Abstract

Sanghuangporus baumii (Pilát) (Zhou and Dai), a traditional Chinese medicinal fungus, has been confirmed to produce triterpenoids, which possess a good anti-cancer effect. However, the triterpenoids yield of S. baumii is low and its biosynthetic mechanism is not clearly understood. To investigate the triterpenoids biosynthetic mechanism, the whole cDNA sequence of the HMG-CoA reductase (HMGR) gene was obtained. The open reading frame of HMGR gene cDNA named SbHMGR, comprised 4170 bp, encoding a polypeptide of 1389 amino acids with a predicted protein molecular weight of 147.53 kDa. The HMGR gene includes four exons and three introns. Subsequently, the triterpenoids content was determined at different developmental stages of S. baumii. It showed a dynamic tendency—increased first, then decreased and peaked on 14th day in the mycelium stage. In addition, the changing trend of SbHMGR enzyme activity and transcript level of SbHMGR gene were measured. They were nearly consistent with the triterpenoids content, suggesting that SbHMGR gene may play a key regulatory role in triterpenoids biosynthetic pathway. Similarly, changes in the transcript level of downstream key genes (SbMVD, SbIDI, SbLSD) of SbHMGR gene were also positively correlated with the triterpenoids content, indicating those three genes were also involved in triterpenoids biosynthetic pathway. Moreover, SbHMGR gene had been observed to be expressed in prokaryote. The results will be valuable for elucidation of the biosynthetic pathway of triterpenoids in S. baumii, lay a foundation for genetic modifications to improve triterpenoids yield.

摘要

Sanghuangporus baumii (Pilát) (Zhou and Dai) 是一种传统的中药真菌，已被证实产生具有良好抗癌作用的三萜类化合物。然而，鲍鱼三萜类化合物的产量较低，其生物合成机制尚不清楚。为了研究三萜类化合物的生物合成机制，获得了HMG-CoA 还原酶( HMGR ) 基因的全 cDNA 序列。HMGR基因 cDNA的开放阅读框SbHMGR，包含 4170 bp，编码 1389 个氨基酸的多肽，预测蛋白质分子量为 147.53 kDa。该HMGR基因包括四个外显子和三个内含子。随后，测定了鲍鱼不同发育阶段的三萜含量。呈动态趋势——在菌丝期第14天先增加后减少并达到峰值。此外，还测量了SbHMGR酶活性和SbHMGR基因转录水平的变化趋势。它们与三萜含量几乎一致，表明SbHMGR基因可能在三萜生物合成途径中起关键调控作用。类似地，在改变的下游关键基因的转录水平（SbMVD，SbIDI，SbLSD的）SbHMGR基因也与三萜含量呈正相关，表明这三个基因也参与了三萜生物合成途径。此外，已观察到SbHMGR基因在原核生物中表达。研究结果对阐明鲍鱼三萜类化合物的生物合成途径具有重要意义，为遗传修饰提高三萜类化合物产量奠定基础。