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Abstract：
Oncogene MYCN is closely related with malignant progression and poor prognosis of neuroblastoma (NB). Recently, long non-coding RNAs (lncRNAs) have been recognized as crucial regulators in various cancers. However, whether lncRNAs contribute to the overexpression of MYCN in NB is unclear.
Microarray analysis were applied to analyze the differentially expressed lncRNAs between MYCN-amplified and MYCN-non-amplified NB cell lines. Bioinformatic analyses were utilized to identify lncRNAs nearby MYCN locus. qRT-PCR was used to detect the expression level of lncRNA AC142119.1 in NB cell lines and tissues. Gain- and loss-of-function assays were conducted to investigate the biological effect of AC142119.1 in NB. Fluorescence in situ hybridization, RNA pull-down, RNA immunoprecipitation, mass spectrometry, RNA electrophoretic mobility shift, chromatin immunoprecipitation and chromatin isolation by RNA purification assays were performed to validate the interaction between AC142119.1 and WDR5 protein as well as MYCN promoter.
AC142119.1 was significantly elevated in NB tissues with MYCN amplification, advanced INSS stage and high risk, and associated with poor survival of NB patients. Moreover, enforced expression of AC142119.1 reinforced the proliferation of NB cells in vitro and in vivo. Additionally, AC142119.1 specifically recruited WDR5 protein to interact with MYCN promoter, further initiating the transcription of MYCN and accelerating NB progression.
We identified a novel lncRNA AC142119.1, which promoted the progression of NB through epigenetically initiating the transcription of MYCN via interacting with both WDR5 protein and the promoter of MYCN, indicating that AC142119.1 might be a potential diagnostic biomarker and therapeutic target for NB.
Microarray analysis were applied to analyze the differentially expressed lncRNAs between MYCN-amplified and MYCN-non-amplified NB cell lines. Bioinformatic analyses were utilized to identify lncRNAs nearby MYCN locus. qRT-PCR was used to detect the expression level of lncRNA AC142119.1 in NB cell lines and tissues. Gain- and loss-of-function assays were conducted to investigate the biological effect of AC142119.1 in NB. Fluorescence in situ hybridization, RNA pull-down, RNA immunoprecipitation, mass spectrometry, RNA electrophoretic mobility shift, chromatin immunoprecipitation and chromatin isolation by RNA purification assays were performed to validate the interaction between AC142119.1 and WDR5 protein as well as MYCN promoter.
AC142119.1 was significantly elevated in NB tissues with MYCN amplification, advanced INSS stage and high risk, and associated with poor survival of NB patients. Moreover, enforced expression of AC142119.1 reinforced the proliferation of NB cells in vitro and in vivo. Additionally, AC142119.1 specifically recruited WDR5 protein to interact with MYCN promoter, further initiating the transcription of MYCN and accelerating NB progression.
We identified a novel lncRNA AC142119.1, which promoted the progression of NB through epigenetically initiating the transcription of MYCN via interacting with both WDR5 protein and the promoter of MYCN, indicating that AC142119.1 might be a potential diagnostic biomarker and therapeutic target for NB.
摘要：
背景：癌基因MYCN与神经母细胞瘤（NB）的恶性进展和不良预后密切相关。最近,长链非编码RNA（lncRNAs）已被认为是各种癌症的关键调节因子。然而,lncRNAs是否有助于NB中MYCN的过表达尚不清楚.
方法：微阵列分析用于分析MYCN扩增和MYCN非扩增NB细胞系之间差异表达的lncRNAs。生物信息学分析用于鉴定MYCN基因座附近的lncRNA。qRT-PCR检测lncRNAAC142119.1在NB细胞系和组织中的表达水平。进行了功能增益和功能丧失试验以研究AC142119.1在NB中的生物学效应。荧光原位杂交,RNA下拉，RNA免疫沉淀，质谱,RNA电泳迁移率变化，通过RNA纯化测定进行染色质免疫沉淀和染色质分离以验证AC142119.1和WDR5蛋白以及MYCN启动子之间的相互作用。
结果：在MYCN扩增的NB组织中AC142119.1显著升高，高级INSS阶段和高风险，并与NB患者的低生存率相关。此外，AC142119.1的强制表达增强了NB细胞的体外和体内增殖。此外,AC142119.1专门招募WDR5蛋白与MYCN启动子相互作用，进一步启动MYCN的转录并加速NB的进展。
结论：我们鉴定了一种新的lncRNAAC142119.1，它通过与WDR5蛋白和MYCN启动子相互作用，通过表观遗传学启动MYCN的转录促进NB的进展，表明AC142119.1可能是NB的潜在诊断生物标志物和治疗靶标。
方法：微阵列分析用于分析MYCN扩增和MYCN非扩增NB细胞系之间差异表达的lncRNAs。生物信息学分析用于鉴定MYCN基因座附近的lncRNA。qRT-PCR检测lncRNAAC142119.1在NB细胞系和组织中的表达水平。进行了功能增益和功能丧失试验以研究AC142119.1在NB中的生物学效应。荧光原位杂交,RNA下拉，RNA免疫沉淀，质谱,RNA电泳迁移率变化，通过RNA纯化测定进行染色质免疫沉淀和染色质分离以验证AC142119.1和WDR5蛋白以及MYCN启动子之间的相互作用。
结果：在MYCN扩增的NB组织中AC142119.1显著升高，高级INSS阶段和高风险，并与NB患者的低生存率相关。此外，AC142119.1的强制表达增强了NB细胞的体外和体内增殖。此外,AC142119.1专门招募WDR5蛋白与MYCN启动子相互作用，进一步启动MYCN的转录并加速NB的进展。
结论：我们鉴定了一种新的lncRNAAC142119.1，它通过与WDR5蛋白和MYCN启动子相互作用，通过表观遗传学启动MYCN的转录促进NB的进展，表明AC142119.1可能是NB的潜在诊断生物标志物和治疗靶标。
