Virtanen I, Badley R A, Paasivuo R, Lehto V P
J Cell Biol. 1984 Sep;99(3):1083-91. doi: 10.1083/jcb.99.3.1083.
Antibodies against different cytoskeletal proteins were used to study the cytoskeletal organization of human spermatozoa. A positive staining with actin antibodies was seen in both the acrosomal cap region and the principal piece region of the tail. However, no staining was obtained with nitrobenzoxadiazol-phallacidin, suggesting that most of the actin was in the nonpolymerized form. Most of the myosin immunoreactivity was confirmed to a narrow band in the neck region of spermatozoa. Tubulin was located to the entire tail, whereas vimentin was only seen in a discrete band-like structure encircling the sperm head, apparently coinciding with the equatorial segment region. Surface staining of the spermatozoa with fluorochrome-coupled Helix pomatia agglutinin revealed a similar band-like structure that co-distributed with the vimentin-specific staining. Instead, other lectin conjugates used labeled either the acrosomal cap region (peanut and soybean agglutinins), both the acrosomal cap and the postacrosomal region of the head (concanavalin A), or the whole sperm cell surface membrane (wheat germ and lens culinaris agglutinins and ricinus communis agglutinin l). In lectin blotting experiments, the Helix pomatia agglutinin-binding was assigned to a 80,000-mol-wt polypeptide which, together with vimentin, also resisted treatment with Triton X-100. Only the acrosomal cap and the principal piece of the tail were decorated with rabbit and hydridoma antibodies against an immunoanalogue of erythrocyte alpha-spectrin (p230). p230 appeared to be the major calmodulin-binding polypeptide in spermatozoa, as shown by a direct overlay assay of electrophoretic blots of spermatozoa with 125I-calmodulin. The results indicate that spermatozoa have a highly specialized cytoskeletal organization and that the distribution of actin, spectrin, and vimentin can be correlated with distinct surface specializations of the sperm cells. This suggest that cytoskeleton may regulate the maintenance of these surface assemblies and, hence, affect the spermatozoan function.
利用针对不同细胞骨架蛋白的抗体来研究人类精子的细胞骨架组织。在顶体帽区域和尾部的主段区域均可见肌动蛋白抗体的阳性染色。然而，用硝基苯并恶二唑 - 鬼笔环肽未获得染色，这表明大多数肌动蛋白处于非聚合形式。大多数肌球蛋白免疫反应性在精子颈部区域被确认为一条窄带。微管蛋白分布于整个尾部，而波形蛋白仅见于围绕精子头部的离散带状结构中，显然与赤道段区域重合。用荧光染料偶联的苹果蜗牛凝集素对精子进行表面染色，显示出与波形蛋白特异性染色共分布的类似带状结构。相反，使用的其他凝集素缀合物标记的区域分别为：顶体帽区域（花生和大豆凝集素）、头部的顶体帽和顶体后区域（伴刀豆球蛋白A）或整个精子细胞膜表面（麦芽和菜豆凝集素以及蓖麻凝集素1）。在凝集素印迹实验中，苹果蜗牛凝集素结合物被确定为一种80,000道尔顿分子量的多肽，该多肽与波形蛋白一起也能抵抗Triton X - 100处理。只有顶体帽和尾部的主段被抗红细胞α - 血影蛋白免疫类似物（p230）的兔抗体和杂交瘤抗体所标记。如用125I - 钙调蛋白对精子电泳印迹进行直接覆盖分析所示，p230似乎是精子中主要的钙调蛋白结合多肽。结果表明，精子具有高度特化的细胞骨架组织，肌动蛋白、血影蛋白和波形蛋白的分布可与精子细胞不同的表面特化相关。这表明细胞骨架可能调节这些表面组件的维持，从而影响精子功能。
