Li Qiong, Wu Mingfu, Wang Hui, Xu Gang, Zhu Tao, Zhang Yongtao, Liu Ping, Song Anping, Gang Chen, Han Zhiqiang, Zhou Jianfeng, Meng Li, Lu Yunpin, Wang Shixuan, Ma Ding
Cancer Biology Research Center, Tongji Hospital, Huazhong University of Science and Technology, 1095 Jiefang Anv, Wuhan, Hubei 430030, PR China.
Cancer Lett. 2008 Mar 8;261(1):55-63. doi: 10.1016/j.canlet.2007.11.018. Epub 2007 Dec 26.
Ezrin primarily acts as a linker between the plasma membrane and the cytoskeleton and is a key component in tumor metastasis. In the present study, RNA interference (RNAi) using ezrin small hairpin RNAs (ezrin shRNAs) was used to define the roles of ezrin in the regulation of malignant behaviors of human breast cancer. The highly metastatic human breast cancer cell MDA-MB-231, in which ezrin mRNA and protein levels are the highest, was selected as a cell model in vitro. In addition, we also found that ezrin expression was up-regulated and its immuno-staining trans-located from cell membrane to cytoplasm, whereas E-cadherin expression decreased and showed the same cell distribution as ezrin in lymphatic metastases of human breast carcinomas. After repression of ezrin by more than 85% of G3PDH and 75% of beta-actin in mRNA and protein levels was maintained in the stable expressing ezrin shRNAs MDA-MB-231 cell clones, the abilities of cell motility and invasiveness were obviously inhibited with a 4-fold and 2-fold, respectively, and the altered cell polarity was observed. Western blot analyses further revealed that the silencing of ezrin induced an increased E-cadherin expression and a decreased phosphorylation of beta-catenin by inhibiting phosphorylation levels of c-src. These data indicate that ezrin overexpression positively correlated with metastatic potentials of human breast cancer cells, especially lymphatic system metastasis. Decreased ezrin expression by shRNA reversed metastatic behaviors of human breast cancer cells by inducing c-src-mediated E-cadherin expression, suggesting that ezrin may have potential values in assessing lymphatic metastasis of human breast cancers.
埃兹蛋白主要作为质膜与细胞骨架之间的连接蛋白，是肿瘤转移的关键组成部分。在本研究中，使用埃兹蛋白小发夹RNA（ezrin shRNAs）进行RNA干扰（RNAi），以确定埃兹蛋白在调控人乳腺癌恶性行为中的作用。选择埃兹蛋白mRNA和蛋白水平最高的高转移性人乳腺癌细胞MDA-MB-231作为体外细胞模型。此外，我们还发现，在人乳腺癌的淋巴转移中，埃兹蛋白表达上调，其免疫染色从细胞膜转移至细胞质，而E-钙黏蛋白表达下降，且与埃兹蛋白呈现相同的细胞分布。在稳定表达ezrin shRNAs的MDA-MB-231细胞克隆中，埃兹蛋白在mRNA和蛋白水平上被抑制超过85%的甘油醛-3-磷酸脱氢酶（G3PDH）和75%的β-肌动蛋白后，细胞运动和侵袭能力分别被显著抑制了4倍和2倍，并且观察到细胞极性发生改变。蛋白质印迹分析进一步表明，埃兹蛋白的沉默通过抑制c-src的磷酸化水平，诱导E-钙黏蛋白表达增加和β-连环蛋白磷酸化减少。这些数据表明，埃兹蛋白的过表达与人乳腺癌细胞的转移潜能呈正相关，尤其是与淋巴系统转移相关。通过shRNA降低埃兹蛋白表达可通过诱导c-src介导的E-钙黏蛋白表达来逆转人乳腺癌细胞的转移行为，这表明埃兹蛋白在评估人乳腺癌淋巴转移方面可能具有潜在价值。
