Liu Xiang-Yu, Wang Sha-Sha, Zhong Fan, Zhou Min, Jiang Xin-Yi, Cheng Yi-Sha, Dan Yi-Hao, Hu Gao, Li Can, Tang Bin, Wu Yan
Guizhou Provincial Key Laboratory for Rare Animal and Economic Insect of the Mountainous Region, Department of Biology and Engineering of Environment, Guiyang University, Guiyang, China.
College of Life and Environmental Sciences, Hangzhou Normal University, Hangzhou, Zhejiang, China.
Front Physiol. 2022 Oct 3;13:1034926. doi: 10.3389/fphys.2022.1034926. eCollection 2022.
Chitin is the main component of insect exoskeleton and midgut peritrophic membrane. Insect molting is the result of the balance and coordination of chitin synthesis and degradation in chitin metabolism under the action of hormones. In this study, a 678 bp dsRNA fragment was designed and synthesized according to the known CHI (Chitinase) sequence of . It was injected into the larvae to observe the molting and development of . At the same time, the activities of trehalase and chitinase, the contents of trehalose, chitin and other substances were detected, and the expression of related genes in the chitin synthesis pathway was determined. The results showed that gene was highly expressed at the end of each instar, prepupa and pupal stage before molting; At 12 and 24 h after dsRNA injection of gene of , the expression of gene decreased significantly, and the chitinase activity decreased significantly from 12 to 48 h. The expression of chitin synthase () gene decreased significantly, and the chitin content increased significantly. Some larvae could not molt normally and complete development, leading to certain mortality. Secondly, after RNAi of gene, the content of glucose and glycogen increased first and then decreased, while the content of trehalose decreased significantly or showed a downward trend. The activities of the two types of trehalase and the expression levels of trehalase genes decreased first and then increased, especially the trehalase activities increased significantly at 48 h after dsCHI injection. And trehalose-6-phosphate synthase (), glutamine: fructose-6-phosphate amidotransferase (), UDP-N-acetylglucosamine pyrophosphorylases (), hexokinase (), glucose-6-phosphate isomerase () and phosphoacetylglucosamine mutase () all decreased significantly at 24 h, and then increased or significantly increased at 48 h. These results indicated that when the expression of chitinase gene of was inhibited, it affected the degradation of chitin in the old epidermis and the formation of new epidermis, and the content of chitin increased, which led to the failure of larvae to molt normally. Moreover, the chitin synthesis pathway and trehalose metabolism were also regulated. The relevant results provide a theoretical basis for screening target genes and developing green insecticides to control pests by using the chitin metabolism pathway.
几丁质是昆虫外骨骼和中肠围食膜的主要成分。昆虫蜕皮是在激素作用下几丁质代谢中几丁质合成与降解平衡及协调的结果。在本研究中,根据已知的[昆虫名称]几丁质酶(CHI)序列设计并合成了一段678 bp的dsRNA片段。将其注射到幼虫体内以观察[昆虫名称]的蜕皮和发育情况。同时,检测海藻糖酶和几丁质酶的活性、海藻糖、几丁质等物质的含量,并测定几丁质合成途径中相关基因的表达。结果表明,[昆虫名称]几丁质酶基因在各龄期末、化蛹前和蛹期蜕皮前高表达;在注射[昆虫名称]几丁质酶基因dsRNA后的12 h和24 h时几丁质酶基因表达显著下降,几丁质酶活性在12至48 h显著降低。几丁质合酶([具体名称])基因表达显著下降,几丁质含量显著增加。部分幼虫无法正常蜕皮并完成发育,导致一定死亡率。其次,[昆虫名称]几丁质酶基因RNA干扰后,葡萄糖和糖原含量先升高后降低,而海藻糖含量显著下降或呈下降趋势。两种海藻糖酶活性及海藻糖酶基因表达水平先下降后升高,尤其是在注射dsCHI后的48 h海藻糖酶活性显著升高。并且海藻糖-6-磷酸合酶([具体名称])、谷氨酰胺:果糖-6-磷酸酰胺转移酶([具体名称])、UDP-N-乙酰葡糖胺焦磷酸化酶([具体名称])、己糖激酶([具体名称])、葡萄糖-6-磷酸异构酶([具体名称])和磷酸乙酰葡糖胺变位酶([具体名称])在24 h时均显著下降,然后在48 h时升高或显著升高。这些结果表明,当[昆虫名称]几丁质酶基因表达受到抑制时,影响了旧表皮中几丁质的降解和新表皮的形成,几丁质含量增加,导致幼虫无法正常蜕皮。此外,几丁质合成途径和海藻糖代谢也受到调控。相关结果为利用几丁质代谢途径筛选靶标基因和开发绿色杀虫剂防治害虫提供了理论依据。