Off-target and a portion of target-specific siRNA mediated mRNA degradation is Ago2 'Slicer' independent and can be mediated by Ago1

Nucleic Acids Res. 2009 Nov;37(20):6927-41. doi: 10.1093/nar/gkp735. Epub 2009 Sep 18.


It is known that siRNAs are capable of reducing expression of non-target genes due to the interaction of the siRNA guide strand with a partially complementary site on the 'off-target' mRNA. In the current study, we show that reduction of cellular Ago2 levels has no effect on off-target reduction of endogenous genes and that off-target degradation of mRNA can occur even in an Ago2 knockout cell line. Using antisense mediated reduction of Ago proteins and chemically modified cleavage- and binding-deficient siRNAs, we demonstrate that siRNA mediated off-target reduction is Ago2 cleavage independent, but does require siRNA interaction with either Ago1 or Ago2 and the RISC-loading complex. We also show that depletion of P-body associated proteins results in a reduction of off-target siRNA-mediated degradation of mRNA. Finally, we present data suggesting that a significant portion of on-target siRNA activity is also Ago2 cleavage independent, however, this activity does not appear to be P-body associated.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Argonaute Proteins
  • Cells, Cultured
  • Class I Phosphatidylinositol 3-Kinases
  • Eukaryotic Initiation Factor-2 / genetics
  • Eukaryotic Initiation Factors / metabolism*
  • Gene Expression Profiling
  • Humans
  • Mice
  • Mice, Knockout
  • Phosphatidylinositol 3-Kinases / metabolism
  • RNA Interference*
  • RNA Stability
  • RNA, Messenger / metabolism*
  • RNA, Small Interfering / metabolism*
  • Ribonuclease III / metabolism


  • AGO1 protein, human
  • Ago2 protein, mouse
  • Argonaute Proteins
  • Eukaryotic Initiation Factor-2
  • Eukaryotic Initiation Factors
  • RNA, Messenger
  • RNA, Small Interfering
  • Class I Phosphatidylinositol 3-Kinases
  • PIK3CB protein, human
  • Ribonuclease III