X-ray tomography of Schizosaccharomyces pombe
X-ray tomography of Schizosaccharomyces pombe
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DOI：
10.1111/j.1432-0436.2007.00180.x
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发表时间：
2007-07-01
期刊：
影响因子：
2.9
通讯作者：
Larabell, Carolyn A.
中科院分区：
文献类型：
--
作者：
Gu, Weiwei;Etkin, Laurence D.;Larabell, Carolyn A.
单细胞酵母裂殖酵母的遗传可塑性使其成为研究许多真核细胞过程，特别是细胞分裂的重要模式生物。在过去的几年里，人们在了解真核细胞分裂的调控机制和发生的细胞变化方面取得了很大的进展，例如，细胞动力学收缩环的形成。然而，一个完整的理解需要鉴定所涉及的蛋白质，并将这些信息与显示分子在细胞结构中的位置的图像相关联。电子显微镜分析揭示了细胞结构的精细超微结构图像，但这种技术通常需要大量的处理程序，这些程序是劳动密集型和耗时的。成像技术，可以更迅速地获得更好的分辨率比光学显微镜需要推进使用该模型系统进行精确的分子定位分析。在这份手稿中，我们研究了S。pombe使用软X射线断层扫描，成像技术，产生三维（3-D）图像的完整的水合细胞在优于50纳米的各向同性分辨率。这种技术在“水窗”中使用X射线，其中有机材料的吸收比水强大约一个数量级，产生细胞结构的高对比度图像。当细胞在没有任何化学固定剂、染色剂或对比增强试剂的情况下被检查时，图像反映了接近天然状态的细胞结构。我们对S.在细胞分裂前和分裂过程中的庞贝氏细胞，揭示了亚细胞器，肌动球蛋白环和分裂细胞的隔膜。这些图像揭示了胞质分裂过程的诱人细节，并且是我们生成断层图像组合的目标的第一步，该组合将标记分子的位置映射到细胞的高分辨率3-D重建中。
The genetic tractability of the unicellular yeast Schizosaccharomyces pombe has resulted in it becoming an important model organism for the study of many eukaryotic cellular processes, in particular cell division. Over the past few years much progress has been made toward understanding the mechanisms that regulate eukaryotic cell division and the cellular changes that occur-for example, the formation of the cytokinetic contractile ring. However, a full understanding requires both identification of the proteins involved and correlation of this information with images showing the location of molecules in context of the cell architecture. Electron microscopic analyses have revealed exquisite ultrastructural images of cell structure, but this technique typically requires extensive processing-procedures that are labor intensive and time consuming. Imaging techniques that can more rapidly obtain better resolution than light microscopy are needed to advance the use of this model system for precise molecular localization analyses. In this manuscript, we examined S. pombe using soft X-ray tomography, an imaging technique that generates three-dimensional (3-D) images of intact hydrated cells at better than 50 nm isotropic resolution. This technique uses X-rays in the "water window," where organic material absorbs approximately an order of magnitude more strongly than water, producing high-contrast images of cellular structures. As cells are examined in the absence of any chemical fixatives, stains, or contrast enhancement reagents, the images reflect cellular structures in the near-native state. We conducted preliminary soft X-ray imaging of S. Pombe cells before and during cell division that revealed subcellular organelles, the actomyosin ring, and the septum of dividing cells. These images reveal tantalizing details of the cytokinesis process and are the first steps in our goal of generating a portfolio of tomographic images that map the location of labeled molecules into high-resolution 3-D reconstructions of the cell.