Functional characterization of the main integral protein components of the parasite-host cell interface of Plasmodium falciparum blood stages

人疟原虫恶性疟原虫在红细胞中的细胞内发育是疟疾临床症状的原因。在红细胞中，寄生虫与宿主细胞胞质溶胶被称为寄生虫空泡膜（PVM）的膜隔开。这种膜在寄生虫与其宿主细胞的相互作用中起着关键作用，但其许多功能知之甚少或未知。PVM含有短的、高度带电的和高度丰富的被称为EXP1和ETRAMPs（14种蛋白质的家族）的整合膜蛋白。这些分子是迄今为止唯一已知的桥接这种膜的蛋白质（它们包含一个进入宿主细胞的结构域和一个面向寄生虫的结构域）。包括EXP1在内的几种蛋白被认为是寄生虫生存所必需的，它们可能在寄生虫与宿主细胞的相互作用中起关键作用。然而，由于技术限制，迄今为止尚未对它们在恶性疟原虫寄生虫中进行功能分析。使用新技术，我们产生了EXP1的条件敲除。在这里，我们建议使用它来研究EXP1，并使用相同的技术来功能分析ETRAMP。此外，我们开发了一种新的改进的邻近生物素化方法，以确定相互作用的合作伙伴和区室蛋白质组，我们建议使用它来确定与EXP1和ETRAMP的宿主细胞面对域相互作用的因素。这也将产生PVM外表面的蛋白质组，这是用其他方法难以获得的。我们希望这项工作能够解开寄生虫宿主细胞界面的主要组成部分的功能，并有助于了解寄生虫如何利用和与宿主红细胞相互作用。

The intracellular development of the human malaria parasite Plasmodium falciparum in red blood cells is responsible for the clinical symptoms of malaria. In the red blood cell, the parasite is separated from the host cell cytosol by a membrane called the parasitophorous vacuolar membrane (PVM). This membrane plays a key role in the interaction of the parasite with its host cell but many of its functions are poorly understood or unknown. The PVM contains short, highly charged and highly abundant integral membrane proteins termed EXP1 and ETRAMPs (a family of 14 proteins). These molecules are the only so far known proteins bridging this membrane (they contain a domain reaching into the host cell and one facing the parasite). Several, including EXP1, are indicated to be essential for parasite survival and they likely play critical roles in the interaction of the parasite with the host cell. However, due to technical limitations, they have so far not been functionally analysed in P. falciparum parasites. Using new technology we generated a conditional knock out of EXP1. Here we propose to use this to study EXP1 and use the same technology to functionally analyse ETRAMPs. In addition we developed a new modified proximity biotinylation method to identify interaction partners and compartment proteomes that we propose to use to identify factors interacting with the host cell facing domain of EXP1 and ETRAMPs. This will also generate a proteome of the outer face of the PVM that with other means is difficult to obtain. We expect this work to unravel the function of the major integral components of the parasite host cell interface and to contribute to an understanding how the parasite exploits and interacts with the host red blood cell.