MicroRNAs (miRNAs) are non-coding endogenous single-stranded RNAs that regulate target gene expression by reducing their transcription and translation. Several miRNAs in plants function in secondary metabolism. The dried root of Scutellaria baicalensis Georgi is a traditional Chinese medicine that contains flavonoids (baicalin, wogonoside, and baicalein) as its main active ingredients. Although the S. baicalensis genome sequence has been published, information regarding its miRNAs is lacking. In this study, 12 small RNA libraries of different S. baicalensis tissues were compiled, including roots, stems, leaves, and flowers. A total of 129 miRNAs were identified, including 99 miRNAs from 27 miRNA families and 30 predicted miRNAs. Furthermore, 46 reliable target genes of 15 miRNA families were revealed using psRNAtarget and confirmed by degradome sequencing. It was speculated that the microRNA858 (miR858)–SbMYB47 module might be involved in flavonoid biosynthesis. Transient assays in Nicotiana benthamiana leaves indicated that miR858 targets SbMYB47 and suppresses its expression. Artificial miRNA-mediated knockdown of miR858 and overexpression of SbMYB47 significantly increased the flavonoid content in S. baicalensis hairy roots, while SbMYB47 knockdown inhibited flavonoid accumulation. Yeast one-hybrid and dual-luciferase assays indicated that SbMYB47 directly binds to and activates the S. baicalensis phenylalanine ammonia-lyase 3 (SbPAL-3) and flavone synthase II (SbFNSⅡ-2) promoters. Our findings reveal the link between the miR858–SbMYB47 module and flavonoid biosynthesis, providing a potential strategy for the production of flavonoids with important pharmacological activities through metabolic engineering.

中文翻译：

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microRNA858 抑制黄芩转录因子基因 SbMYB47 并调节黄酮类化合物的生物合成


MicroRNA （miRNA） 是非编码内源性单链 RNA，通过减少靶基因的转录和翻译来调节靶基因表达。植物中的几种 miRNA 在次生代谢中发挥作用。黄芩 （Scutellaria baicalensis Georgi） 的干根是一种中药，其主要活性成分是黄酮类化合物（黄芩苷、沃戈诺苷和黄芩素）。尽管 S. baicalensis 基因组序列已经发表，但缺乏有关其 miRNA 的信息。在这项研究中，汇编了 12 个不同 S. baicalensis 组织的小 RNA 文库，包括根、茎、叶和花。共鉴定出 129 个 miRNA，包括来自 27 个 miRNA 家族的 99 个 miRNA 和 30 个预测的 miRNA。此外，使用 psRNAtarget 揭示了 15 个 miRNA 家族的 46 个可靠靶基因，并通过降解组测序证实。据推测，microRNA858 （miR858）–SbMYB47 模块可能参与类黄酮的生物合成。在本氏烟草叶片中的瞬时测定表明，miR858 靶向 SbMYB47 并抑制其表达。人工 miRNA 介导的 miR858 敲低和 SbMYB47 过表达显著提高了黄酮含量，而 SbMYB47 敲低抑制了黄酮积累。酵母单杂交和双荧光素酶测定表明，SbMYB47 直接结合并激活黄酮苯丙氨酸解氨酶 3 （SbPAL-3） 和黄酮合酶 II （SbFNSII.-2） 启动子。我们的研究结果揭示了 miR858-SbMYB47 模块与类黄酮生物合成之间的联系，为通过代谢工程生产具有重要药理活性的类黄酮提供了一种潜在的策略。