The Conoid Associated Motor MyoH Is Indispensable for Toxoplasma gondii Entry and Exit from Host Cells

Many members of the phylum of Apicomplexa have adopted an obligate intracellular life style and critically depend on active invasion and egress from the infected cells to complete their lytic cycle. Toxoplasma gondii belongs to the coccidian subgroup of the Apicomplexa, and as such, the invasive tachyzoite contains an organelle termed the conoid at its extreme apex. This motile organelle consists of a unique polymer of tubulin fibres and protrudes in both gliding and invading parasites. The class XIV myosin A, which is conserved across the Apicomplexa phylum, is known to critically contribute to motility, invasion and egress from infected cells. The MyoA-glideosome is anchored to the inner membrane complex (IMC) and is assumed to translocate the components of the circular junction secreted by the micronemes and rhoptries, to the rear of the parasite. Here we comprehensively characterise the class XIV myosin H (MyoH) and its associated light chains. We show that the 3 alpha-tubulin suppressor domains, located in MyoH tail, are necessary to anchor this motor to the conoid. Despite the presence of an intact MyoA-glideosome, conditional disruption of TgMyoH severely compromises parasite motility, invasion and egress from infected cells. We demonstrate that MyoH is necessary for the translocation of the circular junction from the tip of the parasite, where secretory organelles exocytosis occurs, to the apical position where the IMC starts. This study attributes for the first time a direct function of the conoid in motility and invasion, and establishes the indispensable role of MyoH in initiating the first step of motility along this unique organelle, which is subsequently relayed by MyoA to enact effective gliding and invasion.顶复门（Apicomplexa）的多数类群演化出专性胞内寄生的生活方式，其完成裂解周期高度依赖主动入侵宿主细胞以及从感染细胞中逸出的过程。刚地弓形虫（Toxoplasma gondii）隶属于顶复门的球虫亚类，其侵袭性速殖子（tachyzoite）的极端顶端处存在一类名为顶锥（conoid）的细胞器。该运动细胞器由独特的微管蛋白纤维聚合物构成，可在滑行运动与入侵宿主细胞的寄生虫中伸出。在顶复门中保守存在的第XIV类肌球蛋白A（MyoA），已被证实对寄生虫的运动、入侵以及从感染细胞中逸出发挥关键作用。MyoA滑行体（glideosome）锚定在内膜复合物（IMC）上，被认为可将由微线体（micronemes）与棒状体（rhoptries）分泌的环状连接复合体转运至寄生虫的后端。本研究对第XIV类肌球蛋白H（MyoH）及其相关轻链进行了全面的功能表征。我们证实，位于MyoH尾部的3个α微管蛋白抑制结构域，是该动力蛋白锚定至顶锥所必需的结构基础。尽管MyoA滑行体结构完整，但条件性敲除TgMyoH会严重损害寄生虫的运动、入侵以及从感染细胞中逸出的能力。我们进一步证明，MyoH对于将环状连接复合体从分泌细胞器胞吐作用发生的寄生虫顶端，转运至内膜复合物起始的顶端位置是必不可少的。本研究首次证实顶锥在运动与入侵过程中发挥直接功能，并确立了MyoH在沿该独特细胞器启动运动第一步中的不可或缺的作用，后续该过程将由MyoA接力完成，以实现有效的滑行运动与宿主细胞入侵。