The activity of siRNA in mammalian cells is related to structural target accessibility: a comparison with antisense oligonucleotides
- 1 August 2003
- journal article
- research article
- Published by Oxford University Press (OUP) in Nucleic Acids Research
- Vol. 31 (15) , 4417-4424
- https://doi.org/10.1093/nar/gkg649
Abstract
The biological activity of siRNA seems to be influenced by local characteristics of the target RNA, including local RNA folding. Here, we investigated quantitatively the relationship between local target accessibility and the extent of inhibition of the target gene by siRNA. Target accessibility was assessed by a computational approach that had been shown earlier to be consistent with experimental probing of target RNA. Two sites of ICAM-1 mRNA predicted to serve as accessible motifs and one site predicted to adopt an inaccessible structure were chosen to test siRNA constructs for suppression of ICAM-1 gene expression in ECV304 cells. The local target-dependent effectiveness of siRNA was compared with antisense oligonucleotides (asON). The concentration dependency of siRNA-mediated suppression indicates a >1000-fold difference between active siRNAs (IC50 approximately 0.2-0.5 nM) versus an inactive siRNA (IC50 > or = 1 microM) which is consistent with the activity pattern of asON when relating target suppression to predicted local target accessibility. The extremely high activity of the siRNA si2B (IC50 = 0.24 nM) indicates that not all siRNAs shown to be active at the usual concentrations of >10-100 nM belong to this highly active species. The observations described here suggest an option to assess target accessibility for siRNA and, thus, support the design of active siRNA constructs. This approach can be automated, work at high throughput and is open to include additional parameters relevant to the biological activity of siRNA.Keywords
This publication has 31 references indexed in Scilit:
- The Efficacy of Small Interfering RNAs Targeted to the Type 1 Insulin-like Growth Factor Receptor (IGF1R) Is Influenced by Secondary Structure in the IGF1R TranscriptJournal of Biological Chemistry, 2003
- Efficient Reduction of Target RNAs by Small Interfering RNA and RNase H-dependent Antisense AgentsJournal of Biological Chemistry, 2003
- Comparison of the Suppressive Effects of Antisense Oligonucleotides and siRNAs Directed Against the Same Targets in Mammalian CellsAntisense and Nucleic Acid Drug Development, 2003
- The Effectiveness of Double-Stranded Short Inhibitory RNAs (siRNAs) May Depend on the Method of TransfectionAntisense and Nucleic Acid Drug Development, 2002
- Effects on RNA Interference in Gene Expression (RNAi) in Cultured Mammalian Cells of Mismatches and the Introduction of Chemical Modifications at the 3′-Ends of siRNAsAntisense and Nucleic Acid Drug Development, 2002
- Functional anatomy of siRNAs for mediating efficient RNAi in Drosophila melanogaster embryo lysateThe EMBO Journal, 2001
- Detection of Antisense and Ribozyme Accessible Sites on Native mRNAs: Application to NCOA3 mRNAMolecular Therapy, 2001
- Expanded sequence dependence of thermodynamic parameters improves prediction of RNA secondary structureJournal of Molecular Biology, 1999
- Mapping of RNA accessible sites for antisense experiments with oligonucleotide librariesNature Biotechnology, 1998
- The Association of Complementary Ribonucleic Acids Can Be Strongly Increased without Lowering Arrhenius Activation Energies or Significantly Altering StructuresBiochemistry, 1997