Combining structural biology and genetics to understand the function of a multi-gene family expanded in neglected human malaria parasites

结合结构生物学和遗传学来了解在被忽视的人类疟疾寄生虫中扩展的多基因家族的功能

• 批准号: MR/Y012895/1
• 负责人: Julian Rayner
• 金额: $ 116.09万
• 依托单位: University of Cambridge
• 依托单位国家: 英国
• 项目类别: Research Grant
• 财政年份: 2024
• 资助国家: 英国
• 起止时间: 2024 至 无数据
• 项目状态: 未结题
• 来源: https://gtr.ukri.org/projects?ref=MR%2FY012895%2F1
• 关键词: Combining; structural; biology; genetics; understand

More than a third of the world's population is at risk of contracting malaria and there are more than 200 million cases each year, leading to nearly half a million deaths. Malaria is caused by multiple species of Plasmodium parasite, which are spread from person to person by Anopheles mosquitoes. Most malaria research has focussed on Plasmodium falciparum, the dominant species in Africa and the one that causes the majority of malaria deaths. However, P. falciparum is only relatively distantly related to the species that cause most malaria cases outside Africa. These fall into a different evolutionary group which includes both Plasmodium vivax, the second most significant cause of malaria globally, and Plasmodium knowlesi, a parasite that predominantly infects monkeys in Southeast Asia but can also be transmitted to humans where it can cause severe disease and death. The biology of this group of parasites, collectively referred to as the Plasmodium subgenus, is significantly underexplored, in part because only P. knowlesi can be routinely grown and experimentally manipulated in a lab setting.In this project we will focus on a group of proteins, Tryptophan-rich antigens (TRAgs) that are found in all Plasmodium parasites but are present in significantly higher numbers in species within the Plasmodium subgenus. The P. vivax genome contains 38 genes that encode TRAgs, while P. knowlesi contains 29 TRAgs - by contrast the major African malaria parasite P. falciparum contains only 3. The function of this protein family is unknown, but the fact that they are expanded in the Plasmodium subgenus suggests that they are particularly important in these neglected human malaria parasite species. We have recently solved the three-dimensional protein structure of one Plasmodium vivax TRAG, which revealed similarities between it and proteins in other organisms that bind to the lipids that make up the membranes of all cells, including the red blood cells that Plasmodium parasites grow inside. We subsequently confirmed that both P. vivax and P. knowlesi TRAgs can bind directly to lipids, and in the case of one P. knowlesi TRAg, that this interaction is involved in the process by which these parasites recognise and invade human red blood cells. This for the first time raises a clear hypothesis for TRAg function - that they bind to lipids and are involved in manipulating human red blood cell membranes, either during or after the process of invasion.We will explore this hypothesis by carrying out the first systematic study of a large number of TRAgs in a Plasmodium subgenus parasite species. We will focus on P. knowlesi, which can be grown in the lab and genetically manipulated, which makes it possible to probe the location and function of individual TRAgs. We will extend our structural studies, exploring whether different TRAgs have specificity for different lipids, and solving the structure of TRAg proteins in complex with lipids found in the membrane of human red blood cells. We will identify which TRAg genes are expressed in the lab strain of P. knowlesi and establish the specific localisation of the most abundant TRAg proteins using advanced microscopy. We will also leverage recent technical advances to create P. knowlesi parasite lines in which these highly expressed TRAg genes have been deleted using CRISPR-Cas9 genome editing and monitor the effect on parasite growth and invasion. Overall, this research will provide critical information about the function of an understudied protein family in a neglected malaria parasite species.

全世界三分之一以上的人口处于患疟疾的风险，每年有超过2亿个案件，导致近50万人死亡。疟疾是由多种疟原虫寄生虫引起的，这些疟原虫由蚊子散布在人到人之间。大多数疟疾研究都集中在恶性疟原虫上，这是非洲的主要物种，也是导致大多数疟疾死亡的一种。但是，恶性疟原虫仅与在非洲以外的大多数疟疾病例的物种相对较远。这些属于不同的进化群，其中包括疟原虫，全球疟疾的第二重要原因和疟原虫，这是一种寄生虫，这种寄生虫主要会感染东南亚的猴子，但也可以传播到可能引起严重疾病和死亡的人类。这组寄生虫的生物学（统称为疟原虫亚属）被显着不流失，部分原因是，只有诺斯蒂（P. knotlesi）才能常规生长并在实验室环境中进行实验性操纵。在该项目中，我们将集中在较高的蛋白质中，在较高的蛋白质中，在所有蛋白质中均具有较高的质量（trags），但在所有plastiiuim中均具有大量的质量，但在所有plastiiuim中都具有plassodious（trags），但在所有蛋白质中都属于所有plastious，但在所有蛋白质中都属于所有质量，但在所有蛋白质中都属于所有蛋白质（trags）。亚属。维瓦克斯基因组包含38个编码trags的基因，而P. knowlesi则包含29个trags-相比之下，非洲主要的疟疾寄生虫只包含3个。该蛋白质家族的功能尚不清楚，但事实是，它们在这些疟原虫中却在这些质量化的疟原虫中特别重要。最近，我们解决了一个疟原虫的体内trag的三维蛋白质结构，该结构揭示了它与构成所有细胞膜的脂质结合的其他生物体中的相似性，包括所有细胞的膜，包括质子寄生虫的红细胞。随后，我们证实了植发疟原虫和诺尔斯氏菌trags都可以直接与脂质结合，并且对于一个诺尔斯氏菌trag，这种相互作用涉及这些寄生虫识别并侵犯人类红细胞的过程。这首先提出了对Trag功能的明确假设 - 它们与脂质结合并参与了在入侵过程中或之后或之后或之后或之后操纵人类红细胞膜的作用。我们将通过对疟原虫帕拉斯菌群中大量trags进行首次系统研究来探索这一假设。我们将专注于P. knowlesi，可以在实验室中生长并进行遗传操纵，这使得可以探究单个trag的位置和功能。我们将扩展我们的结构研究，探索不同的trags对不同脂质的特异性，并求解与人类红细胞膜中发现的脂质中的trag蛋白结构。我们将确定哪些trag基因在P. knowlesi的实验室菌株中表达，并使用晚期显微镜建立最丰富的Trag蛋白的特定定位。我们还将利用最近的技术进步来创建诺尔斯寄生虫线​​，其中这些高度表达的TRAG基因已使用CRISPR-CAS9基因组编辑删除，并监测对寄生虫生长和入侵的影响。总体而言，这项研究将提供有关被忽视的疟疾寄生虫物种中研究蛋白质家族的功能的关键信息。