lncRNA CCAT2调控SIRT1蛋白表达激活Wnt/β-catenin通路影响非小细胞肺癌细胞增殖和转移

lncRNA CCAT2 Affects Proliferation and Metastasis of NSCLC Cells by Activating Wnt/β-catenin Pathway via Regulating SIRT1

该文旨在分析长链非编码RNA(lncRNA) CCAT2通过调控SIRT1蛋白的表达激活Wnt/β-catenin信号通路进而影响非小细胞肺癌细胞增殖和转移的机制。采用qRT-PCR检测肺癌组织及肺癌细胞株中lncRNA CCAT2的表达。利用卡方检验分析lncRNA CCAT2表达与肺癌患者临床病理特征的关系。CCK-8实验、划痕实验及Transwell实验观察敲低lncRNA CCAT2表达对肺癌细胞增殖、迁移及浸润能力的影响。Western blot检测敲低lncRNA CCAT2表达对H1975细胞株中SIRT1蛋白及Wnt/β-catenin信号通路蛋白表达的影响;以及敲低或过表达SIRT1对Wnt/β-catenin信号通路蛋白表达的影响。利用RNA免疫共沉淀(RIP)及RNA pull-down实验验证lncRNA CCAT2与SIRT1之间的相互作用。该研究得出,癌组织及肺癌细胞株中lncRNA CCAT2表达显著较高(P<0.05),敲低lncRNA CCAT2表达能够抑制H1975细胞的增殖、迁移及浸润。敲低lncRNA CCAT2表达后,H1975细胞株中SIRT1、β-catenin、Cyclin D1、myc蛋白的表达降低;敲低SIRT1后,细胞核β-catenin蛋白、Cyclin D1、myc蛋白的表达均降低。与非特异性抗体比较,SIRT1抗体呈明显的lncRNA CCAT2富集;lncRNA CCAT2的截短突变组细胞中SIRT1相对表达量明显低于lncRNA CCAT2组。lncRNA CCAT2通过调控SIRT1蛋白表达激活Wnt/β-catenin信号通路进而促进肺癌细胞增殖、迁移及浸润,有望成为新的肿瘤标志物。

lncRNA CCAT2 affects the proliferation and metastasis of NSCLC cells by regulating the expression of SIRT1 protein and activating Wnt/β-catenin signaling pathway.In this study,the expression of lncRNA CCAT2 in lung cancer tissues and lung cancer cell lines was detected by qRT-PCR.The relationship between lncRNA CCAT2 expression and clinicopathological features of lung cancer was analyzed by chi-square test.CCK-8 assay,scratch test and Transwell assay were used to observe the effect of lncRNA CCAT2 knockdown on the proliferation,migration and infiltration of lung cancer cells.After knocking down the expression of lncRNA CCAT2 expression in H1975 cell line,Western blot was used to detect the expression change of SIRT1 protein and Wnt/β-catenin signaling pathway.The effect of knockdown or overexpression of SIRT1 on Wnt/β-catenin signaling pathway were also detected.RNA immunoprecipitation and RNA pull-down experiments were used to verify the interaction between lncRNA CCAT2 and SIRT1.The results showed that the expression of lncRNA CCAT2 was significantly higher in cancer tissues and lung cancer cell lines (P<0.05).lncRNA CCAT2 knockdown inhibited the proliferation,migration and infiltration of H1975 cells.The expression of SIRT1,β-catenin,Cyclin D1,and myc proteins in H1975 cell line was decreased after the knockdown of lncRNA CCAT2.After knockdown of SIRT1,the expression of β-catenin,Cyclin D1 and myc proteins decreased.Compared with non-specific antibodies,SIRT1 antibodies were significantly enriched to lncRNA CCAT2.The expression of SIRT1 in truncated mutant cells of lncRNA CCAT2 was significantly lower than that of lncRNA CCAT2.lncRNA CCAT2 promotes lung cancer cell proliferation,migration and infiltration by activating Wnt/β-catenin signaling pathway via SIRT1,and lncRNA CCAT2 may become a new tumor marker.