Basigin促进猪圆环病毒2型复制机制的研究

Porcine circovirus type 2 (PCV2) is the causative agent of post-weaning multi-systemic wasting syndrome and triggers giant economic loss in the swine industry. Basigin is also called extracellular matrix metalloproteinase inducer, and is a transmembrane glycoprotein, plays an important role in the process of some viruses’ entry and infection. Our previous experiments results showed a change in Basigin expression in PK-15 cells after infected with PCV2, suggesting that Basigin may play an important role in the course of PCV2 infection. The function of Basigin in elevating PCV2 replication had be elucidated by up or down regulate the Basigin expression in host cells. In this study, in order to further confirm the function of Basigin in PCV2 replication, stable-expression cell line of Basigin is constructed. The course of Basigin in elevating PCV2 replication will be confirmed. Also the interaction of PCV2 protein with Basigin will be detected by laser scanning confocal microscope technique and immunoprecipitation assay. For further elucidating the mechanisms of Basigin in supporting PCV2 replication, the potential mediate proteins CyPA or CyPB’s function in Basigin elevating PCV2 replication will be studied. The research plans to found out the mechanisms of Basigin in elevating PCV2 replication and enriches the pathopoiesis mechanisms of PCV2.

猪圆环病毒2型是断奶仔猪多系统衰竭综合征的主要病原，给世界养猪业造成了巨大的经济损失；Basigin又称细胞外基质金属蛋白酶诱导蛋白，为一种跨膜糖蛋白，影响多种病毒的入侵和复制。前期研究发现，PCV2能够显著影响Basigin的表达水平，提示Basigin可能参与了PCV2感染宿主的过程；通过转染过表达质粒以及RNA干扰技术上调或者下调该蛋白在宿主细胞中的表达量，发现Basigin能够促进PCV2的复制。本研究拟构建稳定表达Basigin的细胞系，进一步验证Basigin促进PCV2复制的作用；发现Basigin影响PCV2复制的环节；采用激光共聚焦显微技术和免疫共沉淀技术探究Basigin与病毒编码蛋白之间的相互作用；同时，探索中间蛋白CyPA和CyPB是否参与了Basigin促进病毒复制的过程。通过本项目研究，发现Basigin促进PCV2复制的机制，为阐明PCV2致病机理奠定基础。

猪圆环病毒2型（PCV2）是断奶仔猪断奶后多系统消耗综合症（PMWS）的主要病原，并给养猪业造成大量经济损失。Basigin为免疫球蛋白超家族的跨膜糖蛋白；这是也是亲环蛋白CyP的受体，被证明可以影响许多病原入侵宿主的过程。在这项研究中我们发现Basigin可以提高PCV2的复制，利用抗Basigin多克隆抗体以不同浓度封闭细胞或利用siRNA干扰Basigin并感染PCV2，观察Basigin参与PCV2 感染过程的具体环节，发现Basigin影响PCV2的复制阶段，但是不影响其吸附和入胞的过程。为了排除Basigin抗体对PCV2复制的作用，利用PCV2 Cap杆状病毒表达的病毒样颗粒作用细胞，仍然发现Basigin仅影响PCV2的复制阶段而不是吸附或胞吞阶段。此外，Basigin的中间蛋白CyPA和CyPB也提高了PCV2的复制，CyP被证明可以通过激活ERK信号来促进PCV2的复制，另一方面，CsA能显著够降低PCV2的复制，也能抑制CypA与Basigin的相互作用，进一步证明Basigin-CyP相互作用对于提高PCV2复制是必需的。