Knockdown of stromal interaction molecule 1 (STIM1) suppresses store-operated calcium entry, cell proliferation and tumorigenicity in human epidermoid carcinoma A431 cells.
Knockdown of stromal interaction molecule 1 (STIM1) suppresses store-operated calcium entry, cell proliferation and tumorigenicity in human epidermoid carcinoma A431 cells.
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DOI：
10.1016/j.bcp.2012.09.021
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发表时间：
2012-12
影响因子：
5.8
通讯作者：
J. Yoshida;Kuniyoshi Iwabuchi;T. Matsui;T. Ishibashi;T. Masuoka;M. Nishio
中科院分区：
文献类型：
--
作者：
J. Yoshida;Kuniyoshi Iwabuchi;T. Matsui;T. Ishibashi;T. Masuoka;M. Nishio
钙库操纵的钙离子内流（SOCE）对大多数非兴奋细胞的基因转录、细胞周期进程和增殖等细胞活动非常重要。基质相互作用分子1（Stromal interaction molecule 1，STIM 1）是一种新发现的钙敏感蛋白，可监测内质网（endoplasmic reticulum，ER）钙库的耗竭，并激活质膜上钙库操纵的钙通道，诱导SOCE。为了研究STIM 1在肿瘤生长中与SOCE相关的可能作用，我们利用RNA干扰技术建立了人表皮样癌A431细胞STIM 1敲低（KD）克隆。与阴性对照克隆相比，两个STIM 1 KD克隆中的毒胡萝卜素（ER Ca 2 +-ATP酶抑制剂）诱导的SOCE和磷脂酶C偶联受体激动剂诱导的SOCE减少。KD-抗性全长STIM 1的重新表达，但不是Ca 2+释放激活的Ca 2+通道激活结构域（CAD）缺失的STIM 1突变体，在KD克隆恢复了SOCE的幅度，表明STIM 1敲低的特异性。STIM 1 KD克隆的细胞生长慢于阴性对照克隆。DNA合成评估BrdU掺入，以及EGF刺激的EGF受体活化，减少在STIM 1 KD克隆。与阴性对照相比，STIM 1 KD克隆的异种移植物生长显著延迟。在STIM 1 KD克隆中细胞迁移减弱，并且STIM 1沉默效应通过全长STIM 1而不是CAD缺失突变体的瞬时再表达而逆转。这些结果表明，STIM 1在人表皮样癌A431细胞的SOCE、细胞生长和致瘤性中起重要作用，提示STIM 1靶向药物在表皮样癌治疗中的潜在应用。
Store-operated calcium (Ca2+) entry (SOCE) is important for cellular activities such as gene transcription, cell cycle progression and proliferation in most non-excitable cells. Stromal interaction molecule 1 (STIM1), a newly identified Ca2+-sensing protein, monitors the depletion of endoplasmic reticulum (ER) Ca2+stores and activates store-operated Ca2+channels at the plasma membrane to induce SOCE. To investigate the possible roles of STIM1 in tumor growth in relation to SOCE, we established STIM1 knockdown (KD) clones of human epidermoid carcinoma A431 cells by RNA interference. Thapsigargin, an inhibitor of ER Ca2+-ATPase, -induced and phospholipase C-coupled receptor agonist-induced SOCEs were reduced in two STIM1 KD clones compared to a negative control clone. Re-expression of a KD-resistant full-length STIM1, but not a Ca2+release-activated Ca2+channel activation domain (CAD)-deleted STIM1 mutant, in the KD clone restored the amplitude of SOCE, suggesting the specificity of the STIM1 knockdown. The cell growth of the STIM1 KD clones was slower than that of the negative control clone. DNA synthesis assessed by BrdU incorporation, as well as EGF-stimulated EGF receptor activation, decreased in the STIM1 KD clones. Xenograft growth of the STIM1 KD clones was significantly retarded compared with that of the negative control. Cell migration was attenuated in the STIM1 KD clone and the STIM1 silencing effect was reversed by transient re-expression of the full-length STIM1 but not CAD-deletion mutant. These results indicate that STIM1 plays an important role in SOCE, cell-growth and tumorigenicity in human epidermoid carcinoma A431cells, suggesting the potential use of STIM1-targeting agents for treating epidermoid carcinoma.