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Abstract：
Entada phaseoloides stem is known for its high medicinal benefits and ornamental value. Flavonoids are one of the main active constituents in E. phaseoloides stem. However, the regulatory mechanism of flavonoids accumulation in E. phaseoloides is lacking. Here, phytochemical compounds and transcripts from stems at different developmental stages in E. phaseoloides were investigated by metabolome and transcriptome analysis. The metabolite profiling of the oldest stem was obviously different from young and older stem tissues. A total of 198 flavonoids were detected, and flavones, flavonols, anthocyanins, isoflavones, and flavanones were the main subclasses. The metabolome data showed that the content of acacetin was significantly higher in the young stem and older stem than the oldest stem. Rutin and myricitrin showed significantly higher levels in the oldest stem. A total of 143 MYBs and 143 bHLHs were identified and classified in the RNA-seq data. Meanwhile, 34 flavonoid biosynthesis structural genes were identified. Based on the expression pattern of structural genes involved in flavonoid biosynthesis, it indicated that flavonol, anthocyanin, and proanthocyanin biosynthesis were first active during the development of E. phaseoloides stem, and the anthocyanin or proanthocyanin biosynthesis branch was dominant; the flavone biosynthesis branch was active at the late developmental stage of the stem. Through the correlation analysis of transcriptome and metabolome data, the potential candidate genes related to regulating flavonoid synthesis and transport were identified. Among them, the MYBs, bHLH, and TTG1 are coregulated biosynthesis of flavonols and structural genes, bHLH and transporter genes are coregulated biosynthesis of anthocyanins. In addition, the WDR gene TTG1-like (AN11) may regulate dihydrochalcones and flavonol biosynthesis in specific combinations with IIIb bHLH and R2R3-MYB proteins. Furthermore, the transport gene protein TRANSPARENT TESTA 12-like gene is positively regulated the accumulation of rutin, and the homolog of ABC transporter B family member gene is positively correlated with the content of flavone acacetin. This study offered candidate genes involved in flavonoid biosynthesis, information of flavonoid composition and characteristics of flavonoids accumulation, improved our understanding of the MYBs and bHLHs-related regulation networks of flavonoid biosynthesis in E. phaseoloides stem, and provided references for the metabolic engineering of flavonoid biosynthesis in E. phaseoloides stem.
摘要：
Entadaphaseoloides茎以其高药用价值和观赏价值而闻名。黄酮类化合物是菜豆茎中的主要活性成分之一。然而,黄酮类化合物积累的调控机制尚缺乏。这里,通过代谢组和转录组分析研究了E.phaseoloides中不同发育阶段茎的植物化学化合物和转录本。最老茎的代谢产物特征明显不同于年轻和较老的茎组织。共检测到198种黄酮类化合物，和黄酮，黄酮醇，花青素,异黄酮,黄烷酮是主要的亚类。代谢组数据显示，幼茎和老茎中的acacetin含量明显高于最老的茎。芦丁和肉豆蔻素在最古老的茎中显示出显着较高的水平。在RNA-seq数据中鉴定并分类了总共143个MYB和143个bHLH。同时,鉴定了34个类黄酮生物合成结构基因。基于类黄酮生物合成相关结构基因的表达模式,它表明黄酮醇，花青素,原花青素的生物合成在E.phaseoloides茎的发育过程中首先活跃，花青素或原花青素生物合成分支占主导地位；黄酮生物合成分支在茎发育后期活跃。通过对转录组和代谢组数据的相关性分析,确定了与黄酮类化合物合成和运输相关的潜在候选基因。其中,MYBs,bHLH,和TTG1共同调节黄酮醇和结构基因的生物合成，bHLH和转运蛋白基因共同调节花色苷的生物合成。此外,WDR基因TTG1样（AN11）可能与IIIbbHLH和R2R3-MYB蛋白特定组合调节二氢查耳酮和黄酮醇的生物合成。此外，转运基因蛋白TRANSPARENTTESTA12样基因正调控芦丁的积累，ABC转运体B家族成员基因的同源物与黄酮刺槐素的含量呈正相关。这项研究提供了参与类黄酮生物合成的候选基因，类黄酮组成信息和类黄酮积累特征，提高了我们对E.phaseoloides茎中类黄酮生物合成的MYBs和bHLHs相关调节网络的理解，为菜豆茎中黄酮类生物合成的代谢工程提供参考。
