Liaison between SNAI2 and MYOD enhances oncogenesis and suppresses differentiation in Fusion-Negative Rhabdomyosarcoma. Liaison between SNAI2 and MYOD enhances oncogenesis and suppresses differentiation in Fusion-Negative Rhabdomyosarcoma

Rhabdomyosarcoma (RMS) is a pediatric malignancy of mesenchymal origin. Fusion Negative-RMS (FN-RMS) tumors are associated with RAS-pathway activation. RMS tumors express pro-differentiation myogenic transcription factors MYOD and MYOG, yet why they are unable to differentiate is poorly understood. Here we show that SNAI2 is highly expressed in FN-RMS, is regulated by MYOD and blocks myogenic differentiation promoting growth. Molecularly, SNAI2 preferentially binds E-Box-associated enhancer elements and represses expression by dampening enhancer function. SNAI2 inhibits MYOD at a subset of myogenic enhancers associated with terminal differentiation. Functional dissection demonstrates SNAI2 suppresses a MYOG, MEF2 and CDKN1A differentiation program. SNAI2 knockdown transcriptionally mimics a chemical blockade of the mutant RAS signal in FN-RMS, providing new insight connecting the genetic and epigenetic causes of this disease. Overall design: Epignome-wide binding profiles of histone modifications and transcription factors, before and after selected disruption of SNAI2 via chemical or genetic means, in FN_RMS cell lines, by ChIP-seq. RNA-seq in 2 FN-RMS cell lines exposed to either shControl or shSNAI2. H3K27ac HiChIP of RMS cell line.

横纹肌肉瘤（Rhabdomyosarcoma, RMS）是一种起源于间叶组织的儿童恶性肿瘤。融合基因阴性横纹肌肉瘤（Fusion Negative-RMS, FN-RMS）与RAS通路激活密切相关。RMS肿瘤表达促分化肌源性转录因子MYOD与MYOG，但其为何无法完成分化，目前其机制仍未被充分阐明。本研究证实，SNAI2在FN-RMS中高表达，受MYOD调控，并通过阻断肌源性分化促进肿瘤生长。分子机制层面，SNAI2优先结合E盒相关增强子元件，并通过削弱增强子功能抑制基因表达。SNAI2可在一组与终末分化相关的肌源性增强子位点上抑制MYOD的活性。功能解析实验证实，SNAI2可抑制由MYOG、MEF2及CDKN1A介导的分化程序。敲低SNAI2可在转录层面模拟FN-RMS中突变RAS信号的化学阻断效应，为阐明该疾病的遗传与表观遗传致病机制提供了全新视角。整体实验设计：在FN-RMS细胞系中，通过化学或遗传学手段干扰SNAI2表达前后，采用染色质免疫共沉淀测序（ChIP-seq）检测表观基因组范围内的组蛋白修饰与转录因子结合谱；在2株FN-RMS细胞系中分别转染短发夹RNA对照（shControl）与靶向SNAI2的短发夹RNA（shSNAI2），开展RNA测序（RNA-seq）；并对RMS细胞系进行H3K27乙酰化染色质构象捕获测序（H3K27ac HiChIP）。