长非编码RNA调控捕食线虫真菌捕食器官形成的功能与机制

The key of one fungus infects against its hosts is to switch its lifestyles from saprophytic to parasitic. This is the most important scientific question needs more studies in the future. The formation of trapping devices by nematode-trapping fungi is an important indicator of their switch from the saprophytic to the predacious lifestyles. Previous studies suggested that the formation of trap structures of Arthrobotrys oligospora maybe regulated by some long noncoding RNAs. In this study, the samples collected from the four key time points during trap formation of A. oligospora will be high throughput sequenced based on RNA-seq. The lncRNAs that might have functions in trap formation will be predicated through bioinformatic methods. Moreover, the biological functions of lncRNA will be characterized by using molecular biotechnologies. All these studies will help us from the perspective of long noncoding RNAs to understand the molecular mechanism of nematode-trapping fungi switching its lifestyles from saprophytic to predacious, also provide a basis for elucidating the pathogenicity mechanism of fungi and provide theoretical foundation for utilizing these fungi as high effective bio-control agents in the future.

真菌从腐生到寄生的生活方式转换是实现其侵染的关键，是生命科学领域有待加强研究的重要科学问题。捕食器官的形成是捕食线虫真菌实现对线虫侵染，从腐生转向寄生的标志。前期研究提示在捕食线虫真菌寡孢节丛孢中可能存在某些长非编码RNA（lncRNA）参与调控捕食器官的形成。本项目拟以寡孢节丛孢为研究对象，通过高通量测序技术对捕食器官形成过程中的几个关键节点进行测序，利用生物信息学手段筛选可能在寡孢节丛孢捕食器官产生过程中发挥功能的lncRNA，进而利用分子生物学手段对lncRNA进行实验验证及生物学功能研究，探明在寡孢节丛孢等捕食线虫真菌通过产生捕食器官实现从腐生向寄生生活转换过程中lncRNA的功能和调控机制，为理解真菌通过转换生活方式实现侵染的致病机理奠定理论基础，为开发有效的线虫生防产品提供理论指导。

本研究通过RNA-seq技术对寡孢节孢野生型菌株捕食器官形成关键时间点进行测序和分析，成功筛选出1951条候选的lncRNAs，并从中选取了54条经生物信息学预测的有显著表达差异的lncRNAs进行RT-qPCR验证，得到30条具有差异表达的lncRNAs。鉴于lncRNAs可以通过cis作用对其邻近基因进行调控，本研究对具有表达差异的lncRNAs上下游2000 bp范围内的基因进行分析，最终选取了5条lncRNAs（TCONS_00193747，TCONS_00349760，TCONS_00182722，TCONS_00188549，TCONS_00316812）和其邻近的功能基因（AOL_s00080g63，AOL_s00215g323utp215，AOL_s00080g393，AOL_s000193g139）开展功能研究，发现敲除lncRNA及其邻近的编码基因后，寡孢节丛孢的捕食器官或分生孢子的形成受到一定程度的影响，而基于RT-qPCR和/或Northern blot方法发现，敲除了lncRNA后，其邻近功能基因的表达水平也受到影响，提示这些lncRNAs可能对其邻近功能基因具有调控功能。本研究为今后进一步探明lncRNAs调控捕食线虫真菌捕食器官形成的机制奠定了基础。