies and the correlation of the linkage of the genes with resistance and/or P450-mediated resistance in resistant ALHF house flies, this study sheds new light on the functional importance of P450 genes in response to insecticide treatment, detoxification of insecticides, the adaptation of insects to their environment, and the evolution of insecticide resistance.

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Northern blot analysis of differentially expressed patterns of CYP4G2, CYP4D4v2, and CYP6A38 between permethrin treated and untreated susceptible CS and resistant ALHF house flies. mRNAs were isolated from the whole bodies of 20 surviving house flies 24 h after permethrin treatment with 2 μg/fly. Blots were hybridized with the cDNA probes derived from three P450 gene fragments. The ethidium bromide stain of 18S ribosomal RNA in agarose gel is shown at the bottom.
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Figure 1: Northern blot analysis of differentially expressed patterns of CYP4G2, CYP4D4v2, and CYP6A38 between permethrin treated and untreated susceptible CS and resistant ALHF house flies. mRNAs were isolated from the whole bodies of 20 surviving house flies 24 h after permethrin treatment with 2 μg/fly. Blots were hybridized with the cDNA probes derived from three P450 gene fragments. The ethidium bromide stain of 18S ribosomal RNA in agarose gel is shown at the bottom.

Mentions: In order to identify the P450 genes raised in response to the insecticide treatment, we used the PCR technique with degenerated PCR primers [29] to amplify P450 cDNAs from house flies. We initially isolated a total of 19 P450 cDNA fragments from ALHF house flies using three primer pairs, C2/Flyh1, C2/Flyc1, and HemeR1/CYP6AD1 (Table 1). Northern blot analysis was conducted using the 19 P450 cDNA fragments as probes to compare expression levels of these cDNAs in both permethrin treated and untreated susceptible CS and resistant ALHF house flies. Three of the 19 P450 cDNAs were found to be significantly induced in the permethrin treated ALHF house flies after 24 hours treatment (Fig. 1) at the pilot dose of LD50 that caused ~50% mortality of each house fly strain.


Co-up-regulation of three P450 genes in response to permethrin exposure in permethrin resistant house flies, Musca domestica.

Zhu F, Li T, Zhang L, Liu N - BMC Physiol. (2008)

Northern blot analysis of differentially expressed patterns of CYP4G2, CYP4D4v2, and CYP6A38 between permethrin treated and untreated susceptible CS and resistant ALHF house flies. mRNAs were isolated from the whole bodies of 20 surviving house flies 24 h after permethrin treatment with 2 μg/fly. Blots were hybridized with the cDNA probes derived from three P450 gene fragments. The ethidium bromide stain of 18S ribosomal RNA in agarose gel is shown at the bottom.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2567968&req=5

Figure 1: Northern blot analysis of differentially expressed patterns of CYP4G2, CYP4D4v2, and CYP6A38 between permethrin treated and untreated susceptible CS and resistant ALHF house flies. mRNAs were isolated from the whole bodies of 20 surviving house flies 24 h after permethrin treatment with 2 μg/fly. Blots were hybridized with the cDNA probes derived from three P450 gene fragments. The ethidium bromide stain of 18S ribosomal RNA in agarose gel is shown at the bottom.
Mentions: In order to identify the P450 genes raised in response to the insecticide treatment, we used the PCR technique with degenerated PCR primers [29] to amplify P450 cDNAs from house flies. We initially isolated a total of 19 P450 cDNA fragments from ALHF house flies using three primer pairs, C2/Flyh1, C2/Flyc1, and HemeR1/CYP6AD1 (Table 1). Northern blot analysis was conducted using the 19 P450 cDNA fragments as probes to compare expression levels of these cDNAs in both permethrin treated and untreated susceptible CS and resistant ALHF house flies. Three of the 19 P450 cDNAs were found to be significantly induced in the permethrin treated ALHF house flies after 24 hours treatment (Fig. 1) at the pilot dose of LD50 that caused ~50% mortality of each house fly strain.

Bottom Line: Comparison of the deduced protein sequences of these three P450s from resistant ALHF and susceptible aabys and CS house flies revealed identical protein sequences.Genetic linkage analysis located CYP4D4v2 and CYP6A38 on autosome 5, corresponding to the linkage of P450-mediated resistance in ALHF, whereas CYP4G2 was located on autosome 3, where the major insecticide resistance factor(s) for ALHF had been mapped but no P450 genes reported prior to this study.Our study provides the first direct evidence that multiple P450 genes are co-up-regulated in permethrin resistant house flies through the induction mechanism, which increases overall expression levels of P450 genes in resistant house flies.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Entomology and Plant Pathology, Auburn University, Auburn, AL 36849, USA. fangzhudy@uky.edu

ABSTRACT

Background: Insects may use various biochemical pathways to enable them to tolerate the lethal action of insecticides. For example, increased cytochrome P450 detoxification is known to play an important role in many insect species. Both constitutively increased expression (overexpression) and induction of P450s are thought to be responsible for increased levels of detoxification of insecticides. However, unlike constitutively overexpressed P450 genes, whose expression association with insecticide resistance has been extensively studied, the induction of P450s is less well characterized in insecticide resistance. The current study focuses on the characterization of individual P450 genes that are induced in response to permethrin treatment in permethrin resistant house flies.

Results: The expression of 3 P450 genes, CYP4D4v2, CYP4G2, and CYP6A38, was co-up-regulated by permethrin treatment in permethrin resistant ALHF house flies in a time and dose-dependent manner. Comparison of the deduced protein sequences of these three P450s from resistant ALHF and susceptible aabys and CS house flies revealed identical protein sequences. Genetic linkage analysis located CYP4D4v2 and CYP6A38 on autosome 5, corresponding to the linkage of P450-mediated resistance in ALHF, whereas CYP4G2 was located on autosome 3, where the major insecticide resistance factor(s) for ALHF had been mapped but no P450 genes reported prior to this study.

Conclusion: Our study provides the first direct evidence that multiple P450 genes are co-up-regulated in permethrin resistant house flies through the induction mechanism, which increases overall expression levels of P450 genes in resistant house flies. Taken together with the significant induction of CYP4D4v2, CYP4G2, and CYP6A38 expression by permethrin only in permethrin resistant house flies and the correlation of the linkage of the genes with resistance and/or P450-mediated resistance in resistant ALHF house flies, this study sheds new light on the functional importance of P450 genes in response to insecticide treatment, detoxification of insecticides, the adaptation of insects to their environment, and the evolution of insecticide resistance.

Show MeSH