CPR和P450基因在昆虫抗药性中的作用研究进展

P450酶系在昆虫代谢农药中有重要作用，NADPH-细胞色素P450还原酶（NADPH-cytochrome P450 reductase，CPR）和细胞色素P450（P450）在该酶系起核心作用。昆虫具有P450超基因家族，但只有一个单一的CPR基因，CPR是昆虫所有参与农药代谢的P450酶的唯一电子供体，其影响P450活性。P450基因的高水平表达在害虫抗药性中具有重要作用，P450基因介导的昆虫抗药性是最重要的代谢抗性类型。不同P450基因的高表达的调控机制不同，引起P450基因过量表达的原因可能有P450基因的编码区突变、顺式作用元件和反式作用因子变化、基因扩增等。细胞色素P450介导的抗药性存在一定程度的进化可塑性，即同种昆虫不同种群对相同的农药产生抗药性时，导致抗性产生的P450基因不同；同一昆虫品系在某种农药的抗性选择压力下，影响抗性的P450基因的种类和表达特性会随着持续的农药选择而发生变化。最近的研究显示，CPR的变异和昆虫抗药性相关，但是昆虫CPR基因介导抗药性的机制还缺乏深入研究。全面阐释P450酶系介导昆虫抗药性的机制、建立基于P450基因表达量变化与CPR突变的抗性分子标记，对于害虫抗药性治理具有重要意义。     

昆虫抗药性机理：行为和生理改变及解毒代谢增强（英文）

杀虫剂抗性是指“生物的一个品系发展了对该生物正常种群中大多数个体具有致死作用剂量的杀虫药剂的能力”。行为改变、生理学上的变化或代谢解毒等抗性机制能够降低毒物到达靶标的有效剂量。行为抗性是指减少昆虫与毒物接触或使昆虫能够存活于对大多数对正常个体致死（或有害）的环境中的任何行为。生理学改变的机制包括杀虫剂对表皮的穿透性降低、增加对药剂阻隔（sequestration）或储存和加速杀虫剂的排泄。细胞色素P450、水解酶和谷胱甘肽S-转移酶是杀虫药剂代谢解毒的主要3大酶系。细胞色素P450是一个超基因家族,是生物体内对外源性和内源性化合物解毒代谢或活化最重要的酶系。在许多害虫中发现P450介导的解毒代谢增加导致了对杀虫药剂抗性的增加。谷胱甘肽S-转移酶是可溶性的 二聚体蛋白,与代谢解毒、大量内源性和外源性化合物的排泄有关,许多昆虫中证明其抗药性与该酶活性增加有关。水解酶实际上是一组异源的酶类,其对抗药性的作用包括通过基因扩增增加酶量,作为结合蛋白隔离杀虫药剂或通过增加酶的活性加强对药剂的水解作用。     

P450s in plant-insect interactions

Cytochrome P450 monooxygenases (P450s) are integral in defining the relationships between plants and insects. Secondary metabolites produced in plants for protection against insects and other organisms are synthesized via pathways that include P450s in many different families and subfamilies. Survival of insects in the presence of toxic secondary metabolites depends on their metabolism by more limited groups of P450s. Examples of functionally characterized plant and insect P450s known to be involved in these interactions are discussed in terms of their diversities, reactivities and regulators. These and future examples, which will be uncovered as the fields of plant biology and entomology converge on this interesting area, provide much insight into the array of plant metabolites that are mainline defenses against insects, the range of insect monooxygenases that inactivate these compounds and the evolutionary processes occurring as these organisms wage daily battles with one another. Molecular perspectives on these interactions will provide the scientific community with information critical for genetic manipulation of these organisms aimed at enhancing plant resistance to insects and eliminating insect resistance to natural plant toxins and synthetic insecticides.     

