Table_6_Juvenile hormone III induction reveals key genes in general metabolism, pheromone biosynthesis, and detoxification in Eurasian spruce bark beetle.DOCX

IntroductionIn recent years, bark beetle Ips typographus, has caused extensive damage to European Norway spruce forests through widespread outbreaks. This pest employs pheromone-assisted aggregation to overcome tree defense, resulting in mass attacks on host spruce. Many morphological and behavioral processes in I. typographus are under the regulation of juvenile hormone III (JH III), including the biosynthesis of aggregation pheromones and associated detoxification monoterpene conjugates.
Objectives and MethodsIn this study, we topically applied juvenile hormone III (JH III) and performed metabolomics, transcriptomics, and proteomics in I. typographus both sexes, with focused aims; 1. Highlight the JH III-regulated metabolic processes; 2. Identify pheromone biosynthesis-linked genes; and 3. Investigate JH III’s impact on detoxification conjugates linked to pheromonal components.
ResultsNumerous gene families were enriched after JH III treatment, including genes associated with catalytic and oxidoreductase activity, esterases, phosphatases, and membrane transporters. Sex-specific enrichments for reproduction-related and detoxification genes in females and metabolic regulation genes in males were observed. On the protein level were enriched metal ion binding and transferase enzymes in male beetles. After JHIII treatment, mevalonate pathway genes, including terminal isoprenyl diphosphate synthase (IPDS), were exclusively 35- folds upregulated in males, providing evidence of de novo biosynthesis of pheromone components 2-methyl-3-buten-2-ol and ipsdienol. In addition, cytochrome P450 genes likely involved in the biosynthesis of cis/trans-verbenol, detoxification, and formation of ipsdienol, were 3-fold upregulated in the male gut. The increase in gene expression correlated with the heightened production of the respective metabolites. Detoxification conjugates, verbenyl oleate in the beetle fat body and verbenyl diglycosides in the gut, were induced by JHIII application, which confirms the hormone regulation of their formation. The JH III induction also increased the gene contigs esterase and glycosyl hydrolase up to proteins from male gut tissue. The esterase was proposed to release pheromone cis-verbenol in adult males by breaking down verbenyl oleate. The correlating analyses confirmed a reduction in the abundance of verbenyl oleate in the induced male beetle.
ConclusionThe data provide evidence of JH III’s regulatory role in the expression of genes and enzymes related to fundamental beetle metabolism, pheromone biosynthesis, and detoxification in Ips typographus.

引言
近年来，欧洲云杉八齿小蠹（Ips typographus）通过大规模暴发，对欧洲挪威云杉林造成了严重破坏。该害虫借助信息素介导的聚集行为突破寄主树木的防御系统，进而对宿主云杉造成大规模侵染为害。欧洲云杉八齿小蠹的诸多形态与行为过程均受保幼激素III（juvenile hormone III, JH III）调控，其中包括聚集信息素的生物合成以及相关解毒单萜结合物的代谢过程。

研究目标与方法
本研究通过体表涂抹保幼激素III（JH III），对雌雄两性欧洲云杉八齿小蠹开展代谢组学、转录组学与蛋白质组学分析，核心研究目标如下：1. 阐明保幼激素III调控的代谢通路；2. 筛选与信息素生物合成相关的功能基因；3. 探究保幼激素III对信息素组分相关解毒结合物的调控作用。

研究结果
保幼激素III处理后，大量基因家族出现富集现象，其中包括与催化活性、氧化还原酶活性、酯酶、磷酸酶以及膜转运蛋白相关的基因家族。研究观察到性特异性富集特征：雌性个体中与繁殖及解毒相关的基因呈现富集，雄性个体则以代谢调控基因富集为主。在蛋白质组层面，雄性小蠹体内的金属离子结合蛋白与转移酶类呈现富集特征。
保幼激素III处理后，甲羟戊酸通路相关基因（包括末端异戊烯二磷酸合酶（IPDS））仅在雄性个体中出现35倍的上调表达，这为信息素组分2-甲基-3-丁烯-2-醇与艾氏醇（ipsdienol）的从头生物合成提供了实验证据。此外，雄性肠道中参与顺/反式葑醇（cis/trans-verbenol）生物合成、解毒过程以及艾氏醇形成的细胞色素P450基因出现3倍上调表达。基因表达的上调与相应代谢产物的产量提升呈显著正相关。
保幼激素III处理可诱导解毒结合物的产生：小蠹脂肪体内的葑醇油酸酯与肠道内的葑基二糖苷均出现丰度提升，这证实了保幼激素III对这些结合物形成的调控作用。保幼激素III诱导还提升了雄性肠道组织中酯酶与糖苷水解酶的基因表达及蛋白丰度。研究推测，该酯酶可通过分解葑醇油酸酯，在雄性成虫体内释放信息素顺式葑醇。相关性分析证实，经保幼激素III诱导的雄性小蠹体内葑醇油酸酯的丰度显著降低。

结论
本研究数据证实，保幼激素III对欧洲云杉八齿小蠹的基础代谢、信息素生物合成以及解毒过程相关的基因与酶的表达具有明确的调控作用。