State Key Laboratory of Cancer Biology, Department of Biochemistry and Molecular Biology, Fourth Military Medical University, Xi'an, China.
Department of Immunology, Fourth Military Medical University, Xi'an, China.
Theranostics. 2018 Apr 15;8(10):2846-2861. doi: 10.7150/thno.23463. eCollection 2018.
Transforming growth factor-β (TGF-β) has received much attention as a major inducer of epithelial-mesenchymal transition (EMT) during cancer progression, mainly by activating a set of pleiotropic transcription factors including SNAI2/Slug. However, the involvement of long non-coding RNAs (lncRNAs) in TGF-β-induced Slug activation and EMT remains largely unknown. In this study, we used microarray analysis to compare lncRNA expression profiles between TGF-β treated and untreated breast cancer cells. Then, the clinical significance of lncRNAs in breast cancer was investigated by qPCR and Kaplan-Meier survival analysis. The molecular mechanisms and EMT-promoting effects were analyzed by confocal laser microscopy, Western blotting, chromosome conformation capture (3C), chromatin isolation by RNA purification (ChIRP), ChIP, luciferase reporter assay and transwell migration assay. Lastly, the pro-metastatic effects were evaluated by bioluminescent imaging and hematoxylin and eosin (H&E) staining. We observed that TGF-β induced genome-wide changes in lncRNA levels in breast cancer cells, among which AC026904.1 and UCA1 were highly expressed in metastatic breast cancer and closely associated with poor prognosis. Mechanistic study revealed that AC026904.1 and UCA1 were upregulated by non-canonical and canonical TGF-β pathways, respectively. Further analysis showed that AC026904.1 functions as an enhancer RNA in the nucleus, whereas UCA1 exerts a competitive endogenous RNA (ceRNA) activity in the cytoplasm. In addition, the biological functions of these two lncRNAs converged on the activation and maintenance of Slug, constituting a one-two punch in promoting EMT and tumor metastasis. These findings uncover for the first time that AC026904.1 and UCA1 could cooperatively upregulate Slug expression at both transcriptional and post-transcriptional levels, exerting critical roles in TGF-β-induced EMT. The present work provides new evidence that lncRNAs function as key regulators of EMT and hold great promise to be used as novel biomarkers and therapeutic targets for metastatic breast cancer.
转化生长因子-β（TGF-β）作为癌症进展过程中上皮-间充质转化（EMT）的主要诱导因子受到了广泛关注，主要通过激活包括 SNAI2/Slug 在内的一组多效性转录因子。然而，长链非编码 RNA（lncRNA）在 TGF-β诱导的 Slug 激活和 EMT 中的作用在很大程度上仍然未知。在这项研究中，我们使用微阵列分析比较了 TGF-β处理和未处理的乳腺癌细胞之间的 lncRNA 表达谱。然后，通过 qPCR 和 Kaplan-Meier 生存分析研究了 lncRNA 在乳腺癌中的临床意义。通过共聚焦激光显微镜、Western blot、染色体构象捕获（3C）、RNA 纯化染色质分离（ChIRP）、ChIP、荧光素酶报告基因测定和 Transwell 迁移测定分析了分子机制和 EMT 促进作用。最后，通过生物发光成像和苏木精和伊红（H&E）染色评估了促转移作用。我们观察到 TGF-β诱导乳腺癌细胞中 lncRNA 水平的全基因组变化，其中 AC026904.1 和 UCA1 在转移性乳腺癌中高表达，并与不良预后密切相关。机制研究表明，AC026904.1 和 UCA1 分别通过非经典和经典 TGF-β途径上调。进一步分析表明，AC026904.1 在核内作为增强子 RNA 发挥作用，而 UCA1 在细胞质中发挥竞争性内源性 RNA（ceRNA）活性。此外，这两种 lncRNA 的生物学功能集中在 Slug 的激活和维持上，在促进 EMT 和肿瘤转移方面形成了一个一二拳。这些发现首次揭示，AC026904.1 和 UCA1 可以协同上调 Slug 在转录和转录后水平的表达，在 TGF-β诱导的 EMT 中发挥关键作用。本工作为 AC026904.1 和 UCA1 可以协同上调 Slug 在转录和转录后水平的表达，在 TGF-β诱导的 EMT 中发挥关键作用。本工作为 AC026904.1 和 UCA1 可以协同上调 Slug 在转录和转录后水平的表达，在 TGF-β诱导的 EMT 中发挥关键作用。本工作提供了新的证据，表明 lncRNA 作为 EMT 的关键调节剂发挥作用，并有望成为转移性乳腺癌的新型生物标志物和治疗靶点。
