Tsukita S, Oishi K, Sato N, Sagara J, Kawai A, Tsukita S
Department of Information Physiology, National Institute for Physiological Sciences, Aichi, Japan.
J Cell Biol. 1994 Jul;126(2):391-401. doi: 10.1083/jcb.126.2.391.
The ERM family members, ezrin, radixin, and moesin, localizing just beneath the plasma membranes, are thought to be involved in the actin filament/plasma membrane association. To identify the integral membrane protein directly associated with ERM family members, we performed immunoprecipitation studies using antimoesin mAb and cultured baby hamster kidney (BHK) cells metabolically labeled with [35S]methionine or surface-labeled with biotin. The results indicated that moesin is directly associated with a 140-kD integral membrane protein. Using BHK cells as antigens, we obtained a mAb that recognized the 140-kD membrane protein. We next cloned a cDNA encoding the 140-kD membrane protein and identified it as CD44, a broadly distributed cell surface glycoprotein. Immunoprecipitation with various anti-CD44 mAbs showed that ezrin and radixin, as well as moesin, are associated with CD44, not only in BHK cells, but also in mouse L fibroblasts. Furthermore, immunofluorescence microscopy revealed that in both BHK and L cells, the Triton X-100-insoluble CD44 is precisely colocalized with ERM family members. We concluded that ERM family members work as molecular linkers between the cytoplasmic domain of CD44 and actin-based cytoskeletons.
ERM家族成员埃兹蛋白、根蛋白和膜突蛋白定位于质膜下方，被认为参与肌动蛋白丝/质膜的结合。为了鉴定与ERM家族成员直接相关的整合膜蛋白，我们使用抗膜突蛋白单克隆抗体对用[35S]甲硫氨酸进行代谢标记或用生物素进行表面标记的培养的幼仓鼠肾（BHK）细胞进行了免疫沉淀研究。结果表明，膜突蛋白与一种140-kD的整合膜蛋白直接相关。以BHK细胞作为抗原，我们获得了一种识别140-kD膜蛋白的单克隆抗体。接下来，我们克隆了编码140-kD膜蛋白的cDNA，并将其鉴定为CD44，一种广泛分布的细胞表面糖蛋白。用各种抗CD44单克隆抗体进行免疫沉淀表明，不仅在BHK细胞中，而且在小鼠L成纤维细胞中，埃兹蛋白、根蛋白以及膜突蛋白都与CD44相关。此外，免疫荧光显微镜检查显示，在BHK和L细胞中，Triton X-100不溶性CD44与ERM家族成员精确共定位。我们得出结论，ERM家族成员作为CD44胞质结构域与基于肌动蛋白的细胞骨架之间的分子连接物发挥作用。
