title: MYCN controls an alternative RNA splicing program in high-risk metastatic neuroblastoma.
abstract: The molecular mechanisms underlying the aggressive behavior of MYCN driven neuroblastoma (NBL) is under intense investigation; however, little is known about the impact of this family of transcription factors on the splicing program. Here we used high-throughput RNA sequencing to systematically study the expression of RNA isoforms in stage 4 MYCN-amplified NBL, an aggressive subtype of metastatic NBL. We show that MYCN-amplified NBL tumors display a distinct gene splicing pattern affecting multiple cancer hallmark functions. Six splicing factors displayed unique differential expression patterns in MYCN-amplified tumors and cell lines, and the binding motifs for some of these splicing factors are significantly enriched in differentially-spliced genes. Direct binding of MYCN to promoter regions of the splicing factors PTBP1 and HNRNPA1 detected by ChIP-seq demonstrates that MYCN controls the splicing pattern by direct regulation of the expression of these key splicing factors. Furthermore, high expression of PTBP1 and HNRNPA1 was significantly associated with poor overall survival of stage4 NBL patients (p ≤ 0.05). Knocking down PTBP1, HNRNPA1 and their downstream target PKM2, an isoform of pro-tumor-growth, result in repressed growth of NBL cells. Therefore, our study reveals a novel role of MYCN in controlling global splicing program through regulation of splicing factors in addition to its well-known role in the transcription program. These findings suggest a therapeutically potential to target the key splicing factors or gene isoforms in high-risk NBL with MYCN-amplification.
authors.affiliation: Oncogenomics Section, Genetics Branch, National Cancer Institute, National Institute of Health, Bethesda, MD 20892, USA; Program in Bioinformatics, Boston University, Boston, MA 02218, USA.
authors.forename: Shile
authors.initials: S
authors.lastname: Zhang
authors.affiliation: Oncogenomics Section, Genetics Branch, National Cancer Institute, National Institute of Health, Bethesda, MD 20892, USA.
authors.forename: Jun S
authors.initials: JS
authors.lastname: Wei
authors.affiliation: Oncogenomics Section, Genetics Branch, National Cancer Institute, National Institute of Health, Bethesda, MD 20892, USA.
authors.forename: Samuel Q
authors.initials: SQ
authors.lastname: Li
authors.affiliation: Oncogenomics Section, Genetics Branch, National Cancer Institute, National Institute of Health, Bethesda, MD 20892, USA; Pediatric Hematology and Oncology, Kentucky Children's Hospital, Lexington, KY 40536, USA.
authors.forename: Tom C
authors.initials: TC
authors.lastname: Badgett
authors.affiliation: Oncogenomics Section, Genetics Branch, National Cancer Institute, National Institute of Health, Bethesda, MD 20892, USA.
authors.forename: Young K
authors.initials: YK
authors.lastname: Song
authors.affiliation: Texas Children's Cancer Center, Center for Cell and Gene Therapy, Department of Pediatrics, Baylor College of Medicine, Houston, TX 77030, USA.
authors.forename: Saurabh
authors.initials: S
authors.lastname: Agarwal
authors.affiliation: Texas Children's Cancer Center, Center for Cell and Gene Therapy, Department of Pediatrics, Baylor College of Medicine, Houston, TX 77030, USA.
authors.forename: Cristian
authors.initials: C
authors.lastname: Coarfa
authors.affiliation: Oncogenomics Section, Genetics Branch, National Cancer Institute, National Institute of Health, Bethesda, MD 20892, USA.
authors.forename: Catherine
authors.initials: C
authors.lastname: Tolman
authors.affiliation: Oncogenomics Section, Genetics Branch, National Cancer Institute, National Institute of Health, Bethesda, MD 20892, USA.
authors.forename: Laura
authors.initials: L
authors.lastname: Hurd
authors.affiliation: Oncogenomics Section, Genetics Branch, National Cancer Institute, National Institute of Health, Bethesda, MD 20892, USA.
authors.forename: Hongling
authors.initials: H
authors.lastname: Liao
authors.affiliation: Oncogenomics Section, Genetics Branch, National Cancer Institute, National Institute of Health, Bethesda, MD 20892, USA.
authors.forename: Jianbin
authors.initials: J
authors.lastname: He
authors.affiliation: Oncogenomics Section, Genetics Branch, National Cancer Institute, National Institute of Health, Bethesda, MD 20892, USA.
authors.forename: Xinyu
authors.initials: X
authors.lastname: Wen
authors.affiliation: Cell & Molecular Biology Section, Pediatric Oncology Branch, National Cancer Institute, National Institute of Health, Bethesda, MD 20892, USA.
authors.forename: Zhihui
authors.initials: Z
authors.lastname: Liu
authors.affiliation: Cell & Molecular Biology Section, Pediatric Oncology Branch, National Cancer Institute, National Institute of Health, Bethesda, MD 20892, USA.
authors.forename: Carol J
authors.initials: CJ
authors.lastname: Thiele
authors.affiliation: Neuroblastoma Genomics, B030, German Cancer Research Center, Im Neuenheimer Feld 280, 69120 Heidelberg, Germany.
