BACKGROUND The MYCN amplification is a defining hallmark of high-risk neuroblastoma. Due to irregular oncogenes orchestration, tumor cells exhibit distinct fatty acid metabolic features from non-tumor cells. However, the function of MYCN in neuroblastoma fatty acid metabolism reprogramming remains unknown. METHODS Gas Chromatography-Mass Spectrometer (GC-MS) was used to find the potential target fatty acid metabolites of MYCN. Real-time PCR (RT-PCR) and clinical bioinformatics analysis was used to find the related target genes. The function of the identified target gene ELOVL2 on cell growth was detected through CCK-8 assay, Soft agar colony formation assay, flow Cytometry assay and mouse xenograft. Chromatin immunoprecipitation (ChIP) and Immunoprecipitation-Mass Spectrometer (IP-MS) further identified the target gene and the co-repressor of MYCN. RESULTS The fatty acid profile of MYCN-depleted neuroblastoma cells identified docosahexaenoic acid (DHA), an omega-3 polyunsaturated fatty acid with anti-tumor activity, significantly increased after MYCN depletion. Compared with MYCN single-copy neuroblastoma cells, DHA level was significantly lower in MYCN-amplified neuroblastoma cells. RT-PCR and clinical bioinformatics analysis discovered that MYCN interfered DHA accumulation via ELOVL fatty acid elongase 2 (ELOVL2) which is a rate-limiting enzyme of cellular DHA synthesis. Enforced ELOVL2 expression in MYCN-amplified neuroblastoma cells led to decreased cell growth and counteracted the growth-promoting effect of MYCN overexpression both in vitro and vivo. ELOVL2 Knockdown showed the opposite effect in MYCN single-copy neuroblastoma cells. In primary neuroblastoma, high ELOVL2 transcription correlated with favorable clinical tumor biology and patient survival. The mechanism of MYCN-mediated ELOVL2 inhibition contributed to epigenetic regulation. MYCN recruited PRC1 (Polycomb repressive complex 1), catalysed H2AK119ub (histone 2A lysine 119 monoubiquitination) and inhibited subsequent ELOVL2 transcription. CONCLUSIONS The tumor suppressive properties of DHA and ELOVL2 are repressed by the MYCN and PRC1 jointly, which suggests a new epigenetic mechanism of MYCN-mediated fatty acid regulation and indicates PRC1 inhibition as a potential novel strategy to activate ELOVL2 suppressive functions.

中文翻译：

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MYCN和PRC1通过ELOVL2协同抑制神经母细胞瘤中二十二碳六烯酸的合成。

背景技术MYCN扩增是高危神经母细胞瘤的标志性特征。由于不规则的致癌基因编排，肿瘤细胞表现出与非肿瘤细胞不同的脂肪酸代谢特征。但是，MYCN在神经母细胞瘤脂肪酸代谢重编程中的功能仍然未知。方法采用气相色谱-质谱仪（GC-MS）寻找MYCN潜在的目标脂肪酸代谢产物。实时荧光定量PCR（RT-PCR）和临床生物信息学分析被用于寻找相关的靶基因。通过CCK-8试验，软琼脂菌落形成试验，流式细胞术和小鼠异种移植物检测所鉴定的靶基因ELOVL2对细胞生长的功能。染色质免疫沉淀（ChIP）和免疫沉淀质谱仪（IP-MS）进一步鉴定了目标基因和MYCN的共阻遏物。结果贫MYCN的成神经细胞瘤细胞的脂肪酸谱鉴定为二十二碳六烯酸（DHA），一种具有抗肿瘤活性的ω-3多不饱和脂肪酸，在MYCN消耗后显着增加。与MYCN单拷贝神经母细胞瘤细胞相比，MYCN扩增的神经母细胞瘤细胞中的DHA水平显着降低。RT-PCR和临床生物信息学分析发现，MYCN通过ELOVL脂肪酸延伸酶2（ELOVL2）干扰DHA积累，ELOVL 2是细胞DHA合成的限速酶。MYCN扩增的神经母细胞瘤细胞中增强的ELOVL2表达导致细胞生长减少，并抵消了在体外和体内均过表达MYCN的促进生长的作用。ELOVL2抑制在MYCN单拷贝神经母细胞瘤细胞中显示相反的作用。在原发性神经母细胞瘤中，高ELOVL2转录与良好的临床肿瘤生物学和患者生存率相关。MYCN介导的ELOVL2抑制的机制有助于表观遗传调控。MYCN募集了PRC1（多聚体阻抑复合物1），催化了H2AK119ub（组蛋白2A赖氨酸119单泛素化）并抑制了随后的ELOVL2转录。结论MYCN和PRC1共同抑制DHA和ELOVL2的抑癌特性，