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Plasma membrane-Actin filament Association through ERM Family Members

通过 ERM 家族成员建立质膜-肌动蛋白丝协会

基本信息

批准号：
05680627
负责人：
TSUKITA Sachiko
金额：
$ 1.47万
依托单位：
KYOTO UNIVERSITY (1994)Okazaki National Research Institutes (1993)
依托单位国家：
日本
项目类别：
Grant-in-Aid for General Scientific Research (C)
财政年份：
1993
资助国家：
日本
起止时间：
1993 至 1994
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-05680627/
关键词：
CD44 actin filaments ERM radixin ezrin moesin plasma membrane アクチンモノクローナル抗体細胞膜

项目摘要

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.

ERM家族成员ezrin、radixin和膜突蛋白（moesin）位于质膜下方，被认为参与肌动蛋白丝/质膜缔合。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和培养的BHK细胞进行免疫组织化学研究，所述细胞用[^<35>S]甲硫氨酸代谢标记或用生物素表面标记。结果表明，膜突蛋白与一个分子量为140 kD的膜整合蛋白直接结合。以BHK细胞为抗原，获得了识别140 kD膜蛋白的单克隆抗体。我们随后克隆了编码140 kD膜蛋白的cDNA，并将其鉴定为CD 44，一种广泛分布的细胞表面糖蛋白。免疫沉淀与各种抗CD 44单克隆抗体表明，ezrin和radixin以及膜突蛋白与CD 44不仅在BHK细胞，而且在小鼠L成纤维细胞。此外，免疫荧光显微镜显示，在BHK和L细胞中，Triton X-100不溶性CD 44与ERM家族成员精确共定位。我们的结论是，ERM家族成员作为分子之间的CD 44和肌动蛋白为基础的细胞骨架的胞质结构域的接头。