Quick-freeze, deep-etch visualization of the cytoskeleton beneath surface differentiations of intestinal epithelial cells.
Quick-freeze, deep-etch visualization of the cytoskeleton beneath surface differentiations of intestinal epithelial cells.
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DOI：
10.1083/jcb.91.2.399
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发表时间：
1981-11
期刊：
影响因子：
--
通讯作者：
Heuser JE
中科院分区：
文献类型：
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作者：
Hirokawa N;Heuser JE
支持肠上皮细胞中微绒毛的细胞骨架通过快速冷冻、深蚀刻、旋转复制技术可视化（Heuser和Salpeter. 1979. J. Cell Biol.82：150）。在快速冷冻之前，细胞被暴露在洗涤剂中，或者被物理地打开，以清除细胞质中的颗粒物质，否则这些颗粒物质会模糊视野。在这样的提取之后，细胞在其内部仍然显示出细胞骨架丝的特征性组织。这些细胞骨架的铂复制品具有足够的分辨率，使我们能够识别存在的细丝类型，并确定其相互作用的特征模式。最重要的新发现是，这些细胞中的顶端“末端网”（通过其核心肌动蛋白丝束支撑微绒毛）本身并不含有很多肌动蛋白，而是主要由将相邻的肌动蛋白束相互连接以及与中间丝的底层连接的窄股组成。这些链比肌动蛋白稍细，不显示肌动蛋白的53 A周期性，也不装饰肌球蛋白亚片段S1。相反，有两条证据表明，这些链可能包括肌球蛋白分子。首先，其他研究者已经表明肌球蛋白存在于终末网中（Mooseker et al. 1978. 79：444-453），然而我们在该区域中没有发现粗丝。第二，我们发现，在用肌球蛋白亚片段S1装饰核心丝束的过程中，这些股被完全去除，这表明它们已被外源性肌球蛋白竞争性取代。我们的结论是，肌球蛋白可能在这些细胞中发挥结构性作用，通过其交联分布，除了它在微绒毛运动中发挥的作用。
The cytoskeleton that supports microvilli in intestinal epithelial cells was visualized by the quick-freeze, deep-etch, rotary-replication technique (Heuser and Salpeter. 1979. J. Cell Biol. 82: 150). Before quick freezing, cells were exposed to detergents or broken open physically to clear away the granular material in their cytoplasm that would otherwise obscure the view. After such extraction, cells still displayed a characteristic organization of cytoskeletal filaments in their interiors. Platinum replicas of these cytoskeletons had sufficient resolution to allow us to identify the filament types present, and to determine their characteristic patterns of interaction. The most important new finding was that the apical "terminal web" in these cells, which supports the microvilli via their core bundles of actin filaments, does not itself contain very much actin but instead is comprised largely of narrow strands that interconnect adjacent actin bundles with one another and with the underlying base of intermediate filaments. These strands are slightly thinner than actin, do not display actin's 53A periodicity, and do not decorate with myosin subfragment S1. On the contrary, two lines of evidence suggested that these strands, could include myosin molecules. First, other investigators have shown that myosin is present in the terminal web (Mooseker et al. 1978. J. Cell Biol. 79: 444-453), yet we could find no thick filaments in this area. Second, we found that the strands were removed completely in the process of decorating the core filament bundles with the myosin subfragment S1, suggesting that they had been competitively displaced by exogenous myosin. We conclude that myosin may play a structural role in these cells, via its cross-linking distribution, in addition to whatever role it plays in microvillar motility.