Functional Studies of Toxoplasma gondii AMA1 and AMA2

描述(由申请人提供)：弓形虫、恶性疟原虫等弓形虫、恶性疟原虫、人类严重疟疾的病原体是主要的细胞内病原体。宿主细胞入侵、寄生虫增殖和宿主细胞裂解的反复循环是这些病毒致病的中心。 寄生虫，但对宿主细胞入侵的机制或介导这一过程的寄生虫蛋白知之甚少。一种引起人们强烈兴趣的蛋白质是AMA1。AMA1在顶端复合体中保守，而疟原虫AMA1是疟疾疫苗的主要候选者。AMA1在与宿主细胞相互作用的过程中被释放到寄生虫表面，在宿主细胞入侵过程中被认为发挥着重要的-尽管仍未确定-作用。AMA1(TgAMA1)的弓形虫同源基因最近被鉴定出来，一种新的弓形虫条件基因表达系统已经能够创建一种寄生虫系，在其中TgAMA1的表达可以被实验操纵。TgAMA1在这些寄生虫中的表达减少会导致它们的侵袭性显著降低。条件性基因敲除寄生虫为研究模型AMA1蛋白的功能提供了一个前所未有的机会。具体目标1将利用条件性击倒寄生虫和弓形虫丰富的检测手段来阻止TgAMA1在入侵中发挥的特定作用。特定目的2将探讨TgAMA1在入侵过程中从寄生虫表面蛋白水解性切割和释放的功能意义。特异靶3将鉴定和确定TgAMA2的功能，TgAMA2是一种类似AMALL的序列，也在弓形虫速殖子中表达。 弓形虫会在先天感染的胎儿和免疫受损的人中引起危及生命的疾病，包括艾滋病或霍奇金氏病患者，以及接受免疫抑制或癌症化疗的人。鉴于AMA1在侵袭中的明显重要性以及侵袭在发病机制中的核心作用，对AMA1及其相关蛋白功能的更深入了解可能有助于开发基于疫苗或化疗的新方法，以预防或控制弓形虫和其他顶丝复合体寄生虫的感染。

DESCRIPTION (provided by the applicant): Apicomplexan parasites such as Toxoplasma gondii, the causative agent of toxoplasmosis, and Plasmodium falciparum, the causative agent of severe human malaria, are obligate intracellular pathogens. Repeated cycles of host cell invasion, parasite multiplication and host cell lysis are central to the pathogenicity of these parasites, yet little is known about the mechanisms of host cell invasion or the parasite proteins that mediate the process. One protein that has been the focus of intense interest is AMA1. AMA1 is conserved among apicomplexans, and Plasmodium AMA1 is a leading malaria vaccine candidate. AMA1 is released onto the parasite surface during interaction with host cells, where it is thought to play an essential - though still undefined - role in host cell invasion. The T. gondii homolog of AMA1 (TgAMAl) has recently been identified, and a new system for conditional gene expression in T. gondii has enabled the creation of a parasite line in which expression of TgAMAl can be experimentally manipulated. A decrease in TgAMAl expression in these parasites causes a significant decrease in their invasiveness. The conditional knockdown parasites provide an unprecedented opportunity to study the function of a model AMA1 protein. Specific Aim 1 will use the conditional knockdown parasites and the wealth of assays available in T. gondii to deter mine the specific role played by TgAMAl in invasion. Specific Aim 2 will explore the functional significance of proteolytic cleavage and release of TgAMAl from the parasite surface during invasion. Specific Aim 3 will characterize and determine the function of TgAMA2, an AMAl-like sequence that is also expressed in T. gondii tachyzoites. T. gondii causes life-threatening disease in the congenitally-infected fetus and in immunocompromised persons, including those with AIDS or Hodgkins' disease, and those undergoing immunosuppressive or cancer chemotherapy. Given the apparent importance of AMA1 in invasion and the central role invasion plays in pathogenesis, a greater understanding of the function of AMA1 and AMA1-related proteins will likely contribute to the development of new vaccine-based or chemotherapeutic approaches to preventing or controlling infection by T. gondii and other apicomplexan parasites.