authors.forename: Frank
authors.initials: F
authors.lastname: Westermann
authors.affiliation: Division of Hematology/Oncology, The Children's Hospital Los Angeles, Los Angeles, CA 90027, USA; Saban Research Institute, The Children's Hospital Los Angeles, Los Angeles, CA 90027, USA; Keck School of Medicine, University of Southern California, Los Angeles, CA 90089, USA.
authors.forename: Shahab
authors.initials: S
authors.lastname: Asgharzadeh
authors.affiliation: Division of Hematology/Oncology, The Children's Hospital Los Angeles, Los Angeles, CA 90027, USA; Saban Research Institute, The Children's Hospital Los Angeles, Los Angeles, CA 90027, USA; Keck School of Medicine, University of Southern California, Los Angeles, CA 90089, USA.
authors.forename: Robert C
authors.initials: RC
authors.lastname: Seeger
authors.affiliation: Center for Childhood Cancer Research, the Children's Hospital of Philadelphia, Philadelphia, PA 19104, USA; Division of Oncology, the Children's Hospital of Philadelphia, Philadelphia, PA 19104, USA; Department of Pediatrics, Perelman School of Medicine at the University of Pennsylvania, Philadelphia, PA 19104, USA; Abramson Family Cancer Research Institute, Philadelphia, PA 19104, USA.
authors.forename: John M
authors.initials: JM
authors.lastname: Maris
authors.affiliation: Office of Cancer Genomics, National Cancer Institute, Bethesda, MD 20892, USA.
authors.forename: Jamie M
authors.initials: JM
authors.lastname: Guidry Auvil
authors.affiliation: Clinical Investigation Branch, National Cancer Institute, Rockville, MD 20850, USA.
authors.forename: Malcolm A
authors.initials: MA
authors.lastname: Smith
authors.affiliation: Program in Bioinformatics, Boston University, Boston, MA 02218, USA; Department of Mathematics & Statistics, Boston University, Boston, MA 02218, USA.
authors.forename: Eric D
authors.initials: ED
authors.lastname: Kolaczyk
authors.affiliation: Texas Children's Cancer Center, Center for Cell and Gene Therapy, Department of Pediatrics, Baylor College of Medicine, Houston, TX 77030, USA.
authors.forename: Jason
authors.initials: J
authors.lastname: Shohet
authors.affiliation: Oncogenomics Section, Genetics Branch, National Cancer Institute, National Institute of Health, Bethesda, MD 20892, USA. Electronic address: khanjav@mail.nih.gov.
authors.forename: Javed
authors.initials: J
authors.lastname: Khan
mesh_terms: D017398:Alternative Splicing; D001665:Binding Sites; D014408:Biomarkers, Tumor / Q000235:genetics* / Q000378:metabolism; D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism; D045744:Cell Line, Tumor; D049109:Cell Proliferation; D047369:Chromatin Immunoprecipitation; D005784:Gene Amplification; D015972:Gene Expression Regulation, Neoplastic; D000076245:Heterogeneous Nuclear Ribonucleoprotein A1; D034481:Heterogeneous-Nuclear Ribonucleoprotein Group A-B / Q000235:genetics / Q000378:metabolism; D034441:Heterogeneous-Nuclear Ribonucleoproteins / Q000235:genetics / Q000378:metabolism; D059014:High-Throughput Nucleotide Sequencing; D006801:Humans; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism; D000071447:N-Myc Proto-Oncogene Protein; D009367:Neoplasm Staging; D009447:Neuroblastoma / Q000235:genetics* / Q000378:metabolism / Q000401:mortality; D009687:Nuclear Proteins / Q000235:genetics* / Q000378:metabolism; D015513:Oncogene Proteins / Q000235:genetics* / Q000378:metabolism; D038941:Polypyrimidine Tract-Binding Protein / Q000235:genetics / Q000378:metabolism; D011401:Promoter Regions, Genetic; D034622:RNA Interference; D012334:RNA, Neoplasm / Q000235:genetics* / Q000378:metabolism; D012307:Risk Factors; D013963:Thyroid Hormones / Q000235:genetics / Q000378:metabolism; D014158:Transcription, Genetic; D014162:Transfection; D000097810:Thyroid Hormone-Binding Proteins
chemical_list: D014408:Biomarkers, Tumor; D002352:Carrier Proteins; D000076245:Heterogeneous Nuclear Ribonucleoprotein A1; D034481:Heterogeneous-Nuclear Ribonucleoprotein Group A-B; D034441:Heterogeneous-Nuclear Ribonucleoproteins; D008565:Membrane Proteins; D000071447:N-Myc Proto-Oncogene Protein; D009687:Nuclear Proteins; D015513:Oncogene Proteins; D038941:Polypyrimidine Tract-Binding Protein; D012334:RNA, Neoplasm; D013963:Thyroid Hormones; D000097810:Thyroid Hormone-Binding Proteins; C459801:MYCN protein, human; C450006:PTBP1 protein, human; C000619656:hnRNPA1 protein, human
keywords: MYCN-amplification; Neuroblastoma; RNA isoforms; RNA-seq; Splicing