Wang S M, Chen J S, Fong T H, Hsu S Y, Lim S S
Department of Anatomy, College of Medicine, National Taiwan University, Taipei.
Cell Motil Cytoskeleton. 1997;36(3):216-27. doi: 10.1002/(SICI)1097-0169(1997)36:3<216::AID-CM2>3.0.CO;2-5.
We report the presence of a novel filament system in goldfish xanthophores using a monoclonal antibody (A2) made against 40-70 kD proteins derived from cytoskeletal preparations. On Western blots, this antibody recognized a 45 kD protein in xanthophore cell extracts. In cells with dispersed pigment, immunofluorescence staining of xanthophores revealed a uniform distribution of A2-reactive filaments. In cells with aggregated pigment, these filaments assumed a distinctively radial orientation, such that filaments emanated from the central pigment mass (CPM). At the electron microscopic level, immunogold labeling identified a filament system with a diameter of 7 nm. Overall, the cellular distribution of A2-reactive filaments was distinctly different from that of the other known components of the cytoskeleton, such as intermediate filaments, actin filaments, and microtubules. A2-reactive filaments also appeared resistant to agents known to perturb the cytoskeleton such as cytochalasin B, which depolymerized the actin filaments. When xanthophores were treated with vinblastine, shown to depolymerize microtubules and induce the collapse of intermediate filaments (vimentin and keratin) in other cell types, no effect on the A2 filament distribution was observed. On the other hand, treatment with calyculin A, a phosphatase inhibitor, converted A2 filaments into a wavy bundles, the effect of which was completely reversible by the removal of the drug from culture medium. These novel properties of A2 filaments, together with their reorganization in response to pigment translocation suggest that A2 filaments might play a yet unidentified role in intracellular organelle transport in these cells.
我们利用一种针对从细胞骨架制剂中提取的40 - 70 kD蛋白质制备的单克隆抗体（A2），报告了金鱼黄色素细胞中一种新型丝状系统的存在。在蛋白质免疫印迹分析中，该抗体识别出黄色素细胞提取物中的一种45 kD蛋白质。在色素分散的细胞中，对黄色素细胞进行免疫荧光染色显示A2反应性细丝呈均匀分布。在色素聚集的细胞中，这些细丝呈现出独特的放射状取向，即细丝从中央色素团（CPM）发出。在电子显微镜水平上，免疫金标记鉴定出一种直径为7 nm的丝状系统。总体而言，A2反应性细丝的细胞分布与细胞骨架的其他已知成分，如中间丝、肌动蛋白丝和微管，明显不同。A2反应性细丝似乎也对已知会扰乱细胞骨架的试剂具有抗性，例如细胞松弛素B，它会使肌动蛋白丝解聚。当用长春花碱处理黄色素细胞时，长春花碱在其他细胞类型中可使微管解聚并诱导中间丝（波形蛋白和角蛋白）塌陷，但未观察到对A2细丝分布有影响。另一方面，用磷酸酶抑制剂冈田酸处理会使A2细丝转变为波浪状束，从培养基中去除该药物后，这种效应完全可逆。A2细丝的这些新特性，以及它们随色素转运的重组，表明A2细丝可能在这些细胞的细胞内细胞器运输中发挥尚未明确的作用。
