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| Content Provider | World Health Organization (WHO)-Global Index Medicus |
|---|---|
| Author | Liang, Xue-hai Crooke, Stanley T. |
| Description | Author Affiliation: Liang XH ( Department of Core Antisense Research, ISIS Pharmaceuticals, Inc., 2855 Gazelle Court, Carlsbad, CA 92010, USA. Electronic address: lliang@isisph.com.) |
| Abstract | It has been shown that siRNAs can compete with each other or with endogenous miRNAs for RISC components. This competition may complicate the interpretations of phenotypes observed through siRNA-mediated knockdown of genes, especially those genes implicated in the RISC pathway. In this study, we re-examined the function of RNA helicase A (RHA), which has been previously proposed to function in RISC loading based on siRNA-mediated knockdown studies. Here we show that reduced RISC activity or loading of siRNAs was observed only in cells depleted of RHA using siRNA, but not using RNaseH-dependent antisense oligonucleotides (ASOs), suggesting that the impaired RISC function stems from the competition between pre-existing and newly transfected siRNAs, but not from reduction of the RHA protein. This view is further supported by the findings that cells depleted of a control protein, NCL1, using siRNA, but not ASO, exhibited similar defects on the loading and activity of a subsequently transfected siRNA. Transfection of RHA or NCL1 siRNAs, but not ASOs, reduced the levels of endogenous miRNAs, suggesting a competition mechanism. As a positive control, we showed that reduction of MOV10 by either siRNA or ASO decreased siRNA activity, confirming its role in RISC function. Together, our results indicate that RHA is not required for RISC activity or loading, and suggest that proper controls are required when using siRNAs to functionalize genes to avoid competition effects. |
| ISSN | 00063002 |
| Journal | Biochimica et Biophysica Acta (BBA) - Reviews on Cancer |
| Issue Number | 10 |
| Volume Number | 1829 |
| Language | English |
| Publisher | Elsevier |
| Publisher Date | 2013-10-01 |
| Publisher Place | Netherlands |
| Access Restriction | Open |
| Subject Keyword | MicroRNAs Genetics RNA Helicases Metabolism RNA, Small Interfering RNA-Induced Silencing Complex Blotting, Western HeLa Cells Immunoprecipitation Oligonucleotides, Antisense Pharmacology Phosphoproteins Antagonists & Inhibitors RNA, Messenger RNA-Binding Proteins Real-Time Polymerase Chain Reaction Reverse Transcriptase Polymerase Chain Reaction Research Support, Non-U.S. Gov't Biochemistry |
| Content Type | Text |
| Resource Type | Article |
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