Plasma membrane-Actin filament Association through ERM Family Members

位于质膜下方的ERM家族成员Ezrin、Radioxin和Moesin被认为参与了肌动蛋白细丝/质膜的结合。为了鉴定与ERM家族成员直接相关的完整膜蛋白，我们用抗moesin单抗和代谢标记蛋氨酸或生物素标记的培养的BHK细胞进行了免疫沉淀研究。结果表明，Moesin与140kD的完整膜蛋白直接相关。以BHK细胞为抗原，获得一株能识别140kD膜蛋白的单抗。接下来，我们克隆了编码140kD膜蛋白的cDNA，并将其鉴定为CD44，这是一种广泛分布的细胞表面糖蛋白。各种抗CD44mAb的免疫沉淀表明，不仅在BHK细胞中，而且在小鼠L成纤维细胞中，Ezrin、Radixin和moesin都与CD44结合。此外，免疫荧光显微镜显示，在BHK和L细胞中，Triton X-100不溶性CD44与ERM家族成员精确共存。我们的结论是，ERM家族成员作为CD44胞浆结构域和基于肌动蛋白的细胞骨架之间的分子连接物。

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 immunoPrecipitaion studies using anti-moesin mAb and cultured BHK cells metabol-ically-labeled with [^<35>S] methionine or surface-labeled with biotin. The results indicated that moesin is directly associated with a 140kD integral membrane protein. Using BHK cells as antigens, we obtained a mAb which recognized the 140kD membrane protein. We next cloned a cDNA encoding the 140kD 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.