背景与目的:NOB1(NIN1/RPN12 binding protein 1 homolog)是2005年新克隆的一个基因,属于RNA结合蛋白,该类蛋白的功能与恶性肿瘤的发生密切相关.本研究旨在观察利用慢病毒介导的RNAi沉默NOB1基因对人乳腺癌MCF-7细胞增殖及细胞周期的影响.方法:包装表达NOB1短发夹RNA(shRNA)的慢病毒,感染MCF-7细胞,实时定量PCR和蛋白质印迹法(Western blot)验证NOB1的抑制效率;MTT和克隆形成实验检测NOB1对细胞增殖能力的影响;流式细胞术检测细胞周期的变化.结果:NOB1-shRNA慢病毒感染3 d后,可显著下调MCF-7细胞NOB1 mRNA和蛋白的表达水平,显著抑制细胞增殖和体外成瘤能力,并导致细胞周期分布紊乱,G0/G1期及G2/M期细胞增加,S期细胞减少.结论:NOB1基因通过调节细胞周期分布促进乳腺癌细胞恶性增殖,可能是乳腺癌基因治疗的分子靶点.%Background and purpose: NOB1 (N3N1/RPN12 binding protein 1 homolog) is a kind of new gene, cloned in 2005, encodes a UNA-binding protein. This type of protein is often associated with the occurrence of malignant tumors. This study aimed to investigate the functional role of NOBI in the modulating of breast cancer cell proliferation and cell cycle progression by lentivirus-mediated RNA interference. Methods: The lentivirus that expresses NOB1 short harping RNA (shRNA) was constructed and infected into breast MCF-7 cells to silence the expression of NOB1. MTT and colony formation assay were used assess the proliferation ability of MCF-7 cells that transduced with NOB1-shRNA-cxprcssing lentivirus. Flow cytomctry assay wras performed to detect the cell cycle progression. Remits: The expression of NOB1 was significantly downreguiated at both mRNA and protein levels at 3 days after NOB1-shRNA-expressing lentivirus infection. NOB1 suppression resulted in impaired cell proliferation and colony formation ability, and induced the Go/G1 and G2/M phase cell cycle arrest and S phase depression in MCF-7 cells. Conclusion: NOB1 promotes breast cancer proliferation by regulating the cell cycle progression, and may be a potential molecular target in breast cancer gene therapy.
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