敲低 lncRNA-UCA1 通过 miR-193a 介导的 CDK6 下调抑制人神经胶质瘤细胞的活力和迁移。
Knockdown of lncRNA-UCA1 inhibits cell viability and migration of human glioma cells by miR-193a-mediated downregulation of CDK6.
机构信息
Department of Neurosurgery, Anqiu People's Hospital, Anqiu, 262100, China.
Department of Neurology, Anqiu People's Hospital, Anqiu, 262100, China.
出版信息
J Cell Biochem. 2019 Sep;120(9):15157-15169. doi: 10.1002/jcb.28777. Epub 2019 May 20.
BACKGROUND
Long noncoding RNA urothelial carcinoma-associated 1 (lncRNA-UCA1) is generally recognized as an oncogenic molecule in several human malignant tumors. However, the available evidence does not necessarily imply an unequivocal causal function of UCA1 in glioblastoma. The current study was aimed to probe the biological function of lncRNA-UCA1 in human glioblastoma cell lines. Besides, we further investigated the potential mechanisms.
METHODS
Cell viability, apoptosis, as well as migration and invasion were measured using a commercial cell counting kit-8, flow cytometry, and 24-Transwell assay, respectively. LncRNA-UCA1, microRNA-193a (miR-193a), and CDK6 at messenger RNA levels were evaluated by quantitative real-time polymerase chain reaction method. Protein level was examined by Western blot analysis. RNA immunoprecipitation was utilized to validate lncRNA-UCA1 associated with miR-193a. Luciferase activity assay was used to identify the miR-193a-targeted CDK6 3'-untranslated region.
RESULTS
lncRNA-UCA1 knockdown weakened cell viability, augmented apoptosis progression, as well as suppressed migration and invasion behaviors in glioblastoma cells, whereas lncRNA-UCA1 silence exhibited the opposite functions. lncRNA-UCA1 functioned as an endogenous sponge of miR-193a. miR-193a silence reversed the biological function of lncRNA-UCA1 knockdown on U-118 MG cells. miR-193a negatively regulated the expression of CDK6, and it affected the U-118 MG cells through regulating CDK6 expression. CDK6 overexpression abrogated the blockage of PI3K/AKT, mitogen-activated protein kinase (MAPK), and Notch signaling pathways. Furthermore, lncRNA-UCA1 and miR-193a could affect these signaling cascades through regulating CDK6 expression. The regulatory mechanisms of lncRNA-UCA1 were further consolidated in clinical specimens.
CONCLUSION
lncRNA-UCA1 silence reduced cell viability, promoted apoptosis progression, while impeding the migration and invasion of glioblastoma cells by miR-193a-mediated silence of CDK6, with blockage of PI3K/AKT, MAPK, and Notch pathways.
背景
长链非编码 RNA 尿路上皮癌相关 1（lncRNA-UCA1）被普遍认为是几种人类恶性肿瘤中的致癌分子。然而，现有证据并不一定表明 UCA1 在神经胶质瘤中具有明确的因果功能。本研究旨在探讨 lncRNA-UCA1 在人神经胶质瘤细胞系中的生物学功能。此外，我们进一步研究了潜在的机制。
方法
使用商业细胞计数试剂盒-8、流式细胞术和 24 孔 Transwell 测定分别测量细胞活力、凋亡以及迁移和侵袭。通过定量实时聚合酶链反应方法评估 lncRNA-UCA1、微小 RNA-193a（miR-193a）和细胞周期蛋白依赖性激酶 6（CDK6）的信使 RNA 水平。通过 Western blot 分析检测蛋白水平。利用 RNA 免疫沉淀验证 lncRNA-UCA1 与 miR-193a 的关联。使用荧光素酶活性测定鉴定 miR-193a 靶向的 CDK6 3'-非翻译区。
结果
lncRNA-UCA1 敲低削弱了神经胶质瘤细胞的活力，促进了细胞凋亡的进展，并抑制了迁移和侵袭行为，而 lncRNA-UCA1 沉默则表现出相反的功能。lncRNA-UCA1 作为 miR-193a 的内源性海绵起作用。miR-193a 沉默逆转了 U-118 MG 细胞中 lncRNA-UCA1 敲低的生物学功能。miR-193a 负调控 CDK6 的表达，并通过调节 CDK6 表达影响 U-118 MG 细胞。CDK6 过表达阻断了 PI3K/AKT、丝裂原活化蛋白激酶（MAPK）和 Notch 信号通路。此外，lncRNA-UCA1 和 miR-193a 可以通过调节 CDK6 的表达来影响这些信号级联。在临床标本中进一步证实了 lncRNA-UCA1 的调控机制。
结论
lncRNA-UCA1 沉默通过 miR-193a 介导的 CDK6 沉默减少神经胶质瘤细胞的活力，促进细胞凋亡的进展，同时抑制细胞的迁移和侵袭，并阻断 PI3K/AKT、MAPK 和 Notch 通路。
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