RBM24 promotes U1 snRNP recognition of the mutated 5′ splice site in the IKBKAP gene of familial dysautonomia

  1. Masatoshi Hagiwara1
  1. 1Department of Anatomy and Developmental Biology, Kyoto University Graduate School of Medicine, Sakyo-ku, Kyoto 606-8501, Japan
  2. 2Training Program of Leaders for Integrated Medical System for Fruitful Healthy-Longevity Society (LIMS), Kyoto University Graduate School of Medicine, Sakyo-ku, Kyoto 606-8501, Japan
  3. 3Medical Research Support Center, Kyoto University Graduate School of Medicine, Sakyo-ku, Kyoto 606-8501, Japan
  4. 4Laboratory of Gene Expression, School of Biomedical Science, Tokyo Medical and Dental University, Bunkyo-ku, Tokyo 113-8510, Japan
  5. 5Sir William Dunn School of Pathology, University of Oxford, Oxford OX1 3RE, United Kingdom
  6. 6Laboratory for Malignancy Control Research, Medical Innovation Center, Kyoto University Graduate School of Medicine, Sakyo-ku, Kyoto 606-8507, Japan
  7. 7Laboratory of Cell Regulation, Departments of Applied Animal Sciences and Applied Biological Chemistry, Graduate School of Agriculture and Life Sciences, The University of Tokyo, Bunkyo-ku, Tokyo 113-8657, Japan
  1. Corresponding authors: hagiwara.masatoshi.8c{at}kyoto-u.ac.jp, akataoka{at}mail.ecc.u-tokyo.ac.jp
  • 8 Present address: Department of Pharmaceutical Sciences, Fukuoka University, Fukuoka 814-0180, Japan

  • 9 Present address: Frontier Research Core for Life Sciences, University of Toyama, Toyama 930-0194, Japan

  • 10 Present address: Institute for Genetic Medicine, Hokkaido University, Sapporo 060-0815, Japan

Abstract

The 5′ splice site mutation (IVS20+6T>C) of the inhibitor of κ light polypeptide gene enhancer in B cells, kinase complex-associated protein (IKBKAP) gene in familial dysautonomia (FD) is at the sixth intronic nucleotide of the 5′ splice site. It is known to weaken U1 snRNP recognition and result in an aberrantly spliced mRNA product in neuronal tissue, but normally spliced mRNA in other tissues. Aberrantly spliced IKBKAP mRNA abrogates IKK complex-associated protein (IKAP)/elongator protein 1 (ELP1) expression and results in a defect of neuronal cell development in FD. To elucidate the tissue-dependent regulatory mechanism, we screened an expression library of major RNA-binding proteins (RBPs) with our mammalian dual-color splicing reporter system and identified RBM24 as a regulator. RBM24 functioned as a cryptic intronic splicing enhancer binding to an element (IVS20+13–29) downstream from the intronic 5′ splice site mutation in the IKBKAP gene and promoted U1 snRNP recognition only to the mutated 5′ splice site (and not the wild-type 5′ splice site). Our results show that tissue-specific expression of RBM24 can explain the neuron-specific aberrant splicing of IKBKAP exon 20 in familial dysautonomia, and that ectopic expression of RBM24 in neuronal tissue could be a novel therapeutic target of the disease.

Keywords

Footnotes

  • Abbreviations: EMSA, electromobility shift assay; FD, familial dysautonomia; FGFR, fibroblast growth factor receptor; IKBKAP, inhibitor of κ light polypeptide gene enhancer in B cells, kinase complex-associated protein; IP, immunoprecipitation; RBPs, RNA-binding proteins; RF, red fluorescent protein; SPREADD, splicing reporter assay for disease genes with dual color; TAE, Tris–acetate–EDTA; TBE, Tris–borate–EDTA

  • Article is online at http://www.rnajournal.org/cgi/doi/10.1261/rna.059428.116.

  • Received October 6, 2016.
  • Accepted May 25, 2017.

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