Key metabolism pathways and regulatory mechanisms of high polysaccharide yielding in Hericium erinaceus. Key metabolism pathways and regulatory mechanisms of high polysaccharide yielding in Hericium erinaceus

The genome of H. erinaceus original strain HEA was reported 38.16 Mb, encoding 9,780 predicted genes by single-molecule, real-time sequencing technology. The phylogenomic analysis showed that H. erinaceus had the closest evolutionary affinity with Dentipellis sp. The polysaccharide content in the fermented mycelia of mutated strains HEB and HEC, which obtained by ARTP mutagenesis in our previous study, was improved by 23.25% and 47.45%, and a new β-glucan fraction with molecular weight 1.056×10^6 Da was produced in HEC. Integrative analysis of transcriptome and proteomics showed the upregulation of the carbohydrate metabolism pathway modules in HEB and HEC might lead to the increased production of glucose-6P and promote the repeating units synthesis of polysaccharides. qPCR and PRM analysis confirmed that most of the co-enriched and differentially co-expressed genes involved in carbohydrate metabolism shared a similar expression trend with the transcriptome and proteome data in HEB and HEC. Heatmap analysis showed a noticeably decreased protein expression profile of the RAS-cAMP-PKA pathway in HEC with a highly increased 47.45% of polysaccharide content. The S phase progression blocking experiment further verified that the RAS-cAMP-PKA pathway's dysfunction might promote high polysaccharide and β-glucan production in the mutant strain HEC. Overall design: In this study, we reported the original strain genome of H. erinaceus using a single molecule, real-time sequencing technology, and multi-omics analysis was carried out between mutant strains obtained by ARTP mutagenesis and the original strain. Multi-omics analysis indicated that the increased carbohydrate metabolism and the production of glucose-6P constitute the basis of high polysaccharide yield in ARTP mutated strain. Furthermore, the RAS-cAMP-PKA pathway's decreased activity might promote high polysaccharide and β-glucan production through the block of S phase progression. The study revealed the mechanism of the increased polysaccharide synthesis induced by ARTP mutagenesis associated with carbohydrate metabolism and glucose signaling regulation dysfunction and provided the critical theoretical and practical basis for polysaccharide production in H. erinaceus.

本研究中，猴头菌（H. erinaceus）原始菌株HEA的基因组经单分子实时测序（single-molecule, real-time sequencing）解析，大小为38.16 Mb，共编码9780个预测基因。系统发育基因组学分析显示，猴头菌与齿皮菌属未定种（Dentipellis sp.）的进化亲缘关系最近。本团队前期通过常压室温等离子体（ARTP）诱变获得的突变菌株HEB与HEC，其发酵菌丝体的多糖含量分别提升23.25%与47.45%，且HEC中产生了一种分子量为1.056×10^6 Da的新型β-葡聚糖组分。转录组与蛋白质组整合分析表明，HEB与HEC中碳水化合物代谢通路模块的上调可促进葡萄糖-6-磷酸（glucose-6P）的生成，并推动多糖重复单元的合成。定量聚合酶链反应（qPCR）与平行反应监测（PRM）分析证实，参与碳水化合物代谢的多数共富集且差异共表达基因的表达趋势，与HEB和HEC中的转录组、蛋白质组数据一致。热图分析显示，在多糖含量大幅提升47.45%的HEC中，RAS-cAMP-PKA通路的蛋白质表达谱显著下调。S期进程阻滞实验进一步验证，RAS-cAMP-PKA通路功能异常可促进突变菌株HEC中高水平的多糖与β-葡聚糖生成。整体设计：本研究通过单分子实时测序技术解析了猴头菌原始菌株的基因组，并对ARTP诱变获得的突变菌株与原始菌株开展多组学分析。多组学分析结果表明，碳水化合物代谢增强以及葡萄糖-6-磷酸的生成，是ARTP诱变菌株实现高多糖产量的核心基础。此外，RAS-cAMP-PKA通路活性下调可通过阻滞S期进程，促进多糖与β-葡聚糖的高水平合成。本研究揭示了ARTP诱变诱导猴头菌多糖合成增强的分子机制，该机制与碳水化合物代谢及葡萄糖信号调控功能异常密切相关，为猴头菌多糖的生产应用提供了重要的理论与实践依据。