We will probe the function of two potential formin-like actin nucleation factors in malaria parasite adhesion and motility using recently developed quantitative imaging tools

肌动蛋白的聚合和寄生虫的运动性在疟原虫的整个生命周期中是必不可少的。我们主要研究伯氏疟原虫子孢子，这是一种啮齿类疟疾寄生虫，由受感染的蚊子传播给哺乳动物宿主。在蚊子叮咬期间，子孢子被注射到皮肤中，其中它们迁移以找到血管。进入血流后，子孢子继续它们的旅程到肝脏，在那里它们积极地侵入肝细胞以分化。迁移由肌动蛋白-肌球蛋白马达驱动。奇怪的是，寄生虫中聚合的肌动蛋白丝非常短，并且对肌动蛋白丝在哪里形成以及哪些分子对肌动蛋白成核至关重要知之甚少。重要的是，几种疟疾寄生虫的测序基因组仅编码非常有限数量的肌动蛋白结合蛋白，其中根据物种的不同，有两到三个含有同源结构域的肌动蛋白样蛋白。我们最近的研究表明，肌动蛋白聚合不仅是重要的子孢子运动，但也为子孢子粘附过程中的不同步骤。这表明形成了不同的空间分离的肌动蛋白丝组。我们现在想解剖潜在的肌动蛋白成核因子的作用，通过表达的功能结构域作为荧光标记的融合蛋白从阶段特异性强和弱启动子定位在寄生虫和干扰内源性蛋白质的功能。我们将测试野生型和突变体的同源结构域的过表达对寄生虫粘附，运动和宿主细胞侵袭的影响，我们最近适应于子孢子的研究或在实验室新开发的一些工具。

The polymerization of actin and thus parasite motility is essential throughout the life cycle of the malaria parasite. We are mainly working on Plasmodium berghei sporozoites, the forms of a rodent malaria parasite transmitted by an infected mosquito to the mammalian host. During a mosquito bite sporozoites are injected into the skin wherein they migrate to find a blood vessel. After entering the blood stream, sporozoites continue their journey to the liver, where they actively invade liver cells to differentiate. Migration is driven by an actin-myosin motor. Curiously, polymerized actin filaments in parasites are very short and little is known about where actin filaments are formed and which molecules are essential for actin nucleation. Importantly, the sequenced genomes of several malaria parasites encode only a very limited number of actin binding proteins of which there are, depending on species, two to three formin like proteins containing formin homology domains. We have recently shown that actin polymerization is not just important for sporozoite motility, but also for distinct steps during sporozoite adhesion. This suggests that different spatially separated sets of actin filaments are formed. We now want to dissect the roles of the potential actin nucleating factors by expressing the functional domains as fluorescently labeled fusion proteins from stage-specific strong and weak promoters to localize them in the parasite and to interfere with the function of the endogenous proteins. We will test the effect of over-expression of wild type and mutant formin homology domains on parasite adhesion, motility and host cell invasion with a number of tools we recently adapted to the study of sporozoites or developed newly in the lab.