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Improved binding site assignment by high-resolution mapping of RNA–protein interactions using iCLIP

Author

Listed:
  • Christian Hauer

    (Hematology and Immunology, University of Heidelberg
    Molecular Medicine Partnership Unit (MMPU)
    European Molecular Biology Laboratory (EMBL) Heidelberg)

  • Tomaz Curk

    (European Molecular Biology Laboratory (EMBL) Heidelberg
    University of Ljubljana, Faculty of Computer and Information Science)

  • Simon Anders

    (European Molecular Biology Laboratory (EMBL) Heidelberg)

  • Thomas Schwarzl

    (European Molecular Biology Laboratory (EMBL) Heidelberg)

  • Anne-Marie Alleaume

    (European Molecular Biology Laboratory (EMBL) Heidelberg)

  • Jana Sieber

    (Hematology and Immunology, University of Heidelberg
    Molecular Medicine Partnership Unit (MMPU))

  • Ina Hollerer

    (Hematology and Immunology, University of Heidelberg
    Molecular Medicine Partnership Unit (MMPU)
    European Molecular Biology Laboratory (EMBL) Heidelberg)

  • Madhuri Bhuvanagiri

    (Hematology and Immunology, University of Heidelberg
    Molecular Medicine Partnership Unit (MMPU))

  • Wolfgang Huber

    (European Molecular Biology Laboratory (EMBL) Heidelberg)

  • Matthias W. Hentze

    (Molecular Medicine Partnership Unit (MMPU)
    European Molecular Biology Laboratory (EMBL) Heidelberg)

  • Andreas E. Kulozik

    (Hematology and Immunology, University of Heidelberg
    Molecular Medicine Partnership Unit (MMPU))

Abstract

Individual-nucleotide resolution crosslinking and immunoprecipitation (iCLIP) allows the determination of crosslinking sites of RNA-binding proteins (RBPs) on RNAs. iCLIP is based on ultraviolet light crosslinking of RBPs to RNA, reverse transcription and high-throughput sequencing of fragments terminating at the site of crosslinking. As a result, start sites of iCLIP fragments are expected to cluster with a narrow distribution, typically representing the site of direct interaction between the RBP and the RNA. Here we show that for several RBPs (eIF4A3, PTB, SRSF3, SRSF4 and hnRNP L), the start sites of iCLIP fragments show a fragment length-dependent broader distribution that can be shifted to positions upstream of the known RNA-binding site. We developed an analysis tool that identifies these shifts and can improve the positioning of RBP binding sites.

Suggested Citation

  • Christian Hauer & Tomaz Curk & Simon Anders & Thomas Schwarzl & Anne-Marie Alleaume & Jana Sieber & Ina Hollerer & Madhuri Bhuvanagiri & Wolfgang Huber & Matthias W. Hentze & Andreas E. Kulozik, 2015. "Improved binding site assignment by high-resolution mapping of RNA–protein interactions using iCLIP," Nature Communications, Nature, vol. 6(1), pages 1-13, November.
    • Handle: RePEc:nat:natcom:v:6:y:2015:i:1:d:10.1038_ncomms8921
    DOI: 10.1038/ncomms8921
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