RNA looping by PTB: Evidence using FRET and NMR spectroscopy for a role in splicing repression

Proc Natl Acad Sci U S A. 2010 Mar 2;107(9):4105-10. doi: 10.1073/pnas.0907072107. Epub 2010 Feb 16.

Abstract

Alternative splicing plays an important role in generating proteome diversity. The polypyrimidine tract-binding protein (PTB) is a key alternative splicing factor involved in exon repression. It has been proposed that PTB acts by looping out exons flanked by pyrimidine tracts. We present fluorescence, NMR, and in vivo splicing data in support of a role of PTB in inducing RNA loops. We show that the RNA recognition motifs (RRMs) 3 and 4 of PTB can bind two distant pyrimidine tracts and bring their 5' and 3' ends in close proximity, thus looping the RNA. Efficient looping requires an intervening sequence of 15 nucleotides or longer between the pyrimidine tracts. RRM3 and RRM4 bind the 5' and the 3' pyrimidine tracts, respectively, in a specific directionality and work synergistically for efficient splicing repression in vivo.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Alternative Splicing*
  • Blotting, Western
  • Fluorescence Resonance Energy Transfer / methods*
  • HeLa Cells
  • Humans
  • Nuclear Magnetic Resonance, Biomolecular / methods*
  • Polypyrimidine Tract-Binding Protein / chemistry*
  • Protein Conformation
  • RNA / chemistry*
  • Reverse Transcriptase Polymerase Chain Reaction

Substances

  • Polypyrimidine Tract-Binding Protein
  • RNA