The DEAH box ATPases Prp16 and Prp43 cooperate to proofread 5' splice site cleavage during pre-mRNA splicing

Mol Cell. 2010 Aug 13;39(3):385-95. doi: 10.1016/j.molcel.2010.07.014.


To investigate the mechanisms underlying accurate pre-mRNA splicing, we developed an in vitro assay sensitive to proofreading of 5' splice site cleavage. We inactivated spliceosomes by disrupting a metal-ligand interaction at the catalytic center and discovered that, when the DEAH box ATPase Prp16 was disabled, these spliceosomes catalyzed 5' splice site cleavage but at a reduced rate. Although Prp16 does not promote splicing of a genuine substrate until after 5' splice site cleavage, we found that Prp16 can associate with spliceosomes before 5' splice site cleavage, consistent with a role for Prp16 in proofreading 5' splice site cleavage. We established that Prp16-mediated rejection is reversible, necessitating a downstream discard pathway that we found requires the DEAH box ATPase Prp43, a spliceosome disassembly factor. These data indicate that spliceosomes distinguish slow substrates and that the mechanisms for establishing the fidelity of 5' splice site cleavage and exon ligation share a common ATP-dependent framework.

Publication types

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

MeSH terms

  • Adenosine Triphosphatases / genetics
  • Adenosine Triphosphatases / metabolism*
  • Adenosine Triphosphate / genetics
  • Adenosine Triphosphate / metabolism
  • DEAD-box RNA Helicases / genetics
  • DEAD-box RNA Helicases / metabolism*
  • RNA Helicases / genetics
  • RNA Helicases / metabolism*
  • RNA Precursors / genetics
  • RNA Precursors / metabolism*
  • RNA Splice Sites / physiology*
  • RNA Splicing / physiology*
  • RNA Splicing Factors
  • RNA, Fungal / genetics
  • RNA, Fungal / metabolism*
  • Saccharomyces cerevisiae / genetics
  • Saccharomyces cerevisiae / metabolism*
  • Saccharomyces cerevisiae Proteins / genetics
  • Saccharomyces cerevisiae Proteins / metabolism*
  • Spliceosomes / genetics
  • Spliceosomes / metabolism


  • RNA Precursors
  • RNA Splice Sites
  • RNA Splicing Factors
  • RNA, Fungal
  • Saccharomyces cerevisiae Proteins
  • Adenosine Triphosphate
  • Adenosine Triphosphatases
  • PRP16 protein, S cerevisiae
  • PRP43 protein, S cerevisiae
  • DEAD-box RNA Helicases
  • RNA Helicases