Functional analysis of the constrictome in the apicomplexan parasite Toxoplasma gondii: deciphering final events in parasite division.

顶端复合体寄生虫的细胞分裂是由有丝分裂和胞质分裂相互交织的关系决定的。这一过程发生在通过外周形成子代细胞的寄生虫中(例如，疟原虫SSP中的分裂)。或通过内部萌发(例如弓形虫的内生作用)。弓形虫速殖子通过最简单的分裂方式(内源性)分裂，每一次分裂产生两个子细胞，从而使它们成为顶端复合体细胞分裂的可及模型。这种分裂模式的关键是基本复合体，它是在新生的子细胞早期形成的可收缩的细胞骨架组合，在最终的收缩/脱落事件中发挥关键作用，完成胞质分裂。最近，在弓形虫中已经发现了几种基本的复合蛋白，但到目前为止，对其收缩力量的识别尚不清楚。有趣的是，所有已知的参与其他生物胞质分裂相关收缩的关键收缩蛋白都不存在于顶复合体基因组中，这提出了一个问题：这一关键步骤是如何在寄生虫中执行的？本项目的目标是鉴定和表征弓形虫的收缩力量，并揭示其受寄生虫控制的情况。为了实现这一点，我将使用已建立的基本复合蛋白的子集作为起点来分析这个隔室--弓形虫收缩组的完整组成和结构。在这里，我将应用BioID筛选，这是一种基于体内生物素标记感兴趣的复合体中紧密相关的蛋白质的新技术。我将使用反向遗传学来描述已确定的候选基因，重点放在可以执行和/或控制收缩力量的蛋白质上。综上所述，我的工作将突出一种进化的独特的胞质分裂模式的分子基础，从而有助于对胞质分裂的细胞生物学有基本的新见解。

Cell division in apicomplexan parasites is defined by an interwoven relationship of mitosis and cytokinesis. This process occurs in parasites that either form daughter cells by peripheral (e.g. schizogony in Plasmodium ssp.) or by internal budding (e.g. endodyogeny in Toxoplasma gondii). Toxoplasma tachyzoites divide by the simplest division mode (endodyogeny) that produces two daughter cells per division round and therefore leads to their establishment as an accessible model for apicomplexan cell division. An essential linchpin for this division mode is the basal complex, a contractile cytoskeletal assemblage formed early in nascent daughter cells and acting critically in the final constriction/abscission events to complete cytokinesis. Recently, several basal complex proteins have been identified in Toxoplasma, but the identity of the contractile force is unknown so far. Interestingly, all known key contractile proteins involved in cytokinesis related constriction in other organisms are absent from the apicomplexan genomes, raising the question how this essential step is executed in the parasite?The goal of this project is the identification and characterization of the constrictive force in Toxoplasma gondii and revelation of its control by the parasite. To accomplish this, I will use a sub-set of established basal complex proteins as a starting point to analyze the complete composition and architecture of this compartment, the Toxoplasma constrictome. Hereto I will apply BioID screening, a novel technique based on in vivo biotin labeling of closely associated proteins in a complex of interest. I will use reverse genetics to characterize the identified candidates, focusing on proteins that can execute and/or control the constrictive force. Taken together, my work will highlight the molecular basis for an evolutionary distinct mode of cytokinesis thus contributing to fundamental new insights into the cell biology of cytokinesis.