致倦库蚊GSTe1和CYP9J40的时空表达特点
Temporal and spatial expression of the GSTe1 and CYP9J40 genes in Culex pipiens quinquefasciatus
王 炜1** 陈少良1 崔 峰2***
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DOI：10.7679/j.issn.2095-1353.2014.182
作者单位：1. 北京林业大学生物科学与技术学院，北京 100083；2. 中国科学院动物研究所农业虫害鼠害综合治理研究国家重点实验室，北京 100101
中文关键词：致倦库蚊，解毒酶，P450，GST，抗药性，时空表达
英文关键词： Culex pipiens quinquefasciatus, detoxification enzyme, P450, GST, insecticide resistance, temporal and spatial expression
中文摘要：
【目的】细胞色素P450单加氧酶（P450）和谷胱甘肽-S-转移酶（GST）是生物降解外源有毒物质的重要解毒酶家族，在昆虫适应杀虫剂选择的过程中发挥重要的作用。以往在致倦库蚊Culex pipiens quinquefasciatus对硫磷抗性研究中，发现了一个Epsilon GST基因（GSTe1）和一个P450 CYP9家族基因（CYP9J40）在抗性品系中高表达。研究它们在蚊虫中的时空表达模式对理解它们的生物学功能有重要意义。【方法】 本研究利用实时定量PCR和原位杂交的方法探讨了这两个解毒酶基因在蚊虫中的时空表达。【结果】 定量PCR显示GSTe1和CYP9J40在对硫磷抗性蚊虫中的转录水平分别是敏感蚊虫的4.2倍和1.9倍，它们在蚊虫整个生活周期的表达模式相似，均为幼虫期表达量相对较低，从蛹期开始表达量显著升高，到成虫期雌虫达到顶峰，而雄虫与幼虫期表达量相当。原位杂交结果表明这两个基因在肌肉、消化道和卵巢中都有表达，且CYP9J40的表达更广泛，在脑和胸部神经节也有明显表达。【结论】 基因表达的发育阶段特异性和广泛的组织分布说明GSTe1和CYP9J40除了参与蚊虫对杀虫剂的抗性，还可能具有其它生理功能，如对雌虫发育有重要的影响。
英文摘要：
[Objectives] Cytochrome P450 monooxygenase (P450) and glutathione S-transferase (GST) are important enzymes for the degradation of exogenous toxins in eukaryotes and play key roles in insecticide resistance in insects. In a previous study we found that the transcriptional levels of the Epsilon GST (GSTe1) and P450 CYP9 (CYP9J40) genes were upregulated in a parathion-resistant strain of Culex pipiens quinquefasciatus. We here explore the temporal and spatial expression pa-tterns of these two genes in order to further elaborate their biological functions. [Methods] Real-time PCR and in situ hybridization were used to explore the temporal and spatial expression patterns of the two genes. [Results] Real-time PCR showed that the transcription levels of GSTe1 and CYP9J40 in the parathion-resistant strain of C. p. quinquefasciatus were 4.2 and 1.9-fold higher, respectively, than those of a susceptible strain. These two genes had similar expression patterns over the mosquito life cycle; expression was low in larvae, increased in pupae, and peaked in female adults, whereas expre-ssion in male adults was the same as that in larvae. In situ hybridization demonstrated that both genes were expressed in muscle, the digestive tract, and ovary. CYP9J40 was, however, expressed in a greater range of tissues, including the brain and thoracic ganglion, than GSTe1. [Conclusion] The observed changes in expression levels during development and the expression of these genes in a wide range of tissues imply that, in addition to insecticide resistance, GSTe1 and CYP9J40 play a role in other metabolic processes, such as female development.