Molecular Analysis of the PfEMP1 Binding Activity

描述（由申请人提供）：恶性疟原虫感染的红细胞（IEs）在宿主体内持续存在，并通过改变IE表面称为恶性疟原虫红细胞膜蛋白1 （PfEMP1）的细胞粘附蛋白家族的表达而避免在脾脏中被清除。不同的pfemp1结合不同的宿主受体，从而靶向IEs在不同器官的微血管中隔离，这反过来决定了疾病的严重程度。PfEMP1表面转运是一个多步骤的过程，包括蛋白质通过寄生物液泡膜（PVM）输出（早期转运）和蛋白质通过红细胞浆（通过毛勒裂隙细胞器）转移到IE表面（晚期转运）。每个寄生虫基因组编码约60个PfEMP1蛋白，这些蛋白以互斥的方式表达。大多数蛋白与宿主受体CD36结合，除了一种异常保守的介导胎盘中受感染红细胞隔离的PfEMP1变异。该项目旨在阐明PfEMP1在IE表面运输和组装的重要决定因素，并定义PfEMP1蛋白如何在强烈的抗体压力下进化以维持关键的结合相互作用。利用外源重组蛋白和表达转基因miniPfEMP1蛋白的寄生虫系进行体外和体内试验，将用于确定参与PfEMP1转运和功能的辅助蛋白，并绘制细胞粘附中的关键相互作用残基。这些研究将有助于详细描述恶性疟原虫感染红细胞的细胞粘附特性，并有助于更好地了解疟疾发病机制的分子基础。

DESCRIPTION (provided by applicant): Plasmodium falciparum infected erythrocytes (IEs) persist in the host and avoid clearance in the spleen by varying expression of a family of cytoadhesion proteins at the IE surface called P. falciparum erythrocyte membrane protein 1 (PfEMP1). Different PfEMP1s bind to different host receptors and thereby target IEs to sequester in the microvasculature of different organs, which in turn determines disease severity. PfEMP1 surface transport is a multi-step process that involves protein export across the parasitophorus vacuole membrane (PVM) (early transport) and protein transfer through the erythrocyte cytoplasm (via Maurer's cleft organelles) to the IE surface (late transport). Each parasite genome encodes ~60 PfEMP1 proteins that are expressed in a mutually exclusion fashion. Most proteins bind to the host receptor CD36, except for an unusually conserved PfEMP1 variant that mediates infected erythrocyte sequestration in the placenta. This project is designed to elucidate important determinants in PfEMP1 transport and assembly at the IE surface and define how PfEMP1 proteins have evolved to maintain key binding interactions despite intense antibody pressure. A combination of in vitro and in vivo assays employing heterologous recombinant proteins and parasite lines expressing transgenic miniPfEMP1 proteins will be used to define accessory proteins involved in PfEMP1 transport and function and to map critical interaction residues in cytoadhesion. These studies will contribute to a detail characterization of the cytoadhesive properties of P. falciparum infected erythrocytes and enable a greater understanding of the molecular basis of malaria pathogenesis.