Table_4_Integrative Analysis of Elicitor-Induced Camptothecin Biosynthesis in Camptotheca acuminata Plantlets Through a Combined Omics Approach.xls

Treatments with abiotic elicitors can efficiently induce the accumulation of specialized metabolites in plants. We used a combined omics approach to analyze the elicitation effects of MeJa, AgNO3, and PEG on camptothecin (CPT) biosynthesis in Camptotheca acuminata plantlets. Untargeted analyses revealed that treatments with MeJa, AgNO3, and PEG significantly inhibited the photosynthetic pathway and promoted carbon metabolism and secondary metabolic pathways. The CPT levels increased by 78.6, 73.3, and 50.0% in the MeJa, AgNO3, and PEG treatment groups, respectively. Using C. acuminata plantlets after elicitation treatment, we mined and characterized 15 new alkaloids, 25 known CPT analogs and precursors, 9 iridoid biosynthetic precursors, and 15 tryptamine biosynthetic precursors based on their MS/MS fragmentation spectra. Using 32 characterized genes involved in CPT biosynthesis as bait, we mined 12 prioritized CYP450 genes from the 416 CYP450 candidates that had been identified based on co-expression analysis, conserved domain analysis, and their elicitation-associated upregulation patterns. This study provides a comprehensive perspective on CPT biosynthesis in C. acuminata plantlets after abiotic elicitation. The findings enable us to elucidate the previously unexplored CYP450-mediated oxidation steps for CPT biosynthesis.

非生物激发子（abiotic elicitor）处理可高效诱导植物次生代谢产物的积累。本研究采用多组学联用策略，分析了茉莉酸甲酯（methyl jasmonate, MeJa）、硝酸银（AgNO3）与聚乙二醇（PEG）对喜树（Camptotheca acuminata）幼苗中喜树碱（camptothecin, CPT）生物合成的激发效应。非靶向代谢组学分析显示，MeJa、AgNO3及PEG处理均显著抑制光合通路，同时促进碳代谢与次生代谢通路的激活。与对照组相比，MeJa、AgNO3与PEG处理组的喜树碱含量分别提升78.6%、73.3%及50.0%。本研究以经激发子处理的喜树幼苗为材料，基于串联质谱（MS/MS）碎裂光谱，共鉴定并表征了15种新型生物碱、25种已知喜树碱类似物及前体、9种环烯醚萜生物合成前体与15种色胺生物合成前体。本研究以32个已功能注释的喜树碱生物合成相关基因为诱饵，通过共表达分析、保守结构域分析以及激发子处理下的上调表达模式筛选，从416个候选细胞色素P450（cytochrome P450, CYP450）基因中挖掘出12个优先候选基因。本研究为非生物激发子处理后的喜树幼苗喜树碱生物合成机制提供了全面的解析视角，阐明了此前未被揭示的、由CYP450介导的喜树碱生物合成氧化步骤。