The intronic splicing code: multiple factors involved in ATM pseudoexon definition

Authors

  • Ashish Dhir,

    1. Molecular Pathology Group, International Centre for Genetic Engineering and Biotechnology, Trieste, Italy
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  • Emanuele Buratti,

    1. Molecular Pathology Group, International Centre for Genetic Engineering and Biotechnology, Trieste, Italy
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  • Maria A van Santen,

    1. Department of Cellular Biochemistry, Max Planck Institute for Biophysical Chemistry, Göttingen, Germany
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  • Reinhard Lührmann,

    1. Department of Cellular Biochemistry, Max Planck Institute for Biophysical Chemistry, Göttingen, Germany
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  • Francisco E Baralle

    Corresponding author
    1. Molecular Pathology Group, International Centre for Genetic Engineering and Biotechnology, Trieste, Italy
    • Corresponding author. Molecular Pathology Group, International Centre for Genetic Engineering and Biotechnology, Padriciano 99, Trieste 34149, Italy. Tel.: +39 040 3757 337; Fax: +39 040 375 7361; E-mail: baralle@icgeb.org

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Abstract

Abundance of pseudo splice sites in introns can potentially give rise to innumerable pseudoexons, outnumbering the real ones. Nonetheless, these are efficiently ignored by the splicing machinery, a process yet to be understood completely. Although numerous 5′ splice site-like sequences functioning as splicing silencers have been found to be enriched in predicted human pseudoexons, the lack of active pseudoexons pose a fundamental challenge to how these U1snRNP-binding sites function in splicing inhibition. Here, we address this issue by focusing on a previously described pathological ATM pseudoexon whose inhibition is mediated by U1snRNP binding at intronic splicing processing element (ISPE), composed of a consensus donor splice site. Spliceosomal complex assembly demonstrates inefficient A complex formation when ISPE is intact, implying U1snRNP-mediated unproductive U2snRNP recruitment. Furthermore, interaction of SF2/ASF with its motif seems to be dependent on RNA structure and U1snRNP interaction. Our results suggest a complex combinatorial interplay of RNA structure and trans-acting factors in determining the splicing outcome and contribute to understanding the intronic splicing code for the ATM pseudoexon.

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