Identification and functional characterization of proteins of the mitochondrial tRNA import pathway of Toxoplasma gondii

顶复虫是一种寄生虫，在人类和动物中具有极高的感染率，危害极大。由顶复合体引起的最致命的疾病是疟疾，每年在发展中国家造成数百万人死亡。来自该群体的其他寄生虫也会导致免疫系统受损的人患上疾病，比如艾滋病患者。此外，顶复体虫在农业动物中引起疾病，造成重大的经济危害和对人类食物供应的破坏。例如，弓形虫是导致绵羊流产的最常见原因之一。这些小的单细胞寄生虫在人类或动物宿主的大细胞内生存和繁殖。因此，这些寄生虫进化出了精心设计的策略，以尽量减少暴露于宿主防御系统的风险，同时仍能从这种恶劣的环境中获取所需的代谢物。每个寄生虫体内的两个细胞器，线粒体和顶质体，通过提供寄生虫新陈代谢的基本中心，在这场斗争中起着至关重要的作用。因此，它们的功能是必不可少的，它们的维持与寄生虫的增殖密切相关。每个寄生虫都有一个线粒体，其分裂和形态的改变与寄生虫细胞分裂完全同步。这与人类和动物细胞的情况不同，在每个细胞中发现的许多线粒体复制，变化和功能独立于细胞分裂。线粒体是通过内共生获得的，即一个细胞被另一个细胞吞没，并逐渐被奴役成为依赖的细胞器。细胞器和细胞交换物质的分子运输机制必须进化，以允许这种接管。其中一个例子是线粒体tRNA输入（MITI）。由于线粒体保留了其祖先的一些基因组，它需要trna将其基因表达为功能性蛋白质。虽然在大多数生物体中，线粒体仍然保留自己的tRNA基因，但在apiccomplexa中，所有tRNA都在细胞核中编码，线粒体必须从细胞质中输入所需的所有tRNA，这使得MITI对寄生虫的生存至关重要。这一关键作用突出了MITI在控制寄生虫和线粒体分裂之间的紧密同步性方面的关键作用。尽管MITI在线粒体的进化中起着至关重要的作用，但科学家们只在少数生物体中详细描述了MITI系统，导致对其起源和发展的理解非常有限。此外，尽管MITI在顶复虫线粒体功能的控制中起着重要作用，但其在这些寄生虫中的组成和功能机制完全不清楚。在这项研究中，综合和互补的方法将被用于分类和分析MITI机器的组成部分，在最完善的实验系统中，在顶复虫，弓形虫。MITI组成和组织的详细模型将通过标记单个组件和剖析它们彼此之间以及与运输中的RNA的相互作用来生成。单个组成部分的功能，以及通产省作为一个整体的功能，将通过基因一次移除一个组成部分来分析。这不仅提供了MITI在一个新的生物群体中的第一个描述，它也将发展我们对MITI进化的理解，以及在这些寄生虫的情况下，它是如何支持它们致命的生活方式的。对尖端复合体MITI系统的深入了解也将为设计对抗弓形虫病和疟疾等疾病的新型治疗策略提供新的潜在途径。

Apicomplexa is a group of parasites that have extremely high infection rate in humans and animals and that is very harmful. The most deadly disease caused by an apicomplexan is malaria that kills millions yearly in developing countries. Other parasites from the group are also responsible for diseases in people with compromised immune systems, like AIDS patients. Still more, apicomplexans cause disease in agricultural animals, resulting in major economic harm and damage to humans food supply. Toxoplasma, for example, is one of the most frequent causes for abortions in sheep.These small, single-celled parasites live and multiply within the larger cells of their human or animal host. Accordingly, these parasites evolved elaborate tactics to minimize exposure to host defence systems while still harvesting their needed metabolites from this hostile environment. Two organelles within each parasite, called the mitochondrion and the apicoplast, play crucial roles in this struggle by providing essential centres of parasite metabolism. Consequently their function is essential and their maintenance is tightly regulated and synchronized with the parasites' proliferation. Each parasite possesses one mitochondrion that divides and changes morphology in full synchrony with the parasite cell division. This is unlike the situation in human and animal cells where the many mitochondria found in each cell replicate, change and function independent of the cell division.Mitochondria were acquired via endosymbiosis whereby one cell was engulfed by another and was gradually enslaved to become a dependent organelle. Molecular transport mechanisms through which the organelle and the cells could exchange materials had to evolve to allow this take-over. One such example is mitochondrial tRNA import (MITI). Since the mitochondria kept some of the genome of its ancestor it needs tRNAs to express its genes to functional proteins. While in most organisms the mitochondria still retain its own tRNA genes, in Apicomplexa all tRNAs are encoded in the nucleus of the cell and the mitochondrion must import all the tRNAs it needs from the cell cytosol, making MITI absolutely crucial for parasite survival. This pivotal role highlights MITI as a key player in controlling the tight synchrony between parasite and mitochondrion division. Despite the crucial important of MITI in the evolution of mitochondria, scientists have only described MITI systems in detail in a handful of organisms, resulting in a very limited understanding of its origin and development. Moreover, despite the essential role of MITI in the control of mitochondrial function in Apicomplexa, its composition and functional mechanism in these parasites are entirely elusive. In this study integrated and complementary approaches will be used to catalogue and analyse the components of the MITI machinery in the most well developed experimental system in Apicomplexa, Toxoplasma gondii. A detailed model of the MITI composition and organization will be generated via tagging of individual components and dissecting their interaction with each other and with the RNA in transit. The function of the individual components, and of MITI as a whole, will be analysed by genetically removing one component at a time. Not only will this provide the first description of MITI in a new group of organisms, it will also developed our understanding of MITI evolution, and how in the case of these parasites it supports their deadly lifestyle. A thorough understanding of the apicomplexa MITI system will also provide a new potential avenue for the design of novel therapeutic strategies to fight diseases like toxoplasmosis and malaria.