Department of Dermatology, Shandong Provincial Hospital Affiliated to Shandong University, Jinan, Shandong 250021, P.R. China.
Laboratory of Medicine, Shandong Provincial Hospital Affiliated to Shandong University, Jinan, Shandong 250021, P.R. China.
Int J Mol Med. 2018 Mar;41(3):1275-1282. doi: 10.3892/ijmm.2017.3335. Epub 2017 Dec 20.
The present study aimed to investigate the potential role of the long non‑coding RNA (lncRNA) Pvt1 oncogene (non‑protein coding) (PVT1) in the progression and metastasis of malignant melanoma, and to reveal its possible molecular mechanisms. The expression of lncRNA PVT1 in melanoma tissues and adjacent normal skin from patients with melanoma, and in the melanoma A‑375 and sk‑mel‑5 cell lines, was analyzed using reverse transcription‑quantitative polymerase chain reaction and western blot analyses. The effects of PVT1 expression on cell proliferation, the cell cycle, cell migration and cell invasion were analyzed using MTT assay, flow cytometry, Transwell and scratch assays, respectively. The interaction between PVT1 and enhancer of zeste homolog 2 (EZH2) in melanoma cells was analyzed using RNA immunoprecipitation (RIP) assay. The effect of PVT1 on microRNA‑200c (miR‑200c) expression was analyzed by chromatin immunoprecipitation (ChIP) assay. PVT1 was highly expressed in the melanoma tissues and cells. Silencing of PVT1 significantly inhibited cell proliferation, migration and invasion, and arrested the cell cycle at the G0/G1 stage. Additionally, PVT1 silencing significantly decreased the cyclin D1 expression in the melanoma cells. The expression of E‑cadherin was significantly increased and the expression of N‑cadherin and vimentin was significantly decreased in the PVT1‑silenced group. The RIP assay found that endogenous PVT1 was highly enriched by EZH2 RIP compared with that of the negative control. The ChIP assay revealed that the expression of miR‑200c was decreased significantly in the PVT1‑silenced group compared with the controls. Overall, the present study demonstrated that the lncRNA PVT1 may contribute to the tumorigenesis and metastasis of melanoma through binding to EZH2 and regulating the expression of miR‑200c. lncRNA PVT1 may serve as a potential target for the therapy of melanoma.
本研究旨在探讨长链非编码 RNA(lncRNA) Pvt1 癌基因(非蛋白编码)(PVT1)在恶性黑色素瘤进展和转移中的潜在作用,并揭示其可能的分子机制。采用逆转录-定量聚合酶链反应和 Western blot 分析检测黑色素瘤患者肿瘤组织及癌旁正常皮肤、黑色素瘤 A-375 和 sk-mel-5 细胞系中 lncRNA PVT1 的表达。采用 MTT 检测、流式细胞术、Transwell 和划痕实验分别分析 PVT1 表达对细胞增殖、细胞周期、细胞迁移和细胞侵袭的影响。采用 RNA 免疫沉淀(RIP)实验分析 PVT1 与黑色素瘤细胞中增强子的锌指蛋白 2(EZH2)的相互作用。采用染色质免疫沉淀(ChIP)实验分析 PVT1 对 microRNA-200c(miR-200c)表达的影响。结果显示,lncRNA PVT1 在黑色素瘤组织和细胞中高表达。沉默 PVT1 可显著抑制细胞增殖、迁移和侵袭,并使细胞周期停滞在 G0/G1 期。此外,沉默 PVT1 可显著降低黑色素瘤细胞中细胞周期蛋白 D1 的表达。沉默 PVT1 组 E-钙黏蛋白的表达明显增加,N-钙黏蛋白和波形蛋白的表达明显减少。RIP 实验发现,与阴性对照组相比,EZH2 RIP 中内源性 PVT1 高度富集。ChIP 实验显示,与对照组相比,沉默 PVT1 组 miR-200c 的表达明显降低。综上所述,本研究表明,lncRNA PVT1 可能通过与 EZH2 结合并调节 miR-200c 的表达,促进黑色素瘤的发生和转移。lncRNA PVT1 可能成为黑色素瘤治疗的潜在靶点。