Novel rhoptry effector proteins in Toxoplasma host-pathogen interaction

描述（由申请方提供）：弓形虫是一种专性细胞内寄生虫，可导致免疫功能低下个体的严重中枢神经系统疾病和先天性感染新生儿的出生缺陷。这些生物体的细胞内存活依赖于侵入其宿主细胞、建立允许复制的小生境和避免宿主细胞防御的能力。分泌棒状体已经成为调节入侵和劫持宿主细胞功能的关键细胞器。为了劫持宿主功能，棒状体将蛋白质注入宿主细胞质中，这些蛋白质可以被运输到感染细胞内的不同隔室并直接靶向宿主途径。这些宿主“效应物”蛋白的重要性由棒状体激酶家族例证，棒状体激酶被注射并且在调节宿主信号传导途径和调节寄生虫毒力中发挥关键作用。 除了ROP激酶，棒状体还注入一系列功能未知的其他效应蛋白。这些蛋白质中的大多数与已知蛋白质或可识别结构域缺乏同源性，并且是弓形虫和密切相关的寄生虫所特有的。我们已经确定了一组这些新的棒状效应子，并在生物学相关的II型菌株寄生虫中开发了基因敲除。在这个提议中，我们将确定这些新的效应蛋白在调节宿主功能中的作用。具体来说，我们将确定主机响应？在体外使用宿主微阵列分析和在体内通过评估小鼠中的毒力、组织嗜性和缓殖子形成来研究效应菌株。我们还将使用串联亲和标记效应蛋白在宿主细胞中的外源表达来确定它们的目的地并鉴定宿主靶标。总之，这些互补的方法有望揭示弓形虫如何使用这套独特的效应蛋白来调节其哺乳动物宿主细胞的新机制。

DESCRIPTION (provided by applicant): Toxoplasma gondii is an obligate intracellular parasite that causes severe central nervous system disorders in immunocompromised individuals and birth defects in congenitally infected neonates. Intracellular survival of these organisms is dependent on the ability to invade their host cell, establish a replication-permissive niche, and avoid host cell defenses. The secretory rhoptries have emerged as a key organelle that regulates both invasion and hijacking of host cell functions. For hijacking host functions, the rhoptries inject a burst of proteins into the host cell cytoplasm that can be trafficked to distinct compartments within the infected cell and directly target host pathways. The importance of these host "effector" proteins is exemplified by a family of rhoptry kinases that are injected and play critical roles in modulating host signaling pathways and regulating parasite virulence. In addition to ROP kinases, the rhoptries inject an array of other effector proteins that are of unknown function. Most of these proteins lack homology to known proteins or identifiable domains and are unique to Toxoplasma and closely related parasites. We have identified a panel of these novel rhoptry effectors and developed gene knockouts in the biologically relevant type II strain parasites for their study. In this proposal, we will determine the role of these novel effector proteins in regulating host functions. Specifically, we will determine the host response to ?effector strains in vitro using host microarray analysis and in vivo by assessing virulence, tissue tropism, and bradyzoite formation in mice. We will additionally use exogenous expression of tandem affinity tagged effector proteins in host cells to determine their destination and identify host targets. Together, these complementary approaches promise to reveal novel mechanistic insights into how Toxoplasma uses this unique set of effector proteins to modulate its mammalian host cell.