TEAD4 调控的长链非编码 RNA MNX1-AS1 部分通过抑制 BTG2 和激活 BCL2 促进胃癌的进展。

BACKGROUND: Gastric cancer (GC) is the third leading cause of cancer-related mortality globally. Long noncoding RNAs (lncRNAs) are dysregulated in obvious malignancies including GC and exploring the regulatory mechanisms underlying their expression is an attractive research area. However, these molecular mechanisms require further clarification, especially upstream mechanisms. METHODS: LncRNA MNX1-AS1 expression in GC tissue samples was investigated via microarray analysis and further determined in a cohort of GC tissues via quantitative reverse transcription polymerase chain reaction (qRT-PCR) assays. Cell proliferation and flow cytometry assays were performed to confirm the roles of MNX1-AS1 in GC proliferation, cell cycle regulation, and apoptosis. The influence of MNX1-AS1 on GC cell migration and invasion was explored with Transwell assays. A xenograft tumour model was established to verify the effects of MNX1-AS1 on in vivo tumourigenesis. The TEAD4-involved upstream regulatory mechanism of MNX1-AS1 was explored through ChIP and luciferase reporter assays. The mechanistic model of MNX1-AS1 in regulating gene expression was further detected by subcellular fractionation, FISH, RIP, ChIP and luciferase reporter assays. RESULTS: It was found that MNX1-AS1 displayed obvious upregulation in GC tissue samples and cell lines, and ectopic expression of MNX1-AS1 predicted poor clinical outcomes for patients with GC. Overexpressed MNX1-AS1 expression promoted proliferation, migration and invasion of GC cells markedly, whereas decreased MNX1-AS1 expression elicited the opposite effects. Consistent with the in vitro results, MNX1-AS1 depletion effectively inhibited the growth of xenograft tumour in vivo. Mechanistically, TEAD4 directly bound the promoter region of MNX1-AS1 and stimulated the transcription of MNX1-AS1. Furthermore, MNX1-AS1 can sponge miR-6785-5p to upregulate the expression of BCL2 in GC cells. Meanwhile, MNX1-AS1 suppressed the transcription of BTG2 by recruiting polycomb repressive complex 2 to BTG2 promoter regions. CONCLUSIONS: Our findings demonstrate that MNX1-AS1 may be able to serve as a prognostic indicator in GC patients and that TEAD4-activatd MNX1-AS1 can promote GC progression through EZH2/BTG2 and miR-6785-5p/BCL2 axes, implicating it as a novel and potent target for the treatment of GC.

背景：胃癌（GC）是全球癌症相关死亡的第三大主要原因。长链非编码 RNA（lncRNA）在包括 GC 在内的明显恶性肿瘤中失调，探索其表达的调控机制是一个有吸引力的研究领域。然而，这些分子机制需要进一步澄清，特别是上游机制。

方法：通过微阵列分析研究了 GC 组织样本中 lncRNA MNX1-AS1 的表达，并通过定量逆转录聚合酶链反应（qRT-PCR）试验进一步在 GC 组织样本中进行了确定。细胞增殖和流式细胞术检测用于证实 MNX1-AS1 在 GC 增殖、细胞周期调控和细胞凋亡中的作用。通过 Transwell 检测探索了 MNX1-AS1 对 GC 细胞迁移和侵袭的影响。建立异种移植肿瘤模型以验证 MNX1-AS1 对体内肿瘤发生的影响。通过 ChIP 和荧光素酶报告基因检测探索了 MNX1-AS1 的 TEAD4 涉及的上游调控机制。通过亚细胞分离、FISH、RIP、ChIP 和荧光素酶报告基因检测进一步检测了 MNX1-AS1 调节基因表达的机制模型。

结果：研究发现，MNX1-AS1 在 GC 组织样本和细胞系中表现出明显上调，GC 患者中 MNX1-AS1 的过表达预示着不良的临床结局。过表达 MNX1-AS1 表达明显促进 GC 细胞的增殖、迁移和侵袭，而减少 MNX1-AS1 表达则产生相反的效果。与体外结果一致，MNX1-AS1 耗竭有效地抑制了体内异种移植肿瘤的生长。机制上，TEAD4 直接结合 MNX1-AS1 的启动子区域并刺激 MNX1-AS1 的转录。此外，MNX1-AS1 可以通过海绵 miR-6785-5p 来上调 GC 细胞中 BCL2 的表达。同时，MNX1-AS1 通过募集多梳抑制复合物 2 到 BTG2 启动子区域来抑制 BTG2 的转录。

结论：我们的研究结果表明，MNX1-AS1 可能能够作为 GC 患者的预后指标，并且 TEAD4 激活的 MNX1-AS1 可以通过 EZH2/BTG2 和 miR-6785-5p/BCL2 轴促进 GC 进展，表明它是治疗 GC 的一种新的有效靶点。