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Fusion protein EWS-FLI1 is incorporated into a protein granule in cells
bioRxiv - Biochemistry Pub Date : 2021-05-05 , DOI: 10.1101/2020.05.31.122028
Nasiha S. Ahmed , Lucas M. Harrell , Daniel R. Wieland , Michelle A. Lay , Valery F. Thompson , Jacob C. Schwartz

Ewing sarcoma is driven by fusion proteins containing a low complexity (LC) domain that is intrinsically disordered and a powerful transcriptional regulator. The most common fusion protein found in Ewing sarcoma, EWS-FLI1, takes its LC domain from the RNA-binding protein EWSR1 (Ewing Sarcoma RNA-binding protein 1) and a DNA-binding domain from the transcription factor FLI1 (Friend Leukemia Virus Integration 1). EWS-FLI1 can bind RNA polymerase II (RNA Pol II) and self-assemble through its low-complexity (LC) domain. The ability of RNA-binding proteins like EWSR1 to self-assemble or phase separate in cells has raised questions about the contribution of this process to EWS-FLI1 activity. We examined EWSR1 and EWS-FLI1 activity in Ewing sarcoma cells by siRNA-mediated knockdown and RNA-seq analysis. More transcripts were affected by the EWSR1 knockdown than expected and these included many EWS-FLI1 regulated genes. We reevaluated physical interactions between EWS-FLI1, EWSR1, and RNA Pol II, and employed a cross-linking based strategy to investigate protein assemblies associated with the proteins. The LC domain of EWS-FLI1 was required for the assemblies observed to form in cells. These results offer new insights into a protein assembly that may enable EWS-FLI1 to bind its wide network of protein partners and contribute to regulation of gene expression in Ewing sarcoma.

中文翻译:

融合蛋白EWS-FLI1被掺入细胞的蛋白颗粒中

尤文氏肉瘤是由融合蛋白驱动的,该融合蛋白包含固有紊乱的低复杂度(LC)域和强大的转录调节因子。在尤文氏肉瘤中发现的最常见的融合蛋白EWS-FLI1取自RNA结合蛋白EWSR1的LC结构域(尤文氏肉瘤RNA结合蛋白1)和转录因子FLI1的DNA结合结构域(Friend Leukemia Virus Integration) 1)。EWS-FLI1可以结合RNA聚合酶II(RNA Pol II)并通过其低复杂度(LC)域自组装。RNA结合蛋白(如EWSR1)在细胞中自组装或相分离的能力提出了有关此过程对EWS-FLI1活性的贡献的疑问。我们通过siRNA介导的敲低和RNA序列分析检查了尤因肉瘤细胞中的EWSR1和EWS-FLI1活性。EWSR1敲除对转录物的影响比预期的要多,其中包括许多EWS-FLI1调控的基因。我们重新评估了EWS-FLI1,EWSR1和RNA Pol II之间的物理相互作用,并采用了基于交联的策略来研究与蛋白质相关的蛋白质装配。EWS-FLI1的LC结构域是观察到在细胞中形成的装配所必需的。这些结果为蛋白质装配提供了新的见解,可能使EWS-FLI1结合其广泛的蛋白质伴侣网络并有助于调节尤因肉瘤中的基因表达。EWS-FLI1的LC结构域是观察到在细胞中形成的装配所必需的。这些结果为蛋白质装配提供了新的见解,可能使EWS-FLI1结合其广泛的蛋白质伴侣网络并有助于调节尤因肉瘤中的基因表达。EWS-FLI1的LC结构域是观察到在细胞中形成的装配所必需的。这些结果为蛋白质装配提供了新的见解,可能使EWS-FLI1结合其广泛的蛋白质伴侣网络并有助于调节尤因肉瘤中的基因表达。
更新日期:2021-05-06
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