褐黄血蜱HSC70的抗凝机制及免疫原性研究

本项目旨在探明褐黄血蜱HSC70的抗凝机制和免疫原性，为探索控蜱防病提供新材料和策略。基于对褐黄血蜱唾液腺蛋白质组学和转录组学的前期研究，课题组克隆出了hsc70的cDNA；探明了褐黄血蜱吸血与HSC70表达的关系；证实了HSC70的抗凝活性。但HSC70确切的抗凝机制、免疫原性和抗蜱效果强弱还不明确。本研究拟以真核表达系统制备rHSC70，通过分析rHSC70-血凝因子复合物组成，明确HSC70的抗凝机制；rHSC70免疫家兔，测定抗体滴度和CD4+、CD8+ T细胞百分比，评估HSC70的免疫原性；突变hsc70的相应碱基，制备具TKD肽的融合蛋白并免疫家兔，观测增强免疫原性的效果；饲蜱于高抗体效价兔，考察HSC70和具TKD肽融合蛋白的抗蜱效果。通过本项目的研究，理论上将明确HSC70的抗凝机制，实践上对于明确HSC70能否作为优质保护性抗原开发抗蜱疫苗具有重要意义。

The aims of this study are to illuminate the anticoagulant mechanism and immunogenicity of HSC70 in Haemaphysalis flava, further to provide a new clue for controlling the tick infestations. Based on our previous proteomic and transcriptomic datasets of salivary gland of H. flava, we cloned the cDNA of hsc70. It has been proved by us that the expression of HSC70 would increase during blood feeding, and HSC70 possessed anticoagulant activity. However, the exact anticoagulant mechanism, immunogenicity and anti-tick efficacy of HSC70 are not clear. In view of this, we will produce recombinant proteins of HSC70 in eukaryotic expression system. The anticoagulation mechanism of HSC70 will be clarified by analyzing the composition of rHSC70-hemagglutination factor complex. The rabbits will be immunized by rHSC70, then the antibody titer and the CD4+/CD8+ T cells will be detected to assess the immunogenicity of HSC70. Base mutation will be induced in hsc70, the recombinant proteins fused with TKD will be produced, then the recombinant proteins fused with TKD will be used to immunize the rabbits to observe the effect of enhancing immunogenicity. Finally, the ticks will be fed on the high antibody titer rabbits to assess the anti-tick efficacy of HSC70 and the recombinant proteins fused with TKD. By this study, we will clarify the anticoagulant mechanism of HSC70. At the same time, it also will be help to evaluate the availability of HSC70 candidates for universal anti-tick vaccine development.