亚洲玉米螟头壳关键表皮蛋白的发现及功能研究

Insect head capsules, which are covered by hard cuticle, are used to protect brain as well as other inner organs and support the muscles of mouth parts and antennae. In view of its importance, understanding and utilizing of the formation mechanism of head capsule cuticle may provide new ideas for pest control. Cuticular proteins (CP) are one of the main components of insect cuticle. By comparing the transcriptomic data of the head capsule and integument cuticles from the larvae of Ostrinia furnacalis, 16 CP genes that are highly expressed in head capsule have been selected and RNAi of one gene has been performed. As a result, some larvae failed to molt and some molted larvae had abnormal head capsules. Therefore, we speculate that insect head capsules contain CPs that may play key roles in the formation of head capsule cuticle. To identify key CPs, the effects of RNAi of the candidate genes on the insect molting as well as the structures and mechanical properties of their head capsules will be first studied. Then to clarify the action mechanism of the key CPs on the structure and mechanical properties of head capsule cuticle, the key CPs will be recombinantly expressed and characterized, and the forming processes and mechanical properties of the “key CP-chitin” complexes will be investigated. This study will not only find target genes that can destroy the formation of head capsule of O. furnacalis, but also expand our understanding of the mechanism of insect cuticle formation.

昆虫头壳由坚硬表皮构成，用于保护脑等内部器官并支撑口器及触角肌肉。鉴于头壳的重要性，阐明其表皮的形成机制并加以利用，可能为害虫防治提供新思路。表皮蛋白（CP）是昆虫表皮的主要成分之一。通过比较亚洲玉米螟头壳及体壁表皮转录组，申请人选定了16个在头壳高表达的CP基因，并对其中一个进行了RNAi，结果引起幼虫蜕皮失败及头壳形态异常。因此申请人推测昆虫头壳含有特定CP，它们在其表皮形成过程中发挥关键作用。本项目首先研究候选CP基因RNAi对亚洲玉米螟蜕皮及头壳结构及机械性能的影响，从而确定对坚硬头壳表皮形成关键的CP；然后对已确定的头壳关键CP进行重组表达及性质表征，并制备“CP-几丁质”复合物，观察其形成过程并测定其机械性能，从而阐明头壳关键CP影响坚硬头壳表皮结构及机械性能的机制。本研究不仅将找到能够破坏亚洲玉米螟头壳形成的靶标基因，而且也将拓展人类对昆虫表皮形成机制方面的认识。

阐明昆虫头壳形成机制并加以利用，能够为害虫防治提供新思路。本项目首先通过比较亚洲玉米螟头壳和体壁基因组、转录组等组学数据，在头壳中鉴定出20种候选关键表皮蛋白（CP）。接下来，本项目应用了RNA干扰及CRISPR/Cas9技术证明了部分候选关键CP具有重要的生理功能。另一方面，本项目成功重组表达了候选关键CP包括OfCPH-1、OfCPR-9、OfCPR-1和OfCPH-2等；研究了其几丁质结合、共价交联以及疏水团聚等生化性质；进一步制备了几丁质-CP复合膜并测定了机械性能和微观结构，从而阐明了候选关键CP对昆虫头壳形成的关键机制。以上工作不仅找到了害虫防治的候选靶标基因，而且也拓展了人类对昆虫表皮形成机制方面的认识。受本项目资助共发表了学术论文12篇，其中SCI期刊9篇，中文核心期刊3篇，1篇在投。项目负责人参与承办了国际会议1次，培养了博士2人，硕士4人。