RIPchipSRM, a new combinatorial large scale approach identifies a set of translationally regulated bantam/miR58 targets in C. elegans
- Marko Jovanovic1,
- Lukas Reiter1,
- Alejandra Clark2,
- Manuel Weiss1,
- Paola Picotti3,
- Hubert Rehrauer4,
- Andreas Frei3,
- Lukas Neukomm5,
- Ethan Kaufman2,
- Bernd Wollscheid3,
- Martin J. Simard6,
- Eric Miska2,
- Ruedi Aebersold3,
- Andre P. Gerber7 and
- Michael O. Hengartner1,8
- 1 Institute of Molecular Life Sciences, University of Zurich, Zurich, Switzerland;
- 2 Wellcome Trust/Cancer Research UK Gurdon Institute, University of Cambridge, Cambridge, CB2 1QN, Uni;
- 3 Department of Biology, Institute of Molecular Systems Biology, ETH Zurich, Zurich, Switzerland;
- 4 Functional Genomics Center, ETH and University of Zurich, Zurich, Switzerland.;
- 5 Neurobiology Department, Howard Hughes Medical Institute, University of Massachusetts Medical School;
- 6 Laval University Cancer Research Centre, Hotel Dieu de Quebec (CHUQ), Quebec City, Quebec, Canada;
- 7 Institute of Pharmaceutical Sciences, ETH Zurich, Zurich, Switzerland
- ↵* Corresponding author; email: michael.hengartner{at}imls.uzh.ch
Abstract
MicroRNAs (miRNAs) are small, noncoding RNAs that negatively regulate gene expression. As miRNAs are involved in a wide range of biological processes and diseases, much effort has been invested in identifying their mRNA targets. Here we present a novel combinatorial approach, RIPchipSRM (RNA binding protein immunopurification + microarray + targeted protein quantification via selected reaction monitoring), to identify de novo highconfidence miRNA targets in the nematode Caenorhabditis elegans. We used differential RIP-chip analysis of miRNA induced silencing complexes from wildtype and miRNA mutant animals, followed by quantitative targeted proteomics via selected reaction monitoring to identify and validate mRNA targets of the C. elegans bantam homologue miR58. Comparison of total mRNA and protein abundance changes in mir58 mutant and wildtype animals indicated that the direct bantam/miR58 targets identified here are mainly regulated at the level of protein abundance, not mRNA stability.
- Received October 13, 2011.
- Accepted March 19, 2012.
- Copyright © 2012, Cold Spring Harbor Laboratory Press
This manuscript is Open Access.
