Selig Emily E, Sohn Erich J, Stoja Aiola, Moreno-Romero Alma K, Akula Shivani, Xu Xiaoping, Bishop Alexander J R, Libich David S
Greehey Children's Cancer Research Institute, The University of Texas Health Science Center at San Antonio, San Antonio, TX 78229.
Department of Biochemistry and Structural Biology, The University of Texas Health Science Center at San Antonio, San Antonio, TX 78229.
Proc Natl Acad Sci U S A. 2025 May 20;122(20):e2221823122. doi: 10.1073/pnas.2221823122. Epub 2025 May 16.
Ewing sarcoma (EwS) is an aggressive cancer of bone and soft tissue that predominantly affects children and young adults. A chromosomal translocation joins the low-complexity domain (LCD) of the RNA-binding protein EWS (EWS) with the DNA-binding domain of Friend leukemia integration 1 (FLI1), creating EWS::FLI1, a potent fusion oncoprotein essential for EwS development and responsible for over 85% of EwS tumors. EWS::FLI1 forms biomolecular condensates in vivo and promotes tumorigenesis through mediation of aberrant transcriptional changes and by interfering with the normal functions of nucleic acid-binding proteins like EWS through a dominant-negative mechanism. In particular, the expression of EWS::FLI1 in EwS directly interferes with the biological functions of EWS leading to alternate splicing events and defects in DNA-damage repair pathways. Though the EWS is capable of phase separation, here we report a direct interaction between FLI1 and EWS that enhances condensate formation and alters the physical properties of the condensate. This effect was conserved for three related E-twenty-six transformation-specific (ETS) DNA-binding domains (DBDs) while DNA binding blocked the interaction with EWS and inhibited EWS::FLI1 condensate formation. NMR spectroscopy and mutagenesis studies confirmed that ETS DBDs transiently interact with EWS via the ETS DBDs "wings." Together these results revealed that ETS DBDs, particularly FLI1, enhance EWS condensate formation and rigidity, supporting a model in which electrostatic and structural interactions drive condensate dynamics with implications for EWS::FLI1-mediated transcriptional regulation in EwS.
尤因肉瘤（EwS）是一种侵袭性的骨与软组织癌症，主要影响儿童和年轻人。一种染色体易位将RNA结合蛋白EWS（EWS）的低复杂性结构域（LCD）与Friend白血病整合1（FLI1）的DNA结合结构域连接起来，产生EWS::FLI1，这是一种对EwS发展至关重要的强效融合癌蛋白，占EwS肿瘤的85%以上。EWS::FLI1在体内形成生物分子凝聚物，并通过介导异常转录变化以及通过显性负性机制干扰像EWS这样的核酸结合蛋白的正常功能来促进肿瘤发生。特别是，EWS::FLI1在EwS中的表达直接干扰EWS的生物学功能，导致可变剪接事件和DNA损伤修复途径缺陷。虽然EWS能够发生相分离，但我们在此报告FLI1与EWS之间的直接相互作用增强了凝聚物形成并改变了凝聚物的物理性质。这种效应在三个相关的E-26转化特异性（ETS）DNA结合结构域（DBD）中是保守的，而DNA结合则阻断了与EWS的相互作用并抑制了EWS::FLI1凝聚物形成。核磁共振光谱和诱变研究证实，ETS DBD通过ETS DBD的“侧翼”与EWS短暂相互作用。这些结果共同表明，ETS DBD，特别是FLI1，增强了EWS凝聚物的形成和刚性，支持了一种模型，即静电和结构相互作用驱动凝聚物动力学，这对EwS中EWS::FLI1介导的转录调控具有重要意义。
