Table_3_LncRNA FOXD3-AS1 Promotes the Malignant Progression of Nasopharyngeal Carcinoma Through Enhancing the Transcription of YBX1 by H3K27Ac Modification.doc

BackgroundLong noncoding RNAs (lncRNAs) can affect the progression of various tumors, including nasopharyngeal carcinoma (NPC). Here, lncRNA FOXD3-AS1 is highly expressed in NPC tissues through bioinformatics analysis and related to the malignant progression of NPC.
MethodsBioinformatics analysis and real-time reverse transcription quantitative PCR(RT-qPCR) assay were applied to identify the expression of FOXD3-AS1 in NPC tissues and cells. Specific short hairpin RNAs (shRNAs) or overexpression plasmids were used to knockdown or upregulate FOXD3-AS1 in NPC cells. The effect of FOXD3-AS1 on proliferation and metastasis of NPC was confirmed by CCK8, colony formation, transwell assays in vitro and mouse tumor growth and metastasis models in vivo, of which the mechanism was explored by RNA pull down, mass spectrometry (MS), RNA Immunoprecipitation (RIP), chromatin immunoprecipitation (CHIP) and luciferase assays.
ResultsFOXD3-AS1 was highly expressed in NPC tissues and cells. Knockdown of FOXD3-AS1 significantly inhibited proliferation, migration, and invasion of NPC cells in vitro and vivo. FOXD3-AS1 could specifically bind to YBX1 and have a positive effect on the expression of YBX1. Bioinformatics analysis showed that the promoter of YBX1 had a high enrichment of H3K27ac, which promote mRNA transcription and protein translation of YBX1. Moreover, overexpression of YBX1 could reverse the proliferation, migration and invasion arrest caused by FOXD3-AS1 knockdown.
ConclusionLncRNA FOXD3-AS1 is highly expressed and promotes malignant phenotype in NPC, which may provide a new molecular mechanism for NPC.

背景：长链非编码RNA（long noncoding RNAs, lncRNAs）可影响包括鼻咽癌（nasopharyngeal carcinoma, NPC）在内的多种肿瘤的进展。本研究通过生物信息学分析发现，长链非编码RNA FOXD3-AS1在鼻咽癌组织中呈高表达，且与鼻咽癌的恶性进展密切相关。
方法：本研究采用生物信息学分析及实时逆转录定量聚合酶链反应（real-time reverse transcription quantitative PCR, RT-qPCR）检测FOXD3-AS1在鼻咽癌组织及细胞中的表达水平。通过特异性短发夹RNA（short hairpin RNAs, shRNAs）或过表达质粒分别敲低或上调鼻咽癌细胞中FOXD3-AS1的表达。采用CCK8实验、平板克隆形成实验、Transwell实验体外验证FOXD3-AS1对鼻咽癌细胞增殖及转移能力的影响，并通过小鼠肿瘤生长与转移模型开展体内验证；同时借助RNA pull-down实验、质谱分析（mass spectrometry, MS）、RNA免疫沉淀（RNA Immunoprecipitation, RIP）、染色质免疫沉淀（chromatin immunoprecipitation, CHIP）及荧光素酶报告基因实验探索其潜在分子机制。
结果：FOXD3-AS1在鼻咽癌组织及细胞中呈高表达。敲低FOXD3-AS1可显著抑制鼻咽癌细胞的体外增殖、迁移与侵袭能力，该效应在体内实验中同样得到验证。FOXD3-AS1可特异性结合YBX1蛋白，并对YBX1的表达具有正向调控作用。生物信息学分析显示，YBX1的启动子区域存在H3K27ac的高富集修饰，该修饰可促进YBX1的mRNA转录及蛋白翻译过程。此外，过表达YBX1可逆转FOXD3-AS1敲低所导致的细胞增殖、迁移及侵袭受阻表型。
结论：长链非编码RNA FOXD3-AS1在鼻咽癌中呈高表达，并可促进鼻咽癌的恶性表型，该研究结果可为鼻咽癌的诊疗提供全新的分子机制参考。