凋亡是细胞的基本生物学现象，在生命活动和多种疾病过程中发挥重要作用。Caspases是哺乳动物及多种生物中调控细胞凋亡的关键因子。研究发现凋亡诱导剂可以成功诱导弓形虫的凋亡，从感染小鼠腹腔分离的弓形虫也存在凋亡现象，但未见关于弓形虫凋亡关键因子及凋亡机制的报道。本课题组前期已经开展弓形虫凋亡研究，通过生物信息学分析发现，虽然其不存在Caspases，但存在与其类似的Metacaspase，后者在酵母、拟南芥等生物中具有调控凋亡的功能，我们随后证实其是调控弓形虫凋亡的关键因子。本项目拟在此基础上，构建弓形虫Metacaspase（TgMCA）功能域调控虫株及氨基酸突变虫株，比较不同虫株的凋亡，明确TgMCA发挥凋亡作用的分子基础；检测凋亡虫体的线粒体、内质网及死亡受体变化，确认TgMCA被激活的上游路径；挖掘TgMCA作用的靶蛋白，明确其下游路径。全面解析弓形虫凋亡的分子机制。

Apoptosis, a basic phenomenon of physiology, is not only essential to normal development of mammals and other living organisms but also important in the process of diseases. Caspases are central executioners of apoptosis in mammals and many other species. Previous studies showed Toxoplasma gondii exhibited apoptosis upon exposure to apoptosis inducers, and apoptotic parasites were also identified in the peritoneal lavage fluid of mice infected with T. gondii by intraperitoneal injection, but the mechanism of apoptosis in T. gondii has not been elucidated. Our group has conducted preliminary researches on apoptosis in T. gondii. We found that caspases were not existent in T. gondii but metacaspase, which shares structural similarity to caspases, are present in the parasite. Metacaspases are involved in apoptosis in Saccharomyces and Arabidopsis. We have demonstrated that metacaspase is a key factor of apoptosis in T. gondii. Further to our above findings and to elucidate the molecular mechanisms and pathways of apoptosis of T. gondii, we propose to generate metacaspase functional domain knock-out lines and amino acid mutants and compare these genetically modified lines with the wild type. In addition, we will also study the mitochondria，endoplasmic reticulum and death receptor of the parasite lines to understand the pathway and mechanism of metacaspase activation. Downstream molecules of metacaspase will be explored as well to understand the roles of activated metacaspase and downstream molecules in apoptosis. The proposed study findings would enable us to have a comprehensive understanding of the molecular mechanism of and the role of metacaspase in the apoptosis of T. gondii.