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转录因子HOXA5在乳腺癌细胞中的下游信号通路
Identification of the downstream signaling pathways of HOXA5 in breast cancer cells
目的 探究转录因子HOXA5在乳腺癌进展中的作用。方法 构建过表达HOXA5的稳定转染BT549和SUM159细胞系, 在稳转BT549细胞系中利用Transwell检测细胞的转移能力；Western blotting检测上皮-间质转化（EMT）相关蛋白表达水平的变化；平板克隆实验检测细胞的增殖能力；转录组测序技术（RNA-seq）进一步分析HOXA5的调控基因，利用软件Cluster 3.0，Tree View和DAVID生物信息数据库（DAVID Bioinformatics Resources）以及Enrichr分析京都基因与基因组百科全书（KEGG）信号通路，绘制热图（heatmap）。 结果 BT549细胞系中，稳定转染HOXA5后的实验组细胞迁移能力显著降低(P<0.001)，且上皮标志物 E-cadherin蛋白表达水平显著上调，间质标志物 N-cadherin, Twist1、Slug蛋白表达水平显著下调；平板克隆结果表明，实验组形成的克隆数目明显减少(P<0.05)，克隆大小明显降低。 结论 HOXA5通过促进细胞由间质向上皮转化，抑制乳腺癌细胞迁移，并且抑制细胞增殖；HOXA5通过调节糖脂代谢途径调节癌细胞增殖，并且还通过调节细胞运动和黏附相关的基因抑制肿瘤细胞的迁移，通过肿瘤坏死因子（TNF）信号通路发挥抗肿瘤作用。总之，转录因子HOXA5对乳腺癌的发展和恶化起着显著的抑制作用。
Objective To investigate the role of transcriptional factor HOXA5 in breast cancer progression. Methods Stably transfected BT549 and SUM159 cell lines with overexpression of HOXA5 were established. Transwell assay, Western blotting and colony formation were applied to detect cell migration, the expression of epithelial-mesenchymal transition (EMT) markers and cell proliferation respectively. RNA-seq was further performed to investigate the target genes of HOXA5. Software Cluster 3.0, treeview, database DAVID Bioinformatics Resources and Enrichr were used to analyze the pathways in Kyoto Encyclopedia of Genes and Genomes (KEGG) and draw the heatmap. Results In BT549 cells, stable transfection of HOXA5 repressed cell migration(P<0.05), promoted expression of E-cadherin and repressed expression of N-cadherin, Twist1 and Slug. HOXA5 repressed colony formation(P<0.05) and cell proliferation as well. Conclusion Our results reveal that HOXA5 represses cell migration by promoting the transition from mesenchymal to epithelial. RNA-seq results show that HOXA5 may inhibit cancer cell proliferation by regulating glucose and lipid metabolism and may repress cell migration by targeting genes related with cell motility and cell adhension. In addition, HOXA5 may play an anticancer role by regulating tumor necrosis factor (TNF) pathway. Based on these data, we conclude that transcriptional factor HOXA5 represses breast cancer progression and may act as a breast cancer repressor.
HOXA5 / 乳腺癌 / 上皮间质转化 / Transwell / 转录组测序
HOXA5 / Breast cancer / Epithelial-mesenchymal transition / Transwell / RNA-Sequence
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