MicroRNA-like RNA Functions Are Required for the Biosynthesis of Active Compounds in the Medicinal Fungus Sanghuangporus vaninii.
MicroRNA-like RNA Functions Are Required for the Biosynthesis of Active Compounds in the Medicinal Fungus Sanghuangporus vaninii.
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DOI：
10.1128/spectrum.00219-22
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发表时间：
2022-12-21
影响因子：
3.7
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中科院分区：
文献类型：
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MiRNA样RNAs(MilRNAs)被认为是真核生物中转录后基因表达调控的序列特异性调节因子。然而，数以百计的真菌milRNA在代谢成分的生物合成中的功能尚不清楚。双黄孢子产生多种生物活性化合物，在亚洲国家被广泛使用。在这里，编码两个DICER、四个Argavies和四个RdRPs的基因在桑黄孢子虫中被鉴定和鉴定。由于缺乏有效的基因操作系统，喷雾诱导的基因沉默(SIGS)在瓦尼氏链霉菌中表现出高效的双链RNA(DsRNA)摄取和基因沉默效率。SIGS介导的基因敲除表明，SVRDRP-3、SVRDRP-4、SVDICER-1和SVDICER-2对菌丝生物量、类黄酮、三萜类和多糖的合成至关重要。对香草链霉菌菌丝体和子实体中的milRNAs进行了深层测序。共鉴定出31个milRNAs，其中SvmilR10、SvmilR17和SvmilR33依赖于Svrdrp-4和Svdicer-1。重要的是，Sigs介导的SvmilR10和SvmilR33的过表达导致了类黄酮、三萜类和多糖产率的显著变化。进一步的分析表明，编码逆转座子衍生蛋白PEG1和组蛋白赖氨酸N-甲基转移酶的这些milRNA靶基因在milRNA过表达菌株中可能下调。我们的结果表明，S.Vanini具有高的外部dsRNA和小的RNA摄取效率，并且milRNAs可能在生物活性化合物的生物合成中发挥关键的调节作用。重要作用真菌可以结合环境中的RNA，通过RNA干扰沉默真菌基因，从而促进Sigs的发展。瓦尼氏链霉菌对dsRNA和milRNA的高效摄取，dsRNA靶向基因的成功阻断，以及细胞内miRNA丰度的增加，表明Sigs技术是分离真菌基因和milRNAs的有效而有力的工具。我们发现Rdrp、Dird和ArgAerte基因对菌丝生物量和生物活性化合物的生产是至关重要的。我们的研究还表明，过表达的SVRDRP-4和SVDICER-1依赖的milRNAs(SvmilR10和SvmilR33)导致了这三种活性化合物产量的显著变化。本研究不仅为基于Sigs的基因和milRNA功能探索提供了第一个报道，而且为探索参与真菌代谢产物生物合成的milRNAs的功能提供了理论平台。
miRNA-like RNAs (milRNAs) have been recognized as sequence-specific regulators of posttranscriptional regulation of gene expression in eukaryotes. However, the functions of hundreds of fungal milRNAs in the biosynthesis of metabolic components are obscure. Sanghuangporus produces diverse bioactive compounds and is widely used in Asian countries. Here, genes encoding two Dicers, four Argonautes, and four RdRPs were identified and characterized in Sanghuangporus vanini. Due to the lack of an efficient gene manipulation system, the efficacy of spray-induced gene silencing (SIGS) was determined in S. vanini, which showed efficient double-stranded RNA (dsRNA) uptake and gene silencing efficiency. SIGS-mediated gene knockdown showed that SVRDRP-3, SVRDRP-4, SVDICER-1, and SVDICER-2 were critical for mycelial biomass, flavonoid, triterpenoid, and polysaccharide production. Illumina deep sequencing was performed to characterize the milRNAs from S. vanini mycelium and fruiting body. A total of 31 milRNAs were identified, out of which, SvmilR10, SvmilR17, and SvmilR33 were Svrdrp-4- and Svdicer-1-dependent milRNAs. Importantly, SIGS-mediated overexpression of SvmilR10 and SvmilR33 resulted in significant changes in the yields of flavonoids, triterpenoids, and polysaccharides. Further analysis showed that these milRNA target genes encoding the retrotransposon-derived protein PEG1 and histone-lysine N-methyltransferase were potentially downregulated in the milRNA overexpressing strain. Our results revealed that S. vanini has high external dsRNA and small RNA uptake efficiency and that milRNAs may play crucial regulatory roles in the biosynthesis of bioactive compounds. IMPORTANCE Fungi can take up environmental RNA that can silence fungal genes with RNA interference, which prompts the development of SIGS. Efficient dsRNA and milRNA uptake in S. vanini, successful dsRNA-targeted gene block, and the increase in intracellular miRNA abundance showed that SIGS technology is an effective and powerful tool for the functional dissection of fungal genes and millRNAs. We found that the RdRP, Dicer, and Argonaute genes are critical for mycelial biomass and bioactive compound production. Our study also demonstrated that overexpressed SVRDRP-4- and SVDICER-1-dependent milRNAs (SvmilR10 and SvmilR33) led to significant changes in the yields of the three active compounds. This study not only provides the first report on SIGS-based gene and milRNA function exploration, but also provides a theoretical platform for exploration of the functions of milRNAs involved in biosynthesis of metabolic compounds in fungi.
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影响因子：
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