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Rbfox proteins regulate alternative mRNA splicing through evolutionarily conserved RNA bridges.

Michael T Lovci1, Dana Ghanem, Henry Marr

  • 11] Department of Cellular and Molecular Medicine, University of California, San Diego, La Jolla, California, USA. [2] Stem Cell Program, University of California, San Diego, La Jolla, California, USA. [3] Institute for Genomic Medicine, University of California, San Diego, La Jolla, California, USA.

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|November 12, 2013
PubMed
Summary

Rbfox proteins regulate gene expression by binding to distant intronic DNA, influencing alternative splicing. This study reveals a novel RNA-mediated mechanism controlling gene diversity through long-range interactions.

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Area of Science:

  • Molecular Biology
  • Genetics
  • Gene Regulation

Background:

  • Alternative splicing (AS) generates diverse gene products from a single gene, crucial for development and tissue specificity.
  • Rbfox proteins are key regulators of AS, typically binding near exons.
  • The role of Rbfox binding in distal intronic regions remains largely unexplored.

Purpose of the Study:

  • To investigate the role of Rbfox protein binding in distal intronic regions (>500 nt from exons) in regulating alternative splicing.
  • To identify conserved regulatory elements and mechanisms governing distal Rbfox-mediated AS.
  • To demonstrate a novel RNA-mediated mechanism for AS control by distally bound RNA-binding proteins.

Main Methods:

  • Analysis of Rbfox binding sites in distal intronic regions across species.
  • Experimental manipulation of Rbfox abundance in human and mouse cell lines.
  • Validation of Rbfox enhancer activity in specific gene introns (KIF21A, ENAH).
  • Investigation of RNA-RNA base-pairing interactions (RNA bridges) in ENAH AS regulation.

Main Results:

  • Rbfox proteins bind extensively to conserved GCATG sequences in distal intronic regions.
  • Distal Rbfox binding actively regulates alternative splicing, causing both activation and repression.
  • Rbfox enhancer activity was confirmed in KIF21A and ENAH distal introns.
  • A conserved long-range RNA-RNA base-pairing interaction (RNA bridge) is essential for Rbfox-mediated exon inclusion in ENAH.

Conclusions:

  • Rbfox proteins regulate alternative splicing through binding in distal intronic regions, extending beyond previously known proximal binding sites.
  • This distal binding involves conserved sequences and actively modulates gene expression.
  • A novel RNA-mediated mechanism, involving RNA bridges, underlies Rbfox control of AS, highlighting a new layer of gene regulation.