钙离子调控樟芝深层发酵无性产孢及其分子机制

Effect of calcium on sporulation of Taiwanofungus camphoratus in submerged fermentation

摘要:

樟芝是原产于台湾的珍稀药用真菌，具有保肝、抗癌等活性。樟芝在深层发酵过程中能够产生大量无性孢子，但是目前对樟芝无性产孢的影响因素及其分子机制尚不明确。本研究报道了樟芝发酵培养基中Ca2+浓度能够有效调控樟芝无性产孢的现象；采用双向电泳 (2-DE) 技术，对不添加Ca2+培养 (对照) 的菌丝体、添加1 mmol/L Ca2+培养 (促进产孢) 的菌丝体及添加200 mmol/L Ca2+培养 (抑制产孢) 的菌丝体进行差异蛋白质组学分析，鉴定了参与Ca2+/钙调素信号通路的CaM蛋白和HSP90蛋白，以及参与FluG调控产孢信号通路的AbaA蛋白；进一步通过生物信息学分析，预测了Ca2+/钙调素和FluG介导的樟芝无性产孢信号通路模型图；采用实时定量PCR (RT-qPCR) 技术，对该通路上23个功能基因的转录水平进行了分析，发现了受Ca2+调控最为灵敏的7个产孢相关功能基因：crz1、hsp90、flbB、brlA、abaA、wetA及fadA。本研究结果为解析樟芝无性产孢机制提供了实验依据。

Abstract:

Taiwanofungus camphoratus is a valuable and rare medicinal mushroom with various bioactivities, such as liver protection and anti-cancer. T. camphoratus can produce many arthroconidia at the end of submerged fermentation, but molecular mechanism underlying this submerged conidiation remains unknown. In this study, we found that Ca2+ concentration in culture medium significantly affected the arthroconidium production of T. camphoratus. Then, we identified two proteins (CaM and HSP90) involved in Ca2+/calmodulin signaling pathway and one protein (AbaA) involved in FluG-mediated conidiation pathway by two-dimensional electrophoresis analyses. Furthermore, we proposed a Ca2+/calmodulin- and FluG-mediated signaling pathway by bioinformatics analysis. By real-time quantitative PCR analyses of 23 key genes in the Ca2+/calmodulin- and FluG-mediated conidiation pathway, we found that expression levels of 7 genes (crz1, hsp90, flbB, brlA, abaA, wetA and fadA) showed significant responses to Ca2+ concentration in fermentation medium. Our research is beneficial for elucidating the underlying mechanism of submerged fermentation conidiation for T. camphoratus.