Fusion protein EWS-FLI1 is incorporated into a protein granule in cells
- Nasiha S. Ahmed1,2,
- Lucas M. Harrell2,
- Daniel R. Wieland2,
- Michelle A. Lay2,
- Valery F. Thompson2 and
- Jacob C. Schwartz2
- 1Department of Molecular and Cellular Biology, The University of Arizona, Tucson, Arizona 85719, USA
- 2Department of Chemistry and Biochemistry, The University of Arizona, Tucson, Arizona 85719, USA
- Corresponding author: jcschwartz{at}email.arizona.edu
Abstract
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.
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Footnotes
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Article is online at http://www.rnajournal.org/cgi/doi/10.1261/rna.078827.121.
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Freely available online through the RNA Open Access option.
- Received May 5, 2021.
- Accepted May 18, 2021.
This article, published in RNA, is available under a Creative Commons License (Attribution 4.0 International), as described at http://creativecommons.org/licenses/by/4.0/.