Double-stranded RNA-dependent ATPase DRH-3: Insight Into Its Role in RNAsilencing in Caenorhabditis Elegans

J Biol Chem. 2010 Aug 13;285(33):25363-71. doi: 10.1074/jbc.M110.117010. Epub 2010 Jun 7.

Abstract

RNA helicases are proteins essential to almost every facet of RNA metabolism, including the gene-silencing pathways that employ small RNAs. A phylogenetically related group of helicases is required for the RNA-silencing mechanism in Caenorhabditis elegans. Dicer-related helicase 3 (DRH-3) is a Dicer-RIG-I family protein that is essential for RNA silencing and germline development in nematodes. Here we performed a biochemical characterization of the ligand binding and catalytic activities of DRH-3 in vitro. We identify signature motifs specific to this family of RNA helicases. We find that DRH-3 binds both single-stranded and double-stranded RNAs with high affinity. However, the ATPase activity of DRH-3 is stimulated only by double-stranded RNA. DRH-3 is a robust RNA-stimulated ATPase with a k(cat) value of 500/min when stimulated with short RNA duplexes. The DRH-3 ATPase may have allosteric regulation in cis that is controlled by the stoichiometry of double-stranded RNA to enzyme. We observe that the DRH-3 ATPase is stimulated only by duplexes containing RNA, suggesting a role for DRH-3 during or after transcription. Our findings provide clues to the role of DRH-3 during the RNA interference response in vivo.

Publication types

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

MeSH terms

  • Adenosine Triphosphatases / genetics
  • Adenosine Triphosphatases / metabolism*
  • Animals
  • Caenorhabditis elegans / enzymology*
  • Caenorhabditis elegans / genetics*
  • Caenorhabditis elegans Proteins / genetics
  • Caenorhabditis elegans Proteins / metabolism*
  • DEAD-box RNA Helicases / genetics
  • DEAD-box RNA Helicases / metabolism*
  • Protein Binding
  • RNA Interference / physiology*
  • RNA, Double-Stranded / metabolism*
  • Ultracentrifugation

Substances

  • Caenorhabditis elegans Proteins
  • RNA, Double-Stranded
  • Drh-3 protein, C elegans
  • Adenosine Triphosphatases
  • RNA-dependent ATPase
  • DEAD-box RNA Helicases