Journal
NATURE METHODS
Volume 7, Issue 10, Pages 837-U100Publisher
NATURE PUBLISHING GROUP
DOI: 10.1038/NMETH.1504
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Funding
- University of Zurich
- Swiss National Science Foundation
- Gerbert Ruf Foundation
- Swiss initiative for systems biology
- SystemsX
- Ernst Hadorn Foundation
- ETH Zurich
- European Research Council [ERC-2008-AdG 233226]
- Research Foundation of the University of Zurich
- Roche Research Foundation
- Marie Curie Intra-European fellowship
- F. Hoffmann-La Roche Ltd.
- Swiss National Research Foundation
- Novartis Research Foundation
- ERC
- Boehringer Ingelheim
- Cancer Research UK
- Cancer Research UK [11832] Funding Source: researchfish
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Efficient experimental strategies are needed to validate computationally predicted microRNA (miRNA) target genes. Here we present a large-scale targeted proteomics approach to validate predicted miRNA targets in Caenorhabditis elegans. Using selected reaction monitoring (SRM), we quantified 161 proteins of interest in extracts from wild-type and let-7 mutant worms. We demonstrate by independent experimental downstream analyses such as genetic interaction, as well as polysomal profiling and luciferase assays, that validation by targeted proteomics substantially enriched for biologically relevant let-7 interactors. For example, we found that the zinc finger protein ZTF-7 was a bona fide let-7 miRNA target. We also validated predicted miR-58 targets, demonstrating that this approach is adaptable to other miRNAs. We propose that targeted mass spectrometry can be applied generally to validate candidate lists generated by computational methods or in large-scale experiments, and that the described strategy should be readily adaptable to other organisms.
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