Zhu Kai, Gao Tingting, Wang Zhiru, Zhang Liaoran, Tan Kezhe, Lv Zhibao
Department of General Surgery, Shanghai Children's Hospital, School of Medicine, Shanghai Jiao Tong University, Shanghai, PR China.
Cell Death Discov. 2023 May 8;9(1):151. doi: 10.1038/s41420-023-01449-3.
Neuroblastoma (NB) is a kind of typical life-threatening extracranial tumor in children. N6-methyladenosine (m6A) modification is closely related to multiple cancer pathological processes. Insulin-like growth factor 2 mRNA binding protein 3 (IGF2BP3) is a top-ranked prognostic risk gene in NB; however, its function is uncertain. The expression of m6A-associated enzymes in patients with NB was analyzed using the Gene Expression Omnibus (GEO) and Therapeutically Applicable Research to Generate Effective Treatments (TARGET) database. The IGF2BP3 level in NB cell lines and primary samples was tested using quantitative real-time polymerase chain reaction (qRT-PCR), western blot method, and immunohistochemical analysis. The IGF2BP3 function in cell proliferation was clarified based on many functional in vitro and in vivo experiments. The interaction between IGF2BP3 and N-myc was researched via RNA immunoprecipitation (RIP), m6A RNA immunoprecipitation (MeRIP), and chromatin immunoprecipitation (ChIP) assays. The 16 m6A-regulated enzymes in NB were researched, and the result indicated that IGF2BP3 overexpression was related to cancer progression, COG risk, and survival based on the GEO and TARGET databases. Besides, the IGF2BP3 and MYCN levels were positively correlated. IGF2BP3 expression levels increased in MYCN-amplified NB clinical samples and cells. Knockdown of IGF2BP3 inhibited N-myc expression and NB cell proliferation in vitro and in vivo. IGF2BP3 regulates MYCN RNA stability by modifying m6A. In addition, we demonstrated that N-myc is a transcription factor that directly promotes IGF2BP3 expression in NB cells. IGF2BP3 regulates the proliferation of NB cells via m6A modification of MYCN. N-myc also acts as a transcription factor that regulates IGF2BP3 expression. A positive feedback loop between IGF2BP3 and N-myc facilitates NB cell proliferation.
神经母细胞瘤（NB）是儿童中一种典型的危及生命的颅外肿瘤。N6-甲基腺苷（m6A）修饰与多种癌症病理过程密切相关。胰岛素样生长因子2 mRNA结合蛋白3（IGF2BP3）是NB中排名靠前的预后风险基因；然而，其功能尚不确定。利用基因表达综合数据库（GEO）和生成有效治疗方法的治疗应用研究（TARGET）数据库分析了NB患者中m6A相关酶的表达情况。采用定量实时聚合酶链反应（qRT-PCR）、蛋白质免疫印迹法和免疫组织化学分析检测了NB细胞系和原代样本中的IGF2BP3水平。基于许多体外和体内功能实验阐明了IGF2BP3在细胞增殖中的功能。通过RNA免疫沉淀（RIP）、m6A RNA免疫沉淀（MeRIP）和染色质免疫沉淀（ChIP）试验研究了IGF2BP3与N-myc之间的相互作用。对NB中的16种m6A调节酶进行了研究，结果表明，基于GEO和TARGET数据库，IGF2BP3的过表达与癌症进展、COG风险和生存相关。此外，IGF2BP3和MYCN水平呈正相关。在MYCN扩增的NB临床样本和细胞中，IGF2BP3表达水平升高。敲低IGF2BP3可在体外和体内抑制N-myc表达和NB细胞增殖。IGF2BP3通过修饰m6A调节MYCN RNA稳定性。此外，我们证明N-myc是一种转录因子，可直接促进NB细胞中IGF2BP3的表达。IGF2BP3通过对MYCN进行m6A修饰来调节NB细胞的增殖。N-myc还作为一种转录因子调节IGF2BP3的表达。IGF2BP3和N-myc之间的正反馈环促进了NB细胞的增殖。
