RNAi on or off target?

From: News and Views, Nature Genetics 34, 133 (2003)

Two new reports examine the specificity of RNA interference (RNAi) in human cells using genome-wide expression profiling. Targeting exogenous GFP in human embryonic kidney cells, Jen-Tsan Chi and colleagues report efficient and specific knockdown using two different siRNAs (Proc. Natl. Acad. Sci USA 10.1073/pnas.1037853100). Analysis of approximately 20,000 genes identified no consistent gene expression profile and no statistically significant difference in global gene expression patterns associated with the two siRNAs. Further analyses did not identify spreading of the RNAi effect to similar exogenous sequences, leading the authors to conclude that the RNAi effect is on target in mammalian cells. In a related paper, Peter Linsley and colleagues targeted endogenous genes (Nat. Biotechnol. 10.1038/nbt831). They examined the effect of 16 different siRNAs against IGF1R and 8 different siRNAs targeted against MAPK14. In each case, the gene expression profile was siRNA-specific. A group of 9 genes with partial sequence identity to the siRNA duplex targeted against MAPK14 were downregulated with similar kinetics. The authors conclude that at least some cross-hybridization of siRNAs to transcripts of similar sequences occurs. These results suggest that ensuring siRNA-specific effects requires well controlled experiments.