Downregulated PITX1 Modulated by MiR-19a-3p Promotes Cell Malignancy and Predicts a Poor Prognosis of Gastric Cancer by Affecting Transcriptionally Activated PDCD5
Downregulated PITX1 Modulated by MiR-19a-3p Promotes Cell Malignancy and Predicts a Poor Prognosis of Gastric Cancer by Affecting Transcriptionally Activated PDCD5
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DOI：
10.1159/000489590
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发表时间：
2018-01-01
影响因子：
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通讯作者：
Fan, Hong
中科院分区：
文献类型：
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作者：
Qiao, Fengchang;Gong, Pihai;Fan, Hong
背景/目的：PITX1基因在多种恶性肿瘤中被认为是一个潜在的抑癌基因。PITX1的分子机制，特别是其作为转录因子在肿瘤发生过程中调节基因表达的功能，目前仍不清楚。方法：采用定量逆转录聚合酶链式反应(qRT-PCR)和免疫组织化学方法检测PITX1在胃癌组织中的表达水平和定位。分析PITX1在体内外对胃癌细胞增殖和成瘤的影响。为了探索PITX1如何抑制细胞增殖，我们使用PITX1芯片测序技术来测量PITX1的全基因组结合位点，并基于其可能的靶基因评估全球功能相关性。芯片聚合酶链式反应、凝胶迁移率改变分析和启动子报告分析检测PITX1是否与PDCD5结合并调节其表达。在体内外进一步研究了PDCD5在胃癌细胞凋亡中的作用。用Kaplan-Meier估计法评估PITX1蛋白水平与GC患者预后的关系。同时，通过荧光素酶报告实验、qRT-PCR和Western blotting检测与PITX1相关的miR-19a-3p的表达水平。结果：PITX1在胃癌组织和细胞系中的表达水平降低。在体内外，PITX1的高表达显著抑制了GC细胞的增殖和肿瘤的形成。PITX1基因敲除可阻断其对GC细胞增殖的抑制作用。PITX1与全基因组结合，这些靶点富含主要与细胞生长和凋亡相关的功能基因。PITX1在肿瘤发生过程中与细胞凋亡相关基因PDCD5结合，并受顺式调控的PDCD5表达。胃癌细胞中PDCD5表达的增加不仅诱导了胃癌细胞的凋亡，而且在体内外抑制了胃癌细胞的生长。此外，PITX1的表达受miR-19a-3p的调控。更重要的是，PITX1蛋白水平降低与GC患者预后不良相关。结论：PITX1表达降低预示着GC患者总生存期缩短。PITX1作为一种转录激活因子，在胃癌发生过程中调控包括PDCD5在内的细胞凋亡相关基因。这些数据表明，PDCD5是一种新的、可行的GC治疗靶点。(C)2018年作者(S)由S.Karger AG，巴塞尔出版
Background/Aims: PITX1 has been identified as a potential tumor-suppressor gene in several malignant tumors. The molecular mechanism underlying PITX1, particularly its function as a transcription factor regulating gene expression during tumorigenesis, is still poorly understood. Methods: The expression level and location of PITX1 were determined by quantitative reverse transcription PCR (qRT-PCR) and immunohistochemical staining in gastric cancer (GC). The effect of PITX1 on the GC cell proliferation and tumorigenesis was analyzed in vitro and in vivo. To explore how PITX1 suppresses cell proliferation, we used PITX1-ChIP-sequencing to measure genome-wide binding sites of PITX1 and assessed global function associations based on its putative target genes. ChIP-PCR, electrophoretic mobility shift assay, and promoter reporter assays examined whether PITX1 bound to PDCD5 and regulated its expression. The function of PDCD5 in GC cell apoptosis was further examined in vitro and in vivo. The relationship between the PITX1 protein level and GC patient prognosis was evaluated by the Kaplan-Meier estimator. Meanwhile, the expression level of miR-19a-3p, which is related to PITX1, was also detected by luciferase reporter assay, qRT-PCR, and western blotting. Results: The expression level of PITX1 was decreased in GC tissues and cell lines. Elevated PITX1 expression significantly suppressed the cell proliferation of GC cells and tumorigenesis in vitro and in vivo. PITX1 knockdown blocked its inhibition of GC cell proliferation. PITX1 bound to whole genomewide sites, with these targets enriched on genes with functions mainly related to cell growth and apoptosis. PITX1 bound to PDCD5, an apoptosis-related gene, during tumorigenesis, and cis-regulated PDCD5 expression. Increased PDCD5 expression in GC cells not only induced GC cell apoptosis, but also suppressed GC cell growth in vitro and in vivo. Moreover, PITX1 expression was regulated by miR-19a-3p. More importantly, a decreased level of PITX1 protein was correlated with poor GC patient prognosis. Conclusion: Decreased expression of PITX1 predicts shorter overall survival in GC patients. As a transcriptional activator, PITX1 regulates apoptosis-related genes, including PDCD5, during gastric carcinogenesis. These data indicate PDCD5 to be a novel and feasible therapeutic target for GC. (C) 2018 The Author(s) Published by S. Karger AG, Basel