Functional Genomic Screening Reveals Splicing of the EWS-FLI1 Fusion Transcript as a Vulnerability in Ewing Sarcoma
Patrick J. Grohar,
Suntae Kim,
Guillermo O. Rangel Rivera,
Nirmalya Sen,
Sara Haddock,
Matt L. Harlow,
Nichole K. Maloney,
Jack Zhu,
Maura O’Neill,
Tamara L. Jones,
Konrad Huppi,
Magdalena Grandin,
Kristen Gehlhaus,
Carleen A. Klumpp-Thomas,
Eugen Buehler,
Lee J. Helman,
Scott E. Martin,
Natasha J. Caplen
Affiliations
Patrick J. Grohar
Department of Pediatrics, Vanderbilt University School of Medicine, Nashville, TN 37232, USA
Suntae Kim
Gene Silencing Section, Genetics Branch, Center for Cancer Research, National Cancer Institute, NIH, Bethesda, MD 20892, USA
Guillermo O. Rangel Rivera
Gene Silencing Section, Genetics Branch, Center for Cancer Research, National Cancer Institute, NIH, Bethesda, MD 20892, USA
Nirmalya Sen
Gene Silencing Section, Genetics Branch, Center for Cancer Research, National Cancer Institute, NIH, Bethesda, MD 20892, USA
Sara Haddock
Gene Silencing Section, Genetics Branch, Center for Cancer Research, National Cancer Institute, NIH, Bethesda, MD 20892, USA
Matt L. Harlow
Department of Cancer Biology, Vanderbilt University School of Medicine, Nashville, TN 37232, USA
Nichole K. Maloney
Department of Pediatrics, Vanderbilt University School of Medicine, Nashville, TN 37232, USA
Jack Zhu
Molecular Genetics Section, Genetics Branch, Center for Cancer Research, National Cancer Institute, NIH, Bethesda, MD 20892, USA
Maura O’Neill
Protein Characterization Laboratory, Leidos Biomedical Research, Inc., Frederick National Laboratory for Cancer Research, Frederick, MD 21702, USA
Tamara L. Jones
Gene Silencing Section, Genetics Branch, Center for Cancer Research, National Cancer Institute, NIH, Bethesda, MD 20892, USA
Konrad Huppi
Gene Silencing Section, Genetics Branch, Center for Cancer Research, National Cancer Institute, NIH, Bethesda, MD 20892, USA
Magdalena Grandin
Gene Silencing Section, Genetics Branch, Center for Cancer Research, National Cancer Institute, NIH, Bethesda, MD 20892, USA
Kristen Gehlhaus
Gene Silencing Section, Genetics Branch, Center for Cancer Research, National Cancer Institute, NIH, Bethesda, MD 20892, USA
Carleen A. Klumpp-Thomas
Trans-NIH RNAi Screening Facility, Division of Preclinical Innovation, National Center for Advancing Translational Sciences, NIH, Rockville, MD 20850, USA
Eugen Buehler
Trans-NIH RNAi Screening Facility, Division of Preclinical Innovation, National Center for Advancing Translational Sciences, NIH, Rockville, MD 20850, USA
Lee J. Helman
Molecular Oncology Section, Pediatric Oncology Branch, Center for Cancer Research, National Cancer Institute, NIH, Bethesda, MD 20892, USA
Scott E. Martin
Trans-NIH RNAi Screening Facility, Division of Preclinical Innovation, National Center for Advancing Translational Sciences, NIH, Rockville, MD 20850, USA
Natasha J. Caplen
Gene Silencing Section, Genetics Branch, Center for Cancer Research, National Cancer Institute, NIH, Bethesda, MD 20892, USA
Ewing sarcoma cells depend on the EWS-FLI1 fusion transcription factor for cell survival. Using an assay of EWS-FLI1 activity and genome-wide RNAi screening, we have identified proteins required for the processing of the EWS-FLI1 pre-mRNA. We show that Ewing sarcoma cells harboring a genomic breakpoint that retains exon 8 of EWSR1 require the RNA-binding protein HNRNPH1 to express in-frame EWS-FLI1. We also demonstrate the sensitivity of EWS-FLI1 fusion transcripts to the loss of function of the U2 snRNP component, SF3B1. Disrupted splicing of the EWS-FLI1 transcript alters EWS-FLI1 protein expression and EWS-FLI1-driven expression. Our results show that the processing of the EWS-FLI1 fusion RNA is a potentially targetable vulnerability in Ewing sarcoma cells.
