Glioblastoma (GBM) is the most common primary malignant intracranial adult brain tumor. Allelic deletion on chromosome 14q plays an essential role in GBM pathogenesis, and this chromosome 14q site was thought to harbor multiple tumor suppressor genes associated with GBM, a region that also encodes microRNA-203 (miR-203). This study was conducted to identify whole transcriptome profile changes associated with miR-203 expression by high-throughput RNA sequencing. Enrichment analyses for gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis revealed that miR-203 expression had a strong, negative effect on a number of fundamental and interconnected biological processes involved in cell growth and proliferation. The biological processes mostly influenced were p53 signaling pathway, FoxO signaling pathway, DNA replication, cell cycle, MAPK signaling pathway, and apoptosis. In total, 847 upregulated and 345 downregulated differentially expressed genes were identified in control versus miR-203 expressing glioma cells. After GO enrichment, the downregulated differentially expressed genes such as BCL2, SPARC were found to be mainly enriched in cell cycle regulation and apoptosis processes, whereas the upregulated differentially expressed genes such as CCND1, E2F1 were involved in the DNA replication and cell cycle regulation. We also performed miR-203 target analysis and found BCL2, AKT, SPARC, ROBO1, c-JUN, PDGFA, and CREB were predicted target of miR-203 and miR-203 expression suppressed the protein and mRNA levels of these target genes by western blotting and qRT-PCR analysis. Moreover, co-transfection experiments using a luciferase-based reporter assay demonstrated that miR-203 directly regulated BCL-2 expression and BCL-2 overexpression suppressed miR-203 mediated glioma cell apoptosis. These results indicate that overexpression of miR-203 coordinately regulates several oncogenic pathways in GBM.

中文翻译：

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通过全面的RNA测序转录组分析来阐明胶质母细胞瘤细胞中的microRNA-203特定生物学过程。

胶质母细胞瘤（GBM）是最常见的原发性恶性颅内成人脑肿瘤。14q染色体上的等位基因缺失在GBM发病机理中起着至关重要的作用，据认为该14q染色体位点包含与GBM相关的多个抑癌基因，该区域也编码microRNA-203（miR-203）。进行了这项研究，以通过高通量RNA测序鉴定与miR-203表达相关的整个转录组图谱变化。基因本体论的富集分析（GO）和《京都议定书》的基因与基因组百科全书（KEGG）途径分析显示，miR-203表达对涉及细胞生长和增殖的许多基本和相互关联的生物过程具有强烈的负面影响。影响最大的生物学过程是p53信号通路，FoxO信号通路，DNA复制，细胞周期，MAPK信号通路和凋亡。在对照表达miR-203的神经胶质瘤细胞中，总共鉴定出847个上调的差异表达基因和345个下调的差异表达基因。GO富集后，发现下调的差异表达基因如BCL2，SPARC主要富集于细胞周期调控和凋亡过程，而上调的差异表达基因如CCND1，E2F1参与DNA复制和细胞周期调控。我们还进行了miR-203靶标分析，发现BCL2，AKT，SPARC，ROBO1，c-JUN，PDGFA和CREB被认为是miR-203和miR-203表达的靶标，通过蛋白质印迹抑制了这些靶标基因的蛋白质和mRNA水平。印迹和qRT-PCR分析。而且，使用基于荧光素酶的报告基因检测的共转染实验表明，miR-203直接调节BCL-2表达，而BCL-2过表达抑制miR-203介导的神经胶质瘤细胞凋亡。这些结果表明，miR-203的过度表达协调调节GBM中的几种致癌途径。