亮氨酸羧基甲基转移酶（LCMT1）/磷酸酯酶（PME1）通过调控蛋白酶（PP2Ac）活性及甲基化进而影响细胞增殖。弓形虫体内同样存在LCMT1/PME1蛋白，但其生物学特性及介导的信号通路尚无报道。申请人前期研究发现Tg-LCMT1主要分布在细胞质，且与PP2Ac存在共定位及相互作用。基于此，本项目拟通过CRISPR/CAS9技术获得基因缺失/过表达虫株；利用超微结构分析、免疫荧光、流式细胞术等探究LCMT1/PME1生物学特性；利用BN-PAGE分析LCMT1/PME1对PP2A全酶复合物形成的影响；采用GST-pulldown、免疫印记等验证LCMT1/PME1与PP2Ac的互作及对其活性与甲基化的调控，阐明LCMT1/PME1调控弓形虫增殖的分子机制。研究结果将为弓形虫lcmt1/pme1基因功能研究奠定理论基础，并为筛选新型抗弓形虫药物靶位点提供新的依据。

The activity and methylation of protease PP2Ac could be controlled by leucine carboxyl methyl transferase (LCMT1)/phosphatase (PME1), and then influenced cells proliferation. LCMT1/PME1 proteins also exist in Toxoplasma gondii (T. gondii). However, there are almost no reports on the biological role of Tg-LCMT1/PME1 around the world, and the signal pathway were also unknown. In previous study, we confirmed that Tg-LCMT1, co-localized and interacted with PP2Ac, mainly distributed in the cytoplasm. In this research, CRISPR/CAS9 technology will be applied to obtain gene knockout/overexpression parasites; Then the biological characteristics were identified by ultrastructure analysis, immunofluorescence and flow cytometry; BN-PAGE was used to analyze the effect of LCMT1 / PME1 on the formation of PP2A holoenzyme complex; GST-pulldown and Western-blot will be applied to verify the interaction of LCMT1/PME1 with PP2Ac, and the regulation of its activity and methylation, we expect to elucidate the molecular mechanism by which LCMT1 / PME1 regulates the proliferation of T. gondii. The results of this study will not only provide important theoretical significance for elucidating the function of lcmt1/pme1 genes and their roles on regulation of T. gondii proliferation, but also provide new ideas for anti-toxoplasmosis drug targets discovery and vaccine improvement.