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)持续存在于宿主体内，通过改变IE表面称为恶性疟原虫红细胞膜蛋白1(PfEMP1)的一系列细胞黏附蛋白的表达来避免脾内清除。不同的PfEMP1与不同的宿主受体结合，从而将IES靶向隔离在不同器官的微血管中，这反过来又决定了疾病的严重程度。PfEMP1的表面转运是一个多步骤的过程，包括蛋白质通过寄生虫液泡膜(PVM)的早期转运(早期转运)和蛋白质通过红细胞胞浆(通过毛雷氏裂隙细胞器)到IE表面的转运(后期转运)。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。大多数蛋白质与宿主受体CD36结合，除了一个异常保守的PfEMP1变体，它介导感染的红细胞在胎盘中的隔离。该项目旨在阐明PfEMP1在IE表面运输和组装的重要决定因素，并确定PfEMP1蛋白如何进化以在强大的抗体压力下维持关键的结合相互作用。使用异源重组蛋白和表达转基因minIPfEMP1蛋白的寄生虫系的体外和体内检测相结合的方法，将被用于确定参与PfEMP1运输和功能的辅助蛋白，并定位细胞黏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黏附特性，并使人们能够更好地了解疟疾发病的分子基础。

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.