title: Functional Genomic Screening Reveals Splicing of the EWS-FLI1 Fusion Transcript as a Vulnerability in Ewing Sarcoma.
abstract: 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.
authors.affiliation: Department of Pediatrics, Vanderbilt University School of Medicine, Nashville, TN 37232, USA; Department of Pharmacology, Vanderbilt University School of Medicine, Nashville, TN 37232, USA; Department of Cancer Biology, Vanderbilt University School of Medicine, Nashville, TN 37232, USA.
authors.forename: Patrick J
authors.initials: PJ
authors.lastname: Grohar
authors.affiliation: Gene Silencing Section, Genetics Branch, Center for Cancer Research, National Cancer Institute, NIH, Bethesda, MD 20892, USA.
authors.forename: Suntae
authors.initials: S
authors.lastname: Kim
authors.affiliation: Gene Silencing Section, Genetics Branch, Center for Cancer Research, National Cancer Institute, NIH, Bethesda, MD 20892, USA; NIH Academy, Office of Intramural Training and Education, NIH, Bethesda, MD 20892, USA.
authors.forename: Guillermo O
authors.initials: GO
authors.lastname: Rangel Rivera
authors.affiliation: Gene Silencing Section, Genetics Branch, Center for Cancer Research, National Cancer Institute, NIH, Bethesda, MD 20892, USA.
authors.forename: Nirmalya
authors.initials: N
authors.lastname: Sen
authors.affiliation: Gene Silencing Section, Genetics Branch, Center for Cancer Research, National Cancer Institute, NIH, Bethesda, MD 20892, USA.
authors.forename: Sara
authors.initials: S
authors.lastname: Haddock
authors.affiliation: Department of Cancer Biology, Vanderbilt University School of Medicine, Nashville, TN 37232, USA.
authors.forename: Matt L
authors.initials: ML
authors.lastname: Harlow
authors.affiliation: Department of Pediatrics, Vanderbilt University School of Medicine, Nashville, TN 37232, USA.
authors.forename: Nichole K
authors.initials: NK
authors.lastname: Maloney
authors.affiliation: Molecular Genetics Section, Genetics Branch, Center for Cancer Research, National Cancer Institute, NIH, Bethesda, MD 20892, USA.
authors.forename: Jack
authors.initials: J
authors.lastname: Zhu
authors.affiliation: Protein Characterization Laboratory, Leidos Biomedical Research, Inc., Frederick National Laboratory for Cancer Research, Frederick, MD 21702, USA.
authors.forename: Maura
authors.initials: M
authors.lastname: O'Neill
authors.affiliation: Gene Silencing Section, Genetics Branch, Center for Cancer Research, National Cancer Institute, NIH, Bethesda, MD 20892, USA.
authors.forename: Tamara L
authors.initials: TL
authors.lastname: Jones
authors.affiliation: Gene Silencing Section, Genetics Branch, Center for Cancer Research, National Cancer Institute, NIH, Bethesda, MD 20892, USA.
authors.forename: Konrad
authors.initials: K
authors.lastname: Huppi
authors.affiliation: Gene Silencing Section, Genetics Branch, Center for Cancer Research, National Cancer Institute, NIH, Bethesda, MD 20892, USA.
authors.forename: Magdalena
authors.initials: M
authors.lastname: Grandin
authors.affiliation: Gene Silencing Section, Genetics Branch, Center for Cancer Research, National Cancer Institute, NIH, Bethesda, MD 20892, USA.
authors.forename: Kristen
authors.initials: K
authors.lastname: Gehlhaus
authors.affiliation: Trans-NIH RNAi Screening Facility, Division of Preclinical Innovation, National Center for Advancing Translational Sciences, NIH, Rockville, MD 20850, USA.
authors.forename: Carleen A
authors.initials: CA
authors.lastname: Klumpp-Thomas
authors.affiliation: Trans-NIH RNAi Screening Facility, Division of Preclinical Innovation, National Center for Advancing Translational Sciences, NIH, Rockville, MD 20850, USA.
authors.forename: Eugen
authors.initials: E
authors.lastname: Buehler
authors.affiliation: Molecular Oncology Section, Pediatric Oncology Branch, Center for Cancer Research, National Cancer Institute, NIH, Bethesda, MD 20892, USA.
authors.forename: Lee J
authors.initials: LJ
authors.lastname: Helman
authors.affiliation: Trans-NIH RNAi Screening Facility, Division of Preclinical Innovation, National Center for Advancing Translational Sciences, NIH, Rockville, MD 20850, USA.
authors.forename: Scott E
authors.initials: SE
authors.lastname: Martin
authors.affiliation: Gene Silencing Section, Genetics Branch, Center for Cancer Research, National Cancer Institute, NIH, Bethesda, MD 20892, USA. Electronic address: ncaplen@mail.nih.gov.
authors.forename: Natasha J
authors.initials: NJ
authors.lastname: Caplen
mesh_terms: D001483:Base Sequence; D001665:Binding Sites; D002148:Calmodulin-Binding Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D045744:Cell Line, Tumor; D002470:Cell Survival; D005091:Exons; D015972:Gene Expression Regulation, Neoplastic; D034541:Heterogeneous-Nuclear Ribonucleoprotein Group F-H / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D006801:Humans; D008840:Microfilament Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D015514:Oncogene Proteins, Fusion / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D010750:Phosphoproteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D051789:Proto-Oncogene Protein c-fli-1 / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D034622:RNA Interference; D012322:RNA Precursors / Q000378:metabolism; D012326:RNA Splicing; D000072260:RNA Splicing Factors; D034741:RNA, Small Interfering / Q000378:metabolism; D034802:RNA-Binding Protein EWS / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D016601:RNA-Binding Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D018160:Receptors, Cytoplasmic and Nuclear / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D017413:Ribonucleoprotein, U2 Small Nuclear / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D012512:Sarcoma, Ewing / Q000473:pathology; D015534:Trans-Activators; D014157:Transcription Factors / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism
chemical_list: D002148:Calmodulin-Binding Proteins; C106031:EWS-FLI fusion protein; C500986:EWSR1 protein, human; C098179:FLII protein, human; D034541:Heterogeneous-Nuclear Ribonucleoprotein Group F-H; D008840:Microfilament Proteins; D015514:Oncogene Proteins, Fusion; D010750:Phosphoproteins; D051789:Proto-Oncogene Protein c-fli-1; D012322:RNA Precursors; D000072260:RNA Splicing Factors; D034741:RNA, Small Interfering; D034802:RNA-Binding Protein EWS; D016601:RNA-Binding Proteins; D018160:Receptors, Cytoplasmic and Nuclear; D017413:Ribonucleoprotein, U2 Small Nuclear; C112491:SF3B1 protein, human; C100927:SUPT6H protein, human; D015534:Trans-Activators; D014157:Transcription Factors