森林革蜱低温响应的转录调控及重要基因的功能研究

Ticks are obligate blood-sucking ectoparasites, and can transmit a great variety of pathogens which cause lots of diseases, thus resulting in severe harmful to the health of human and animals. Low temperature affects the occurrence, distribution and dispersion of the tick population, and therefore, restricts the incidence and transmission of the tick-borne diseases. Hence, elucidation of the responsive mechanism of ticks against cold temperature will be helpful to understand the rule in their population adaptation and limitation in their geographical range, and may also play important role for thorough undestanding of the epidemic regularity of tick-borne diseases, whereas the systematic researches have not yet been developed. Based on our previous knowledge on the cold hardiness and biological characteristics of the important vector tick-Dermacentor silvarum which distributes widely in China, this work objects to systematically investigate the transcriptomic differences before and after cold hardening. Through high throughput sequencing, de novo assembly and complex bioinformatics analysis, we are intending to construct the gene regulation networks, discover the cell pathways, and analyze the functions of those important genes in the adaptation of D. silvarum to cold stress. Combining with the results accumulated in our previous projects, we will reveal the adaptation strategies of the tick D. silvarum to winter temperatures, and deepen the research on the mechanisms of ticks to cold stress into cellular and molecular level. The above results will establish foundation for accurate understanding and effective utilization of the key functional genes of the tick against cold stress, and provide us theoretical and practical suggestions on the integrate control and future forecast of ticks and tick-borne diseases.

蜱类是专性吸血的外寄生动物，传播多种病原体而导致疾病，严重危害人畜健康。低温影响蜱类的种群发生、分布与扩散，进而制约蜱媒疾病的传播与扩散，因此明确蜱类低温响应机制有助于掌握其种群适应规律和地理分布极限，同时为了解蜱媒疾病流行规律提供基础资料，其系统研究尚未开展。本研究以我国广泛分布的重要媒介蜱种——森林革蜱Dermacentor silvarum为研究对象，在前期对其生物学特性和耐寒特性深入研究基础上，通过引入转录组学研究策略，结合高通量转录组测序和de novo 组装及深度的生物信息学分析，系统研究驯化前后转录谱差异，构建相关基因的调控网络和信号通路，并对重要响应基因进行功能研究，结合前期项目积累的资料，深入阐明森林革蜱越冬适应对策，将其低温响应机制研究深入到分子与细胞水平。研究结果为深入挖掘和有效利用蜱类抵御低温的关键功能基因奠定基础，并为后续蜱及蜱媒疾病的综合防控与预测提供理论依据。

低温影响蜱类的种群发生、分布与扩散，进而制约蜱媒疾病的传播与扩散。因此本项目以我国广泛分布的重要媒介蜱种——森林革蜱为研究对象，利用高通量测序、蛋白质组学、生物信息学分析及RNAi技术系统研究森林革蜱低温响应的分子调控机制，并对重要响应基因进行功能研究，目前取得的主要研究结果包括：（1）明确森林革蜱成蜱耐寒性高于若蜱，分析了低温处理后森林革蜱成蜱甘油、多元糖、脂和蛋白质等生化物质变化，揭示了甘油在森林革蜱低温响应中的重要作用；（2）对低温驯化前后森林革蜱进行转录组测序，组装共得到136,740条unigene，筛选得到差异基因2196条，其中上调基因745条，下调基因1451条。同时对差异表达基因进行Cluster聚类、GO功能富集及COG分类；（3）利用LC-MS/MS蛋白质组学技术鉴定到蛋白序列337条，其中驯化前鉴定到蛋白136条，驯化后鉴定到蛋白267条，并分析差异蛋白的生物学功能；（4）利用RNAi技术对重要的差异基因进行功能分析，明确了HSP70、HSP90和Tubulin等基因对森林革蜱低温存活的影响。研究结果对后续深入挖掘和有效利用蜱类抵御低温的关键功能基因奠定基础，并为后续蜱及蜱媒疾病的综合防控与预测提供理论依据。