Please use this identifier to cite or link to this item:
https://doi.org/10.1186/1758-907X-2-3
DC Field | Value | |
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dc.title | Off-target effects dominate a large-scale RNAi screen for modulators of the TGF-? pathway and reveal microRNA regulation of TGFBR2 | |
dc.contributor.author | Schultz, N | |
dc.contributor.author | Marenstein, D.R | |
dc.contributor.author | De Angelis, D.A | |
dc.contributor.author | Wang, W.-Q | |
dc.contributor.author | Nelander, S | |
dc.contributor.author | Jacobsen, A | |
dc.contributor.author | Marks, D.S | |
dc.contributor.author | Massagué, J | |
dc.contributor.author | Sander, C | |
dc.date.accessioned | 2020-10-20T08:14:17Z | |
dc.date.available | 2020-10-20T08:14:17Z | |
dc.date.issued | 2011 | |
dc.identifier.citation | Schultz, N, Marenstein, D.R, De Angelis, D.A, Wang, W.-Q, Nelander, S, Jacobsen, A, Marks, D.S, Massagué, J, Sander, C (2011). Off-target effects dominate a large-scale RNAi screen for modulators of the TGF-? pathway and reveal microRNA regulation of TGFBR2. Silence 2 (1) : 3. ScholarBank@NUS Repository. https://doi.org/10.1186/1758-907X-2-3 | |
dc.identifier.issn | 1758-907X | |
dc.identifier.uri | https://scholarbank.nus.edu.sg/handle/10635/178179 | |
dc.description.abstract | Background: RNA interference (RNAi) screens have been used to identify novel components of signal-transduction pathways in a variety of organisms. We performed a small interfering (si)RNA screen for novel members of the transforming growth factor (TGF)-? pathway in a human keratinocyte cell line. The TGF-? pathway is integral to mammalian cell proliferation and survival, and aberrant TGF-? responses have been strongly implicated in cancer.Results: We assayed how strongly single siRNAs targeting each of 6,000 genes affect the nuclear translocation of a green fluorescent protein (GFP)-SMAD2 reporter fusion protein. Surprisingly, we found no novel TGF-? pathway members, but we did find dominant off-target effects. All siRNA hits, whatever their intended direct target, reduced the mRNA levels of two known upstream pathway components, the TGF-? receptors 1 and 2 (TGFBR1 and TGFBR2), via micro (mi)RNA-like off-target effects. The scale of these off-target effects was remarkable, with at least 1% of the sequences in the unbiased siRNA library having measurable off-target effects on one of these two genes. It seems that relatively minor reductions of message levels via off-target effects can have dominant effects on an assay, if the pathway output is very dose-sensitive to levels of particular pathway components. In search of mechanistic details, we identified multiple miRNA-like sequence characteristics that correlated with the off-target effects. Based on these results, we identified miR-20a, miR-34a and miR-373 as miRNAs that inhibit TGFBR2 expression.Conclusions: Our findings point to potential improvements for miRNA/siRNA target prediction methods, and suggest that the type II TGF-? receptor is regulated by multiple miRNAs. We also conclude that the risk of obtaining misleading results in siRNA screens using large libraries with single-assay readout is substantial. Control and rescue experiments are essential in the interpretation of such screens, and improvements to the methods to reduce or predict RNAi off-target effects would be beneficial. © 2011 Schultz et al; licensee BioMed Central Ltd. | |
dc.publisher | BMC | |
dc.rights | Attribution 4.0 International | |
dc.rights.uri | http://creativecommons.org/licenses/by/4.0/ | |
dc.source | Unpaywall 20201031 | |
dc.subject | green fluorescent protein | |
dc.subject | microRNA | |
dc.subject | microRNA 20a | |
dc.subject | microRNA 34a | |
dc.subject | microRNA 373 | |
dc.subject | Smad2 protein | |
dc.subject | small interfering RNA | |
dc.subject | transforming growth factor beta | |
dc.subject | transforming growth factor beta receptor 1 | |
dc.subject | transforming growth factor beta receptor 2 | |
dc.subject | unclassified drug | |
dc.subject | 3' untranslated region | |
dc.subject | article | |
dc.subject | CDKN1A gene | |
dc.subject | cell proliferation | |
dc.subject | cell survival | |
dc.subject | controlled study | |
dc.subject | gene | |
dc.subject | gene identification | |
dc.subject | gene sequence | |
dc.subject | gene targeting | |
dc.subject | gene translocation | |
dc.subject | genetic analysis | |
dc.subject | genetic regulation | |
dc.subject | genetic screening | |
dc.subject | genetic transcription | |
dc.subject | human | |
dc.subject | human cell | |
dc.subject | keratinocyte | |
dc.subject | nuclear localization signal | |
dc.subject | open reading frame | |
dc.subject | PAI 1 gene | |
dc.subject | phenotype | |
dc.subject | priority journal | |
dc.subject | protein expression | |
dc.subject | protein phosphorylation | |
dc.subject | protein targeting | |
dc.subject | RNA interference | |
dc.subject | sequence analysis | |
dc.subject | signal transduction | |
dc.subject | SMAD7 gene | |
dc.subject | TGFBR1 gene | |
dc.subject | TGFBR2 gene | |
dc.type | Article | |
dc.contributor.department | DEPARTMENT OF COMPUTER SCIENCE | |
dc.description.doi | 10.1186/1758-907X-2-3 | |
dc.description.sourcetitle | Silence | |
dc.description.volume | 2 | |
dc.description.issue | 1 | |
dc.description.page | 3 | |
dc.published.state | published | |
Appears in Collections: | Staff Publications Elements |
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