Atlas of Plasmodium falciparum intraerythrocytic development using expansion microscopy.

Apicomplexan parasites exhibit tremendous diversity in much of their fundamental cell biology, but study of these organisms using light microscopy is often hindered by their small size. Ultrastructural expansion microscopy (U-ExM) is a microscopy preparation method that physically expands the sample ~4.5x. Here, we apply U-ExM to the human malaria parasite Plasmodium falciparum during the asexual blood stage of its lifecycle to understand how this parasite is organized in three-dimensions. Using a combination of dye-conjugated reagents and immunostaining, we have catalogued 13 different P. falciparum structures or organelles across the intraerythrocytic development of this parasite and made multiple observations about fundamental parasite cell biology. We describe that the outer centriolar plaque and its associated proteins anchor the nucleus to the parasite plasma membrane during mitosis. Furthermore, the rhoptries, Golgi, basal complex, and inner membrane complex, which form around this anchoring site while nuclei are still dividing, are concurrently segregated and maintain an association to the outer centriolar plaque until the start of segmentation. We also show that the mitochondrion and apicoplast undergo sequential fission events while maintaining an association with the outer centriolar plaque during cytokinesis. Collectively, this study represents the most detailed ultrastructural analysis of P. falciparum during its intraerythrocytic development to date, and sheds light on multiple poorly understood aspects of its organelle biogenesis and fundamental cell biology.

顶复门寄生虫在其大部分基础细胞生物学方面表现出极大的多样性，但使用光学显微镜对这些生物进行研究常常因它们体积小而受阻。超微结构膨胀显微镜技术（U - ExM）是一种能将样本物理膨胀约4.5倍的显微镜样本制备方法。在此，我们在恶性疟原虫生命周期的无性血液阶段将U - ExM应用于该寄生虫，以了解这种寄生虫是如何在三维空间中组织的。通过结合使用染料偶联试剂和免疫染色，我们对这种寄生虫在红细胞内发育过程中的13种不同的恶性疟原虫结构或细胞器进行了分类，并对寄生虫基础细胞生物学有了多项观察结果。我们描述了在有丝分裂期间，外中心粒斑块及其相关蛋白质将细胞核锚定在寄生虫质膜上。此外，在细胞核仍在分裂时，围绕这个锚定位点形成的棒状体、高尔基体、基底复合体和内膜复合体同时被分离，并保持与外中心粒斑块的关联，直到分裂开始。我们还表明，在胞质分裂期间，线粒体和类囊体在保持与外中心粒斑块的关联的同时经历连续的分裂事件。总体而言，这项研究是迄今为止对恶性疟原虫在红细胞内发育过程中最详细的超微结构分析，并阐明了其细胞器生物发生和基础细胞生物学中多个鲜为人知的方面。