Ye Hua, Li Wende, Wu Kefeng, Liu Yi, Lv Yingnian, Zhu Yuzhen, Luo Hui, Cui Liao
Guangdong Key Laboratory for Research and Development of Natural Drugs, Guangdong Medical University, Zhanjiang, People's Republic of China.
Institute of Marine Biomedical Research, Guangdong Medical University, Zhanjiang, People's Republic of China.
Onco Targets Ther. 2020 Nov 16;13:11711-11724. doi: 10.2147/OTT.S277254. eCollection 2020.
Accumulating evidence has indicated that long noncoding RNAs (lncRNAs) are pivotal regulators involved in the pathogenesis of cancer; however, the molecular mechanism of LINC00339 in colorectal cancer (CRC) remains unclear.
The quantitative real-time polymerase chain reaction for the expression of LINC00339 and miR-378a-3p and Western blots for were performed. A dual-luciferase assay was used to investigate the interaction between LIN00339 and miR-378a-3p, as well as between miR-378a-3p and . Cell proliferation was determined by 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) and 5-ethynyl-2'-deoxyuridine (EdU) assay. The cell cycle was analyzed by propidium iodide staining followed by flow cytometry analysis. The wound-healing and transwell invasion assays were used to evaluate cell migration and invasion.
The expression of LINC00339 was significantly upregulated in CRC cells and tissues, and high LINC00339 expression indicated an advanced tumor stage. Further experiments demonstrated that activated LINC00339 expression by binding to its promoter region. Luciferase activity and RNA pull-down assays demonstrated a direct interaction between LINC00339 and miR-378a-3p. miR-378a-3p expression was decreased in CRC samples and negatively correlated with LINC00339 expression in tumors. Gain- and loss-of-function assays indicated that LINC00339 contributed to cell proliferation, cell cycle progression, migration, and invasion, while miR-378a-3p reversed these effects. Furthermore, cotransfection of wild-type 3'-UTR reporters and miR-378a-3p significantly reduced luciferase activity. mRNA and protein expression was inhibited and enhanced by miR-378a-3p and LINC00339, respectively. overexpression reversed the effect of miR-378a-3p on cellular processes. Moreover, LINC00339 promoted tumor growth in vivo and induced epithelial-mesenchymal transition (EMT) and activated the Wnt/β-catenin signaling pathway in cells.
Our findings demonstrate the regulatory role of the SP1/LINC00339/miR-378a-3p/MED19 axis in CRC tumorigenesis and provide novel insight into the molecular mechanism underlying CRC.
越来越多的证据表明，长链非编码RNA（lncRNA）是参与癌症发病机制的关键调节因子；然而，LINC00339在结直肠癌（CRC）中的分子机制仍不清楚。
进行了定量实时聚合酶链反应以检测LINC00339和miR-378a-3p的表达，并进行了蛋白质免疫印迹分析。采用双荧光素酶报告基因检测法研究LINC00339与miR-378a-3p之间以及miR-378a-3p与……之间的相互作用。通过3-(4,5-二甲基-2-噻唑基)-2,5-二苯基-2H-四唑溴盐（MTT）和5-乙炔基-2'-脱氧尿苷（EdU）检测法测定细胞增殖。通过碘化丙啶染色后进行流式细胞术分析来分析细胞周期。采用伤口愈合实验和Transwell侵袭实验评估细胞迁移和侵袭能力。
LINC00339在CRC细胞和组织中的表达显著上调，高LINC00339表达表明肿瘤分期较晚。进一步实验表明……通过结合其启动子区域激活LINC00339的表达荧光素酶活性和RNA下拉实验证明LINC00339与miR-378a-3p之间存在直接相互作用。miR-378a-3p在CRC样本中的表达降低，且与肿瘤中LINC00339的表达呈负相关。功能获得和功能丧失实验表明，LINC00339促进细胞增殖、细胞周期进程、迁移和侵袭，而miR-378a-3p可逆转这些作用。此外，共转染野生型……3'-UTR报告基因和miR-378a-3p可显著降低荧光素酶活性。miR-378a-3p和LINC00339分别抑制和增强……的mRNA和蛋白表达。……过表达逆转了miR-378a-3p对细胞过程的影响。此外，LINC00339在体内促进肿瘤生长，并诱导上皮-间质转化（EMT），激活细胞中的Wnt/β-连环蛋白信号通路。
我们的研究结果证明了SP1/LINC00339/miR-378a-3p/MED19轴在CRC肿瘤发生中的调节作用，并为CRC潜在的分子机制提供了新的见解。
