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 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 used 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：

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融合蛋白 EWS-FLI1 被整合到细胞内的蛋白颗粒中


尤文肉瘤由融合蛋白驱动，该融合蛋白含有本质上无序的低复杂性 (LC) 结构域和强大的转录调节因子。尤文肉瘤中最常见的融合蛋白 EWS-FLI1，其 LC 结构域取自 RNA 结合蛋白 EWSR1（尤文肉瘤 RNA 结合蛋白 1），DNA 结合结构域取自转录因子 FLI1（Friend 白血病病毒整合） 1). EWS-FLI1 可以结合 RNA 聚合酶 II (RNA Pol II) 并通过其 LC 结构域进行自组装。像 EWSR1 这样的 RNA 结合蛋白在细胞中自组装或相分离的能力引发了人们对该过程对 EWS-FLI1 活性的贡献的疑问。我们通过 siRNA 介导的敲低和 RNA-seq 分析检查了尤文肉瘤细胞中 EWSR1 和 EWS-FLI1 的活性。受 EWSR1 敲低影响的转录本比预期更多，其中包括许多 EWS-FLI1 调节基因。我们重新评估了 EWS-FLI1、EWSR1 和 RNA Pol II 之间的物理相互作用，并使用基于交联的策略来研究与蛋白质相关的蛋白质组装体。 EWS-FLI1 的 LC 结构域对于观察到在细胞中形成的组装是必需的。这些结果为蛋白质组装提供了新的见解，该蛋白质组装可能使 EWS-FLI1 能够结合其广泛的蛋白质伴侣网络，并有助于调节尤文肉瘤中的基因表达。