Soochow University Laboratory of Cancer Molecular Genetics, Medical College of Soochow University, 199 Ren'ai Road, Suzhou, 215123, Jiangsu, China.
Laboratory Animal Center, Medical College of Soochow University, Suzhou, 215123, Jiangsu, China.
Mol Cancer. 2018 Sep 27;17(1):140. doi: 10.1186/s12943-018-0889-7.
TGF-β promotes tumor invasion and metastasis through inducing epithelial-mesenchymal transition (EMT) in non-small cell lung cancer (NSCLC). Circular RNAs (circRNAs) are recognized as functional non-coding RNAs involved in human cancers. However, whether and how circRNAs contribute to TGF-β-induced EMT and metastasis in NSCLC remain vague. Here, we investigated the regulation and function of Circular RNA hsa_circ_0008305 (circPTK2) in TGF-β-induced EMT and tumor metastasis, as well as a link between circPTK2 and transcriptional intermediary factor 1 γ (TIF1γ) in NSCLC.
Circular RNAs were determined by human circRNA Array analysis, real-time quantitative reverse transcriptase PCR and northern blot. Luciferase reporter, RNA-binding protein immunoprecipitation (RIP), RNA pull-down and fluorescence in situ hybridization (FISH) assays were employed to test the interaction between circPTK2 and miR-429/miR-200b-3p. Ectopic overexpression and siRNA-mediated knockdown of circPTK2, TGF-β-induced EMT, Transwell migration and invasion in vitro, and in vivo experiment of metastasis were used to evaluate the function of circPTK2. Transcription and prognosis analyses were done in public databases.
CircPTK2 and TIF1γ were significantly down-regulated in NSCLC cells undergoing EMT induced by TGF-β. CircPTK2 overexpression augmented TIF1γ expression, inhibited TGF-β-induced EMT and NSCLC cell invasion, whereas circPTK2 knockdown had the opposite effects. CircPTK2 functions as a sponge of miR-429/miR-200b-3p, and miR-429/miR-200b-3p promote TGF-β-induced EMT and NSCLC cell invasion by targeting TIF1γ. CircPTK2 overexpression inhibited the invasion-promoting phenotype of endogenous miR-429/miR-200b-3p in NSCLC cells in response to TGF-β. CircPTK2 overexpression significantly decreased the expression of Snail, an important downstream transcriptional activator of TGF-β/Smad signaling. In an in vivo experiment of metastasis, circPTK2 overexpression suppressed NSCLC cell metastasis. Moreover, circPTK2 expression was dramatically down-regulated and positively correlated with TIF1γ expression in human NSCLC tissues. Especially, circPTK2 was significantly lower in metastatic NSCLC tissues than non-metastatic counterparts.
Our findings show that circPTK2 (hsa_circ_0008305) inhibits TGF-β-induced EMT and metastasis by controlling TIF1γ in NSCLC, revealing a novel mechanism by which circRNA regulates TGF-β-induced EMT and tumor metastasis, and suggesting that circPTK2 overexpression could provide a therapeutic strategy for advanced NSCLC.
TGF-β 通过诱导非小细胞肺癌（NSCLC）中的上皮-间充质转化（EMT）促进肿瘤侵袭和转移。环状 RNA（circRNA）被认为是参与人类癌症的功能性非编码 RNA。然而，circRNA 是否以及如何促进 NSCLC 中的 TGF-β 诱导的 EMT 和转移仍然不清楚。在这里，我们研究了环状 RNA hsa_circ_0008305（circPTK2）在 TGF-β 诱导的 EMT 和肿瘤转移中的调节和功能，以及 circPTK2 和转录中介因子 1γ（TIF1γ）在 NSCLC 中的关系。
通过人环状 RNA 阵列分析、实时定量逆转录 PCR 和 northern blot 确定环状 RNA。荧光素酶报告、RNA 结合蛋白免疫沉淀（RIP）、RNA 下拉和荧光原位杂交（FISH）实验用于测试 circPTK2 与 miR-429/miR-200b-3p 之间的相互作用。外源性过表达和 siRNA 介导的 circPTK2 下调、体外 TGF-β 诱导的 EMT、Transwell 迁移和侵袭以及转移的体内实验用于评估 circPTK2 的功能。在公共数据库中进行转录和预后分析。
在 TGF-β 诱导的 EMT 过程中，NSCLC 细胞中 circPTK2 和 TIF1γ 的表达明显下调。circPTK2 过表达增加了 TIF1γ 的表达，抑制了 TGF-β 诱导的 EMT 和 NSCLC 细胞侵袭，而 circPTK2 下调则产生相反的效果。circPTK2 作为 miR-429/miR-200b-3p 的海绵，miR-429/miR-200b-3p 通过靶向 TIF1γ 促进 TGF-β 诱导的 EMT 和 NSCLC 细胞侵袭。circPTK2 过表达抑制了 TGF-β 刺激的内源性 miR-429/miR-200b-3p 在 NSCLC 细胞中的侵袭促进表型。circPTK2 过表达显著降低了 TGF-β/Smad 信号转导下游重要转录激活因子 Snail 的表达。在转移的体内实验中，circPTK2 过表达抑制了 NSCLC 细胞的转移。此外，circPTK2 的表达在人类 NSCLC 组织中明显下调，并与 TIF1γ 的表达呈正相关。特别是，转移性 NSCLC 组织中的 circPTK2 明显低于非转移性 NSCLC 组织。
我们的研究结果表明，circPTK2（hsa_circ_0008305）通过控制 NSCLC 中的 TIF1γ 抑制 TGF-β 诱导的 EMT 和转移，揭示了 circRNA 调节 TGF-β 诱导的 EMT 和肿瘤转移的新机制，并表明 circPTK2 过表达可为晚期 NSCLC 提供治疗策略。
