Department of Pathogen Biology, Guangdong Provincial Key Laboratory of Tropical Disease Research, School of Public Health, Southern Medical University, Guangzhou, Guangdong Province510515, P.R. China.
Department of Biological Sciences, Dietrich School of Arts and Sciences, University of Pittsburgh, Pittsburgh, Pennsylvania15260, USA.
Parasitology. 2020 Nov;147(13):1433-1442. doi: 10.1017/S0031182020001249. Epub 2020 Jul 30.
Toxoplasma gondii rhoptry protein TgROP18 is a polymorphic virulence effector that targets immunity-related GTPases (IRGs) in rodents. Given that IRGs are uniquely diversified in rodents and not in other T. gondii intermediate hosts, the role of TgROP18 in manipulating non-rodent cells is unclear. Here we show that in human cells TgROP18I interacts with the interferon-gamma-inducible protein N-myc and STAT interactor (NMI) and that this is a property that is unique to the type I TgROP18 allele. Specifically, when expressed ectopically in mammalian cells only TgROP18I co-immunoprecipitates with NMI in IFN-γ-treated cells, while TgROP18II does not. In parasites expressing TgROP18I or TgROP18II, NMI only co-immunoprecipitates with TgROP18I and this is associated with allele-specific immunolocalization of NMI on the parasitophorous vacuolar membrane (PVM). We also found that TgROP18I reduces NMI association with IFN-γ-activated sequences (GAS) in the IRF1 gene promoter. Finally, we determined that polymorphisms in the C-terminal kinase domain of TgROP18I are required for allele-specific effects on NMI. Together, these data further define new host pathway targeted by TgROP18I and provide the first function driven by allelic differences in the highly polymorphic ROP18 locus.
刚地弓形虫棒状体蛋白 TgROP18 是一种多态性毒力效应因子，可靶向啮齿动物中的免疫相关 GTPases（IRGs）。鉴于 IRGs 在啮齿动物中是独特多样化的，而在其他刚地弓形虫中间宿主中则没有，因此 TgROP18 在操纵非啮齿动物细胞中的作用尚不清楚。在这里，我们表明在人类细胞中 TgROP18I 与干扰素-γ诱导的蛋白质 N-myc 和 STAT 相互作用因子（NMI）相互作用，这是 I 型 TgROP18 等位基因所特有的特性。具体而言，当在哺乳动物细胞中外源表达时，只有 TgROP18I 在 IFN-γ 处理的细胞中与 NMI 共免疫沉淀，而 TgROP18II 则不会。在表达 TgROP18I 或 TgROP18II 的寄生虫中，NMI 仅与 TgROP18I 共免疫沉淀，这与 NMI 在寄生泡膜（PVM）上的等位基因特异性免疫定位相关。我们还发现 TgROP18I 降低了 NMI 与 IFN-γ 激活序列（GAS）在 IRF1 基因启动子中的结合。最后，我们确定 TgROP18I 的 C 末端激酶结构域中的多态性是其对 NMI 产生等位基因特异性影响所必需的。总之，这些数据进一步定义了 TgROP18I 靶向的新宿主途径，并提供了第一个由高度多态性 ROP18 基因座中等位基因差异驱动的功能。
