RNA helicase A interacts with RISC in human cells and functions in RISC loading

Mol Cell. 2007 May 25;26(4):523-37. doi: 10.1016/j.molcel.2007.04.016.

Abstract

RNA interference is a conserved pathway of sequence-specific gene silencing that depends on small guide RNAs and the action of proteins assembled in the RNA-induced silencing complex (RISC). Minimally, the action of RISC requires the endonucleolytic slicer activity of Argonaute2 (Ago2) directed to RNA targets whose sequences are complementary to RISC-incorporated small RNA. To identify RISC components in human cells, we developed an affinity-purification strategy to isolate siRNA-programmed RISC. Here we report the identification of RNA helicase A (RHA) as a human RISC-associated factor. We show that RHA interacts in human cells with siRNA, Ago2, TRBP, and Dicer and functions in the RNAi pathway. In RHA-depleted cells, RNAi was reduced as a consequence of decreased intracellular concentration of active RISC assembled with the guide-strand RNA and Ago2. Our results identify RHA as a RISC component and demonstrate that RHA functions in RISC as an siRNA-loading factor.

Publication types

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

MeSH terms

  • Argonaute Proteins
  • Carboxypeptidases / chemistry
  • Carboxypeptidases / genetics
  • Carboxypeptidases / metabolism*
  • Cell Line
  • DEAD-box RNA Helicases / chemistry
  • DEAD-box RNA Helicases / genetics
  • DEAD-box RNA Helicases / metabolism*
  • Eukaryotic Initiation Factor-2 / metabolism
  • Gene Silencing*
  • Genes, Reporter
  • HeLa Cells
  • Humans
  • Kidney
  • Neoplasm Proteins / chemistry
  • Neoplasm Proteins / genetics
  • Neoplasm Proteins / metabolism*
  • RNA Interference
  • RNA, Messenger / genetics
  • RNA, Messenger / metabolism
  • Recombinant Fusion Proteins / chemistry
  • Recombinant Fusion Proteins / metabolism
  • Transfection

Substances

  • AGO2 protein, human
  • Argonaute Proteins
  • Eukaryotic Initiation Factor-2
  • Neoplasm Proteins
  • RNA, Messenger
  • Recombinant Fusion Proteins
  • Carboxypeptidases
  • SCPEP1 protein, human
  • DHX9 protein, human
  • DEAD-box RNA Helicases