Department of Biology, Emory University, Atlanta, GA, USA.
Insect Mol Biol. 2010 Feb;19(1):121-30. doi: 10.1111/j.1365-2583.2009.00954.x. Epub 2009 Dec 1.
Many Drosophila cytochrome P450 or Cyp genes are induced by caffeine and phenobarbital (PB). To understand the induction mechanism, we created Drosophila S2 cell lines stably transformed with different luciferase reporter plasmids carrying upstream DNAs of Cyp6a8 allele of the resistant 91-R strain, and the 1.1-kb upstream DNAs of Cyp6g1 of the 91-R and the susceptible 91-C strains. Following 24 h treatment with dichlorodiphenyltrichloroethane (DDT), caffeine or PB, luciferase activity of all cell lines was determined. Results showed that the 0.1-kb DNA of Cyp6a8 and the upstream DNAs of Cyp6g1 from both strains are not induced by these chemicals in S2 cells. However, the 0.2-, 0.5- and 0.8-kb DNAs of Cyp6a8 showed 13-24-, 4-5- and 2.2-2.7-fold induction with caffeine, PB and DDT, respectively. These DNAs also showed a 2-3-fold synergistic effect of caffeine and PB but not of caffeine and DDT. The results suggest that the cis-regulatory elements for all three chemicals are located within the -11/-199 DNA of Cyp6a8. Furthermore, caffeine and PB inductions appear to be mediated via different cis-elements, whereas caffeine and DDT induction may involve common regulatory elements. These stably transformed cell lines should help understand the mechanism of resistance-associated Cyp gene overexpression in Drosophila.
许多果蝇细胞色素 P450 或 Cyp 基因受咖啡因和苯巴比妥（PB）诱导。为了了解诱导机制，我们创建了稳定转化的果蝇 S2 细胞系，这些细胞系携带具有抗性 91-R 菌株 Cyp6a8 等位基因的上游 DNA 的不同荧光素酶报告质粒，以及 91-R 和易感 91-C 菌株的 Cyp6g1 的 1.1kb 上游 DNA。用滴滴涕（DDT）、咖啡因或 PB 处理 24 小时后，测定所有细胞系的荧光素酶活性。结果表明，Cyp6a8 的 0.1kb DNA 和来自两种菌株的 Cyp6g1 的上游 DNA 在 S2 细胞中不受这些化学物质的诱导。然而，Cyp6a8 的 0.2、0.5 和 0.8kb DNA 分别被咖啡因、PB 和 DDT 诱导 13-24、4-5 和 2.2-2.7 倍。这些 DNA 还显示出咖啡因和 PB 的 2-3 倍协同作用，但咖啡因和 DDT 没有。结果表明，所有三种化学物质的顺式调节元件都位于 Cyp6a8 的-11/-199 DNA 内。此外，咖啡因和 PB 诱导似乎通过不同的顺式元件介导，而咖啡因和 DDT 诱导可能涉及共同的调节元件。这些稳定转化的细胞系应该有助于理解果蝇中与抗性相关的 Cyp 基因过度表达的机制。
