Katsumoto T, Kurimura T
Laboratory of Electron Microscopy, Tottori University School of Medicine, Yonago, Japan.
Biol Cell. 1988;62(1):1-10.
Whole-mount cell preparations of cultured rat 3Y1 cells were examined by stereo electron microscopy to identify the ultrastructural localization of concanavalin A (Con A) receptors in the plasma membrane, and to clarify the relationship between Con A receptors and cytoskeletal components. Well spread monolayer cells were extracted with saponin, briefly fixed, and then partially broken open with shearing force to facilitate the introduction of antibodies for identification of actin filaments. Stereo electron microscopy of such treated cells revealed a 3-dimensional image of filamentous structures such as fine filaments, microtubules (MT) and endoplasmic reticulum (ER) in the flattened areas of each cell. Just beneath the plasma membrane were meshworks of actin-containing fine filaments, as identified by an immunogold staining method. Microtubules and ER were observed to be either directly or indirectly associated with this meshwork. The broken open part of each cell exhibited a meshwork of filaments which were associated with the cytoplasmic surface of the plasma membrane. Some of the filaments were connected to the plasma membrane either by their ends or by their lateral surfaces. The localization of Con A receptors was examined by binding colloidal gold-labelled Con A to the surface of fixed, saponin-extracted cells. Virtually all gold particles bound externally at the same membrane sites where intracellular actin filaments attached internally. The observations strongly suggest that the distribution of Con A receptors was regulated by the underlying meshwork of actin filaments.
通过立体电子显微镜检查培养的大鼠3Y1细胞的整装细胞制剂，以确定伴刀豆球蛋白A（Con A）受体在质膜中的超微结构定位，并阐明Con A受体与细胞骨架成分之间的关系。将铺展良好的单层细胞用皂素提取，短暂固定，然后用剪切力使其部分破裂，以便引入抗体来鉴定肌动蛋白丝。对经如此处理的细胞进行立体电子显微镜观察，揭示了每个细胞扁平区域中丝状结构（如细丝、微管（MT）和内质网（ER））的三维图像。通过免疫金染色法鉴定，在质膜下方是含肌动蛋白的细丝网络。观察到微管和内质网与该网络直接或间接相关。每个细胞的破裂部分呈现出与质膜细胞质表面相关的细丝网络。一些细丝通过其末端或侧面与质膜相连。通过将胶体金标记的Con A结合到固定的、经皂素提取的细胞表面来检查Con A受体的定位。实际上，所有金颗粒都在细胞内肌动蛋白丝附着的同一膜位点外部结合。这些观察结果强烈表明，Con A受体的分布受肌动蛋白丝潜在网络的调节。
