Unwinding by local strand separation is critical for the function of DEAD-box proteins as RNA chaperones

J Mol Biol. 2009 Jun 19;389(4):674-93. doi: 10.1016/j.jmb.2009.04.043. Epub 2009 Apr 23.


The DEAD-box proteins CYT-19 in Neurospora crassa and Mss116p in Saccharomyces cerevisiae are broadly acting RNA chaperones that function in mitochondria to stimulate group I and group II intron splicing and to activate mRNA translation. Previous studies showed that the S. cerevisiae cytosolic/nuclear DEAD-box protein Ded1p could stimulate group II intron splicing in vitro. Here, we show that Ded1p complements mitochondrial translation and group I and group II intron splicing defects in mss116Delta strains, stimulates the in vitro splicing of group I and group II introns, and functions indistinguishably from CYT-19 to resolve different nonnative secondary and/or tertiary structures in the Tetrahymena thermophila large subunit rRNA-DeltaP5abc group I intron. The Escherichia coli DEAD-box protein SrmB also stimulates group I and group II intron splicing in vitro, while the E. coli DEAD-box protein DbpA and the vaccinia virus DExH-box protein NPH-II gave little, if any, group I or group II intron splicing stimulation in vitro or in vivo. The four DEAD-box proteins that stimulate group I and group II intron splicing unwind RNA duplexes by local strand separation and have little or no specificity, as judged by RNA-binding assays and stimulation of their ATPase activity by diverse RNAs. In contrast, DbpA binds group I and group II intron RNAs nonspecifically, but its ATPase activity is activated specifically by a helical segment of E. coli 23S rRNA, and NPH-II unwinds RNAs by directional translocation. The ability of DEAD-box proteins to stimulate group I and group II intron splicing correlates primarily with their RNA-unwinding activity, which, for the protein preparations used here, was greatest for Mss116p, followed by Ded1p, CYT-19, and SrmB. Furthermore, this correlation holds for all group I and group II intron RNAs tested, implying a fundamentally similar mechanism for both types of introns. Our results support the hypothesis that DEAD-box proteins have an inherent ability to function as RNA chaperones by virtue of their distinctive RNA-unwinding mechanism, which enables refolding of localized RNA regions or structures without globally disrupting RNA structure.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Base Sequence
  • DEAD-box RNA Helicases / genetics
  • DEAD-box RNA Helicases / metabolism*
  • DNA, Mitochondrial / genetics
  • DNA, Mitochondrial / metabolism
  • Genetic Complementation Test
  • Introns
  • Molecular Chaperones / genetics
  • Molecular Chaperones / metabolism*
  • Molecular Sequence Data
  • Neurospora crassa / genetics
  • Neurospora crassa / metabolism
  • Nucleic Acid Conformation*
  • Protein Biosynthesis
  • RNA Splicing*
  • RNA* / chemistry
  • RNA* / genetics
  • RNA* / metabolism
  • Saccharomyces cerevisiae / genetics
  • Saccharomyces cerevisiae / metabolism
  • Saccharomyces cerevisiae Proteins / genetics
  • Saccharomyces cerevisiae Proteins / metabolism


  • DNA, Mitochondrial
  • Molecular Chaperones
  • Saccharomyces cerevisiae Proteins
  • RNA
  • DED1 protein, S cerevisiae
  • MSS116 protein, S cerevisiae
  • DEAD-box RNA Helicases