昆虫对植物次生物质的代谢适应机制及其对昆虫抗药性的意义

植物次生物质(plant secondary metabolites)对昆虫的取食行为、生长发育及繁殖可以产生不利影响,甚至对昆虫可以产生毒杀作用。为了应对植物次生物质的不利影响,昆虫通过对植物次生物质忌避取食、解毒代谢等多种机制,而对寄主植物产生适应性。其中,昆虫的解毒代谢酶包括昆虫细胞色素P450酶系（P450s）及谷胱甘肽硫转移酶（GSTs）等,在昆虫对植物次生物质的解毒代谢及对寄主植物的适应性中发挥了重要作用。昆虫的解毒酶系统不仅可以代谢植物次生物质,还可能代谢化学杀虫剂,因而昆虫对寄主植物的适应性与其对杀虫剂的耐药性甚至抗药性密切相关。昆虫细胞色素P450s和GSTs等代谢解毒酶活性及相关基因的表达可以被植物次生物质影响,这不仅使昆虫对寄主植物的防御产生了适应性,还影响了昆虫对杀虫剂的解毒代谢,因而改变昆虫的耐药性或抗药性。掌握昆虫对植物次生物质的代谢适应机制及其在昆虫抗药性中的作用,对于明确昆虫的抗药性机制具有重要的参考意义。本文综述了植物次生物质对昆虫的影响、昆虫对寄主植物次生物质的代谢机制、昆虫对植物次生物质的代谢适应性对昆虫耐药性及抗药性的影响等方面的研究进展。     

Cytochrome P450 MA expression in insecticide-resistant German cockroaches (Dictyoptera: Blattellidae).

Cytochrome P450 monooxygenases are detoxification enzymes commonly involved in insecticide resistance by insects. Recently, an overexpressed form of this enzyme, P450 MA, was purified from an insecticide-resistant strain of German cockroach, Blattella germanica (L.), and polyclonal antisera (anti-P450 MA) was produced. To test hypotheses that the overexpressed condition of P450 MA has evolved in > 1 geographic location and that P450 MA might be involved in insecticide resistance to specific insecticides, investigations were conducted using 4 insecticide-resistant and 1 susceptible German cockroach strains. In western blots that used anti-P450 MA antiserum as a probe, substantial differences in expression of P450 MA were observed. Strains showing the highest P450 MA expression had both the highest tolerance to the organophosphate insecticide chlorpyrifos and cytochrome P450-mediated demethylation activity. Results support the hypothesis that cytochrome P450 MA is potentially overexpressed in insecticide-resistant populations on a global scale.     

Cytochrome P450 monooxygenases and insecticide resistance in insects

Cytochrome P450 monooxygenases are involved in many cases of resistance of insects to insecticides. Resistance has long been associated with an increase in monooxygenase activities and with an increase in cytochrome P450 content. However, this increase does not always account for all of the resistance. In Drosophila melanogaster, we have shown that the overproduction of cytochrome P450 can be lost by the fly without a corresponding complete loss of resistance. These results prompted the sequencing of a cytochrome P450 candidate for resistance in resistant and susceptible flies. Several mutations leading to amino-acid substitutions have been detected in the P450 gene CYP6A2 of a resistant strain. The location of these mutations in a model of the 3D structure of the CYP6A2 protein suggested that some of them may be important for enzyme activity of this molecule. This has been verified by heterologous expression of wild-type and mutated cDNA in Escherichia coli. When other resistance mechanisms are considered, relatively few genetic mutations are involved in insecticide resistance, and this has led to an optimistic view of the management of resistance. Our observations compel us to survey in more detail the genetic diversity of cytochrome P450 genes and alleles involved in resistance.     

细胞色素P450介导抗性的进化可塑性

细胞色素P450是超基因家族,由其介导的杀虫剂代谢解毒的增强是昆虫产生抗药性的普遍而主要的机制。近年的研究表明,细胞色素P450介导的代谢抗性表现出一定程度的进化可塑性:即使是同种昆虫的不同种群在相同种类杀虫剂的胁迫下,进化选择出的抗性相关的细胞色素P450也有所不同,抗性的产生也可以是几种不同细胞色素P450协同作用或控制P450表达的调控因子的不同。