Toxoplasma endocytosis of host cytoplasm

描述（由申请人提供）：先天免疫防御有助于限制原生动物病原体刚地弓形虫的生长，但其功效被寄生虫对抗防御所抵消。例如，被感染的巨噬细胞试图通过表达与免疫相关的gtp酶（IRGs）来破坏寄生虫，IRGs装载到寄生虫的膜上并拆除寄生虫，但寄生虫通过磷酸化Irg蛋白来降低其活性来阻止这一点。由于磷酸化是可逆的，因此是暂时的，我们假设弓形虫通过将这些宿主效应蛋白内化到其内吞系统中，永久性地消除了这些宿主效应蛋白。我们最近意外的发现，寄生虫很容易内化宿主细胞质中表达的报告蛋白，这支持了这一假设。此外，报告蛋白在蛋白酶缺乏的寄生虫中最为丰富，这表明内化的宿主蛋白通常在寄生虫的内吞系统中被消化。此外，寄生液泡内缺乏管状膜生物发生的寄生虫不能有效地内化报告蛋白，这表明需要管状膜来摄取。这些有缺陷的寄生虫也极易被活化的巨噬细胞杀死。具体目的是：(1)确认寄生虫内吞系统是内化蛋白质的目的地；(2)表明弓形虫利用小孔内管膜网络将IRGs内化以破坏其内溶酶体系统。我们希望证明寄生虫通过内吞和降解作为一种免疫逃避策略来消除IRGs。这项工作预计将为减少感染期间寄生虫的细胞内存活开辟令人兴奋的新机会。

DESCRIPTION (provided by applicant): Innate immune defenses help restrict growth of the protozoan pathogen Toxoplasma gondii, but their efficacy is offset by parasite counter defenses. For example, infected macrophages attempt to destroy the parasite by expressing immunity-related GTPases (IRGs) that load onto and dismantle the parasitophorous membrane, but the parasite thwarts this by phosphorylating Irg proteins to diminish their activity. Since phosphorylation is a reversible, and therefore temporary, we hypothesize that T. gondii additionally eliminates these host effector proteins permanently by internalizing them into its endocytic system. Our recent unexpected finding that the parasite readily internalizes a reporter protein expressed in the host cytosol supports this hypothesis. Further, the reporter protein is most abundantly seen in protease-deficient parasites, suggesting that internalized host proteins are normally digested in the parasite endocytic system. Also, parasites deficient in the biogenesis of tubular membranes within the parasitophorous vacuole are unable to efficiently internalize the reporter protein, indicating that the tubular membranes are required for uptake. These deficient parasites are also highly susceptible to killing by activated macrophages. The specific aims are to: (1) confirm the parasite endocytic system as the destination of internalized proteins; and (2) show that Toxoplasma uses the intravacuolar network of tubular membranes to internalize IRGs for destruction in its endolysosomal system. We expect to show that the parasite eliminates IRGs by endocytosis and degradation as an immune evasion strategy. The work is anticipated to open exciting new opportunities to diminish parasite intracellular survival during infection.