PDF(Pubmed)
Abstract：
Wolfiporia cocos is a medicinal mushroom used in China. It biosynthesizes pachymic acid (PA), a main therapeutic triterpene associated with therapies. Nowadays, the unknown PA biosynthesis leads to difficulties in increasing its content in W. cocos. Herein, we report sequencing, assembling, and characterization of the genome and several transcriptomes of W. cocos. Sequence mining determined candidate genes that encode lanosterol synthase, sterol O-acyltransferase, and sterol C-24 methyltransferase likely involved in the steps from lanosterol to PA. Gene cluster analysis identified four CYP450 cDNAs likely involved in the biosynthesis of PA, namely WcCYP64-1, WcCYP64-2, WcCYP52, and WcCYP_FUM15, which were subjected to both overexpression and silencing in mycelia. The overexpression of each of WcCYP64-1, WcCYP52 and WcCYP_FUM15 increased the content of PA, 16α-hydroxytrametenolic acid, eburicoic acid, and tumulosic acid, while the silencing of each gene either significantly or slightly decreased the contents of these four compounds, indicating their involvement in the PA biosynthesis. In addition, different temperatures affected the expression of these genes and the formation of PA. By contrast, the overexpression and silencing of WcCYP64-2 did not alter the formation of these compounds. Taken together, these findings determine more potential steps in the biosynthetic pathway of PA for metabolic engineering.
摘要：
Wolfaporiacocos是在中国使用的药用蘑菇。它生物合成胞嘧啶酸（PA），与治疗相关的主要治疗性三萜。如今,未知的PA生物合成导致难以增加其在W.cocos中的含量。在这里，我们报告测序，装配,W.cocos的基因组和几个转录组的表征。序列挖掘确定了编码羊毛甾醇合酶的候选基因，甾醇O-酰基转移酶，和甾醇C-24甲基转移酶可能参与了从羊毛甾醇到PA的步骤。基因簇分析确定了四个CYP450cDNA可能参与PA的生物合成，即WcCYP64-1，WcCYP64-2，WcCYP52和WcCYP_FUM15，它们在菌丝体中同时经历过表达和沉默。WcCYP64-1，WcCYP52和WcCYP_FUM15的过表达均增加了PA的含量，16α-羟基三甲酚酸，土利酸,和纤维素酸，而每个基因的沉默显著或轻微降低了这四种化合物的含量，表明它们参与PA的生物合成。此外,不同温度影响这些基因的表达和PA的形成。相比之下,WcCYP64-2的过表达和沉默不会改变这些化合物的形成。一起来看,这些发现确定了PA用于代谢工程的生物合成途径中更多的潜在步骤。
