Identification of novel targets for miR-29a using miRNA proteomics
| Autor: | Robin Park, Rhishikesh Bargaje, Mahantappa Halimani, Maharnob Sarkar, Tao Xu, Beena Pillai, John R. Yates, Shivani Gupta, Soumya Sinha Roy, Ali Sarkeshik |
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| Jazyk: | angličtina |
| Rok vydání: | 2012 |
| Předmět: |
Proteomics
Cell Culture Techniques lcsh:Medicine Apoptosis Computational biology Biology Peptide Mapping Stable isotope labeling by amino acids in cell culture Molecular Cell Biology microRNA Humans Computer Simulation lcsh:Science 3' Untranslated Regions Cellular Stress Responses Regulation of gene expression Genetics Multidisciplinary Models Genetic Voltage-Dependent Anion Channel 2 Three prime untranslated region Gene Expression Profiling Voltage-Dependent Anion Channel 1 lcsh:R ATP Synthetase Complexes Gene expression profiling MicroRNAs HEK293 Cells Gene Expression Regulation Proteome lcsh:Q VDAC1 Algorithms Research Article |
| Zdroj: | PLoS ONE, Vol 7, Iss 8, p e43243 (2012) PLoS ONE |
| ISSN: | 1932-6203 |
| Popis: | MicroRNAs (miRNAs) are short regulatory RNA molecules that interfere with the expression of target mRNA by binding to complementary sequences. Currently, the most common method for identification of targets of miRNAs is computational prediction based on free energy change calculations, target site accessibility and conservation. Such algorithms predict hundreds of targets for each miRNA, necessitating tedious experimentation to identify the few functional targets. Here we explore the utility of miRNA-proteomics as an approach to identifying functional miRNA targets. We used Stable Isotope Labeling by amino acids in cell culture (SILAC) based proteomics to detect differences in protein expression induced by the over-expression of miR-34a and miR-29a. Over-expression of miR-29a, a miRNA expressed in the brain and in cells of the blood lineage, resulted in the differential expression of a set of proteins. Gene Ontology based classification showed that a significant sub-set of these targets, including Voltage Dependent Anion Channel 1 and 2 (VDAC1 and VDAC2) and ATP synthetase, were mitochondrial proteins involved in apoptosis. Using reporter assays, we established that miR-29a targets the 3' Untranslated Regions (3' UTR) of VDAC1 and VDAC2. However, due to the limited number of proteins identified using this approach and the inability to differentiate between primary and secondary effects we conclude that miRNA-proteomics is of limited utility as a high-throughput alternative for sensitive and unbiased miRNA target identification. However, this approach was valuable for rapid assessment of the impact of the miRNAs on the cellular proteome and its biological role in apoptosis. |
| Databáze: | OpenAIRE |
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