Hair Molly, Yanase Ryuji, Moreira-Leite Flávia, Wheeler Richard John, Sádlová Jovana, Volf Petr, Vaughan Sue, Sunter Jack Daniel
Department of Biological and Medical Sciences, Oxford Brookes University, Oxford, United Kingdom.
Peter Medawar Building for Pathogen Research, University of Oxford, Oxford, United Kingdom.
PLoS Pathog. 2024 Feb 28;20(2):e1012054. doi: 10.1371/journal.ppat.1012054. eCollection 2024 Feb.
The unicellular parasite Leishmania has a precisely defined cell architecture that is inherited by each subsequent generation, requiring a highly coordinated pattern of duplication and segregation of organelles and cytoskeletal structures. A framework of nuclear division and morphological changes is known from light microscopy, yet this has limited resolution and the intrinsic organisation of organelles within the cell body and their manner of duplication and inheritance is unknown. Using volume electron microscopy approaches, we have produced three-dimensional reconstructions of different promastigote cell cycle stages to give a spatial and quantitative overview of organelle positioning, division and inheritance. The first morphological indications seen in our dataset that a new cell cycle had begun were the assembly of a new flagellum, the duplication of the contractile vacuole and the increase in volume of the nucleus and kinetoplast. We showed that the progression of the cytokinesis furrow created a specific pattern of membrane indentations, while our analysis of sub-pellicular microtubule organisation indicated that there is likely a preferred site of new microtubule insertion. The daughter cells retained these indentations in their cell body for a period post-abscission. By comparing cultured and sand fly derived promastigotes, we found an increase in the number and overall volume of lipid droplets in the promastigotes from the sand fly, reflecting a change in their metabolism to ensure transmissibility to the mammalian host. Our insights into the cell cycle mechanics of Leishmania will support future molecular cell biology analyses of these parasites.
单细胞寄生虫利什曼原虫具有精确界定的细胞结构，这种结构由每一代后续细胞继承，这需要细胞器和细胞骨架结构进行高度协调的复制和分离模式。通过光学显微镜已知核分裂和形态变化的框架，但其分辨率有限，细胞体内细胞器的内在组织及其复制和遗传方式尚不清楚。利用体电子显微镜方法，我们对不同前鞭毛体的细胞周期阶段进行了三维重建，以全面、定量地了解细胞器的定位、分裂和遗传情况。我们的数据集中显示新细胞周期开始的首个形态学迹象是新鞭毛的组装、收缩泡的复制以及细胞核和动基体体积的增加。我们发现胞质分裂沟的进展产生了特定的膜凹陷模式，而我们对表膜下微管组织的分析表明，可能存在新微管插入的优先位点。子细胞在脱离后一段时间内，其细胞体中仍保留这些凹陷。通过比较培养的前鞭毛体和采自沙蝇的前鞭毛体，我们发现采自沙蝇的前鞭毛体中脂滴的数量和总体积有所增加，这反映了它们新陈代谢的变化，以确保能够传播到哺乳动物宿主。我们对利什曼原虫细胞周期机制的深入了解将为今后对这些寄生虫的分子细胞生物学分析提供支持。
