茧蜂病毒调控寄主粒细胞hemichannel与凋亡相互关系的研究

To understand more fully insect immunosuppressive response, we are essential to examine the pathway of immunosuppressive signaling. It is clear that Microplitis bicoloratus bracovirus (MbBV) inhibits the host cellular immune system via inducing the host granulocytes apoptosis, but the mechanism is still unknown. The key question to resolve this problem is how the apoptotic signaling transduces from extracellular to intracellular. Based on the recent research findings that the altering hemichannels contribute to the cell apoptosis through cell death signaling transduction between the extracellular and intracellular, we propose the hypothesis that the permeability of granulocytes hemichannels manipulated by MbBV is the main cause of host granulocyte apoptosis. The further investigation will focus on three questions. The first one is examining the effects of the permeability of hemocytes，the activity of caspase and the level of transcription and expression of host innexins during granulocyte apoptosis. The second one is cloning genes from PTP gene family and Glc gene family of MbBV, which are responsible for interaction with host innexins. The last one is testing if the interaction between MbBV genes and innexins disrupts the permeability of normal hemichannel resulting in the apoptosis of host insect cells.

昆虫免疫抑制信号的传递途径，是当今昆虫免疫学中免疫抑制机制研究寻求新突破的焦点。在茧蜂病毒通过诱导寄主粒细胞发生凋亡的免疫抑制过程中，了解凋亡信号的传递途径是揭示该病毒诱导粒细胞凋亡机理的关键问题。本项目抓住hemichannel (Innexin构成) 通透性变化与细胞凋亡的相关性这一关键问题，提出科学假说：茧蜂病毒调控寄主粒细胞hemichannel通透性传递凋亡信号，诱导粒细胞凋亡。拟采用细胞生物学等方法和手段加以实验验证，即：研究凋亡状态下粒细胞表面hemichannel通透性、凋亡caspase活性和寄主innexin基因的转录及表达的变化规律；筛选克隆与寄主血细胞中innexin基因发生相互作用的基因，如PTP基因族、Glc基因族中的相关基因；最后对双斑侧沟茧蜂多分DNA病毒基因与寄主innexin基因相互作用对hemichannel通透性及其凋亡caspase的影响进行研究。

前期的研究表明, 茧蜂病毒诱导寄主粒细胞凋亡，但并不清楚凋亡信号的传递途径。本研究项目中，主要的研究内容有：1）Innexin_hemichannel传递凋亡信号；2) 茧蜂病毒调控了粒细胞的Innexin_hemichannel; 3) 抑制innexin_hemichannel促进凋亡细胞形成凋亡小体。重要的研究结果：1）细胞凋亡的早期，hemichannel被关闭了，减少凋亡信号的传递，降低了邻近细胞的凋亡；2）茧蜂病毒抑制了粒细胞的通透性，关闭了hemichannel，促进了细胞的凋亡；3）调控了Innexin，促进了凋亡细胞形成凋亡小体。本项目最新的亮点在于，发现Innexin是非常关键的蛋白，维持凋亡细胞的膜的完整性，首次发现，茧蜂病毒抑制Innexin的活性，与凋亡细胞的凋亡小体的形成相关。凋亡小体的形成，有利于吞噬细胞及时清除凋亡细胞，这在无脊椎动物免疫抑制的研究中首次发现，深入的研究将促进无脊椎领域的先天免疫的研究。