微小RNA-1908对肺癌细胞A549细胞周期和靶基因的调控
Role of microRNA-1908 in cell cycle and target gene regulation in lung cancer A549 cells
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摘要:
目的 探讨微小RNA（microRNA，miRNA/miR）1908对肺癌细胞A549的细胞周期及靶基因的调控作用。
方法 通过慢病毒转染方法，上调A549细胞中miR-1908的表达水平，运用噻唑蓝法和流式细胞技术检测miR-1908对A549细胞增殖、周期和凋亡的影响。应用DIANA-TOOLs及KEGG数据库对miR-1908进行生物信息学分析，探讨其可能参与的信号通路和潜在靶基因。应用RT-qPCR和Western blot技术检测miR-1908对预测靶基因mRNA和蛋白表达水平的调控。
结果 与阴性对照组相比，转染组上调miR-1908表达可引起A549细胞G2期阻滞，S期减少（P < 0.05），对细胞增殖和凋亡无明显影响（P > 0.05）。miR-1908主要参与MAPK信号通路的调控，蛋白磷酸酶5（PP5）基因为其重要的潜在靶基因。与阴性对照组相比，转染组上调miR-1908表达可引起PP5的蛋白表达增加（P < 0.05），但其mRNA表达无明显改变（P>0.05）。
结论 miR-1908可能通过对PP5基因正向调控或间接调控影响A549细胞周期，研究结果加深了对miRNAs在肺癌细胞中的作用及其机制的理解，并为miRNAs在肺癌中的进一步研究提供线索。
Abstract:Objective To explore the regulation of cell cycle and target gene in lung cancer A549 cells by microRNA (miRNA/miR) 1908.
Methods Up-regulated miR-1908 expression in A549 cells was induced by lentivirus transfection. Then, MTT and flow cytometry were applied to detect the influence of miR-1908 on cell proliferation, apoptosis, and cell cycle of A549 cells. DIANATOOLs and KEGG databases were used to analyze the bioinformatics of miR-1908 and explore potential signaling pathways and target genes. RT-qPCR and Western blot were applied to detect the miR-1908 regulation on both mRNA and protein expression le vels of its predicted target gene.
Results Compared with the negative control group, the up-regulation of miR-1908 in the transfection group caused arrested G2 phase and shortened S phase of A549 cells (P < 0.05), but did not induce significant changes in cell proliferation and apoptosis (P > 0.05). miR-1908 was mainly related to MAPK signaling pathway, and protein phosphatase 5 (PP5) was an important potential target gene. Compared with the negative control group, the up-regulation of miR-1908 in the transfection group increased PP5 protein level (P < 0.05), but not its mRNA expression level (P>0.05).
Conclusion miR-1908 may affect the cell cycle of A549 cells by regulating PP5 positively or indirectly. The results help further understanding the role and mechanism of miRNAs in lung cancer cells and provide a novel clue for studying miRNAs in lung cancer.
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