title: miR-150, p53 protein and relevant miRNAs consist of a regulatory network in NSCLC tumorigenesis.
abstract: microRNAs (miRNAs) are a class of non-coding small RNAs that act as negative regulators of gene expression by binding to the 3'-untranslated region (3'-UTR) of target mRNAs. Tumor protein p53, a transcriptional factor, plays an important role in the progression of tumorigenesis. miR-150 was the only miRNA predicted to target 3'-UTR of p53 by Targetscan. In order to investigate the function of miR-150, p53 and relevant miRNAs in non-small cell lung cancer (NSCLC), we constructed two expression vectors of p53 (pcDNA3.1-p53 and pcDNA3.1-p53-3'-UTR) and two report vectors (pGL3-p53-3'-UTR and pGL3-p53-3'-mUTR). The activity of luciferase transfected with miR-150 mimics was lower by 30% when compared to that of the miRNA-negative control (miRNA-NC). Moreover, the p53 protein was downregulated by at least 50% when miR-150 mimics were cotransfected with pcDNA3.1-p53-3'-UTR when compared to miRNA-NC. We also determined the expression of miR-150 and p53 in NSCLC patient tissue samples. The expression of miR-150 in T2 stage tissue samples was higher than that in T1 stage tissue samples. The corresponding target gene p53 was correlated with miR-150 expression. In the present study, we further analyzed the cell cycle distribution. The cells transfected with pcDNA3.1-p53 were significantly arrested in the G1 phase when compared to the control cells. When miR-150 mimics were cotransfected with pcDNA3.1-p53-3'-UTR, the percentage of cells in the G1 phase was significantly lower by 4% when compared to miRNA-NC. To identify miRNAs that are regulated by the p53 protein, qRT-PCR was performed after pcDNA3.1-p53 transfection. miR-34a, miR-184, miR-181a and miR-148 were upregulated significantly. However, there was no distinct difference in the expression of miR-10a, miR-182 and miR-34c. Our results showed that miR-150 targets the 3'-UTR of p53, and p53 protein promotes the expression of miRNAs which affect cell cycle progression. These findings suggest that miR-150, p53 protein and relevant miRNAs are members of a regulatory network in NSCLC tumorigenesis.
authors.affiliation: School of Life Sciences, Shanghai University, Shanghai 200444, P.R. China.
authors.forename: De-Tao
authors.initials: DT
authors.lastname: Wang
authors.forename: Zhong-Liang
authors.initials: ZL
authors.lastname: Ma
authors.forename: Yan-Li
authors.initials: YL
authors.lastname: Li
authors.forename: Yue-Qing
authors.initials: YQ
authors.lastname: Wang
authors.forename: Bo-Tao
authors.initials: BT
authors.lastname: Zhao
authors.forename: Jia-Li
authors.initials: JL
authors.lastname: Wei
authors.forename: Xiang
authors.initials: X
authors.lastname: Qi
authors.forename: Xin-Tai
authors.initials: XT
authors.lastname: Zhao
authors.forename: You-Xin
authors.initials: YX
authors.lastname: Jin
mesh_terms: D020413:3' Untranslated Regions / Q000235:genetics; D063646:Carcinogenesis / Q000235:genetics*; D002289:Carcinoma, Non-Small-Cell Lung / Q000235:genetics / Q000378:metabolism*; D002453:Cell Cycle / Q000235:genetics; D045744:Cell Line, Tumor; D059585:G1 Phase Cell Cycle Checkpoints / Q000235:genetics; D015972:Gene Expression Regulation, Neoplastic; D057809:HEK293 Cells; D006801:Humans; D008175:Lung Neoplasms / Q000235:genetics / Q000378:metabolism*; D035683:MicroRNAs / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*; D020411:Oligonucleotide Array Sequence Analysis; D016159:Tumor Suppressor Protein p53 / Q000235:genetics / Q000378:metabolism*
chemical_list: D020413:3' Untranslated Regions; D035683:MicroRNAs; D016159:Tumor Suppressor Protein p53; C550570:MIRN148 microRNA, human; C533097:MIR150, human; C534424:MIRN184 microRNA, human; C522425:MIRN34 microRNA, human; C540193:MIrn181 microRNA, human