TRIM59基因沉默对结直肠癌细胞增殖和细胞周期的影响

Effects of TRIM59 gene silencing on proliferation and cell cycle of colorectal cancer cells

目的探讨三结构域蛋白59(TRIM59)对人结直肠癌HCT116细胞生物学功能的影响及其作用机制。方法构建重组TRIM59干扰质粒RNA(si-TRIM59),采用脂质体转染法将TRIM59敲减质粒转染至结直肠癌细胞HCT116中,RT-qPCR和Western blot验证转染效率;CCK-8法、克隆形成实验、流式细胞术检测TRIM59基因沉默对结直肠癌细胞增殖、克隆形成及细胞周期的影响,Western blot检测CDK4和cyclinD1细胞周期蛋白表达水平。结果TRIM59 mRNA及蛋白在结直肠癌细胞中的表达水平高于正常结直肠黏膜细胞(P<0.05);敲减TRIM59表达后,HCT116细胞的增殖活性和克隆形成能力受到抑制(P<0.05),细胞周期阻滞在G_(0)/G_(1)期;同时,细胞周期相关蛋白CDK4和cyclinD1的表达量降低(P<0.05)。结论TRIM59在结直肠癌细胞中高表达,下调TRIM59表达可抑制结直肠癌细胞的增殖、克隆形成能力,提示TRIM59有望成为结直肠癌基因治疗的新靶点。

Objective To investigate the effect of tripartite motif containing 59(TRIM59)on the biological function of human colorectal cancer HCT116 cells and its possible mechanism.Methods The recombinant TRIM59 interference plasmid(si-TRIM59)was constructed and transfected into colorectal cancer cell line HCT116 by liposome transfection.The transfection efficiency was verified by RT-qPCR and Western blot.The effects of TRIM59 gene silencing on the proliferation,colony-formation ability and cell cycle of colorectal cancer cells were detected by CCK-8 method,colony formation assays and flow cytometry respectively.Western blot was used to detect the expression level of CDK4 and cyclinD1.Results The expression levels of TRIM59 mRNA and protein in colorectal cancer cells were significantly higher than those in normal colorectal mucosa cells(P<0.05).After knockdown of TRIM59 expression,the proliferation activity and colony forming ability of HCT116 cells were significantly inhibited(P<0.05),and the cell cycle was arrested in G0/G1 phase.At the same time,the expression levels of CDK4 and CyclinD1 significantly decreased(P<0.05).Conclusion TRIM59 was highly expressed in colorectal cancer cells.Down regulation of TRIM59 expression can inhibit the proliferation and clone formation of colorectal cancer cells,suggesting that TRIM59 may become a new target for gene therapy of colorectal cancer.