siRNA-induced TRAF6 knockdown promotes the apoptosis and inhibits the invasion of human lung cancer SPC-A1 cells.
siRNA-induced TRAF6 knockdown promotes the apoptosis and inhibits the invasion of human lung cancer SPC-A1 cells.
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DOI：
10.3892/or.2016.4602
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发表时间：
2016-04
期刊：
影响因子：
4.2
通讯作者：
Ye Y
中科院分区：
文献类型：
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作者：
He Z;Huang C;Lin G;Ye Y
肿瘤坏死因子受体相关因子6（TRAF 6）已被发现参与多种癌症。然而，小干扰RNA（siRNA）诱导的TRAF 6敲低对癌细胞生物学行为的影响仍然未知。因此，本研究旨在探讨siRNA诱导TRAF 6基因敲低对人肺癌SPC-A1细胞生物学行为的影响。使用定量RT-PCR（qRT-PCR）和蛋白质印迹法在转录和翻译水平测定TRAF 6在人肺腺癌A549、非小细胞肺癌H1650、人气道上皮Calu-3和人肺癌SPC-A1细胞系中的表达。采用siRNA技术敲低SPC-A1细胞TRAF 6的表达，采用电泳迁移率变动分析（EMSA）、细胞增殖实验、流式细胞术、Transwell侵袭实验和划痕实验检测TRAF 6对SPC-A1细胞NF-κB活性、细胞增殖、凋亡、细胞周期、侵袭和迁移的影响。Western blotting检测CD 24、CXCR 4、MMP 1、MMP 2、MMP 9、TWIST、TIMP-2和Slug蛋白表达。Western blotting和qRT-PCR分析显示，在Calu-3和SPC-A1细胞中，TRAF 6在翻译和转录水平上均上调，并且在SPC-A1细胞中检测到TRAF 6的K63连接的泛素化和组成性NF-κB激活。TRAF 6基因的敲除可抑制SPC-A1细胞的迁移和侵袭，促进细胞凋亡，但对细胞增殖和细胞周期无明显影响。此外，siRNA诱导的TRAF 6敲低导致CD 24和CXCR 4蛋白表达显著降低，但对MMP-1、MMP-2、MMP-9、Twist、TIMP-2或Slug表达几乎没有影响。目前的研究表明，TRAF 6在人肺癌细胞中上调，siRNA诱导的TRAF 6敲低抑制肺癌细胞的侵袭并促进凋亡。提示TRAF 6有可能成为肺癌治疗的一个新靶点。
Tumor necrosis factor receptor-associated factor 6 (TRAF6) has been found to be involved in multiple cancers. However, the effect of small interfering RNA (siRNA)-induced knockdown of TRAF6 on the biological behaviors of cancer cells remains unknown. Thus, the present study aimed to investigate the effect of siRNA-induced knockdown of TRAF6 on the biological behaviors of human lung cancer SPC-A1 cells. The expression of TRAF6 was determined in human lung adenocarcinoma A549, non-small cell lung cancer H1650, human airway epithelial Calu-3 and human lung cancer SPC-A1 cell lines using quantitative RT-PCR (qRT-PCR) and western blotting at the transcriptional and translational levels. TRAF6 expression was knocked down in the SPC-A1 cells using an siRNA technique, and the effects of TRAF6 knockdown on NF-κB activity, cell proliferation, apoptosis, cell cycle, invasion and migration of the SPC-A1 cells were determined using electrophoretic mobility shift assay (EMSA), cell proliferation assay, flow cytometry, Transwell invasion assay and scratch wound assay. In addition, the protein expression of CD24, CXCR4, MMP1, MMP2, MMP9, TWIST, TIMP-2 and Slug was quantified using western blotting assay. Western blotting and qRT-PCR assays showed upregulation of TRAF6 at both the translational and transcriptional levels in the Calu-3 and SPC-A1 cells, and K63-linked ubiquitination of TRAF6 and constitutive NF-κB activation were detected in the SPC-A1 cells. Knockdown of TRAF6 inhibited the migration and invasion and promoted the apoptosis of the SPC-A1 cells, but had little effect on cell proliferation and the cell cycle. In addition, siRNA-induced TRAF6 knockdown caused a marked reduction in the protein expression of CD24 and CXCR4, but had little effect on MMP-1, MMP-2, MMP-9, Twist, TIMP-2 or Slug expression. The present study demonstrated that TRAF6 is upregulated in human lung cancer cells, and siRNA-induced TRAF6 knockdown inhibits the invasion of lung cancer cells and promotes apoptosis. It is suggested that TRAF6 may be a promising target for the therapy of lung cancer.
DOI：
10.3779/j.issn.1009-3419.2015.11.01
发表时间：
2015-11
期刊：
Zhongguo fei ai za zhi = Chinese journal of lung cancer
影响因子：
--
作者：
Lin G;Huang C;Su G;Hu H;Xu H;Huang C
通讯作者：
Huang C