A genome-wide shRNA screen for new OxPhos related genes

• Published in: Mitochondrion 2011-05, Vol.11 (3), p.467-475
• Main Authors: Bayona-Bafaluy, María Pilar, Sánchez-Cabo, Fátima, Fernández-Silva, Patricio, Pérez-Martos, Acisclo, Enríquez, José Antonio
• Format: Article
• Language: English
• Subjects: Animals; Cell Line; Electron Transport Chain Complex Proteins - genetics; Electron Transport Chain Complex Proteins - metabolism; Fibroblasts - metabolism; Genetic Vectors; High-Throughput Screening Assays - methods; Lentivirus - genetics; Mice; Mitochondria - enzymology; Mitochondria - metabolism; Oxidative Phosphorylation; Oxidoreductases - antagonists & inhibitors; Oxidoreductases - genetics; RNA Interference; RNA, Small Interfering - genetics; RNA, Small Interfering - metabolism; Transduction, Genetic
• ISSN: 1567-7249; 1872-8278
• DOI: 10.1016/j.mito.2011.01.007

The mitochondrial oxidative phosphorylation (OxPhos) system produces most of the ATP required by the cell. The structural proteins of the OxPhos holoenzymes are well known, but important aspects of their biogenesis and regulation remain to be uncovered and a significant fraction of mitochondrial proteins have yet to be identified. We have used a high throughput, genome-wide RNA interference (RNAi) approach to identify new OxPhos-related genes. We transduced a mouse fibroblast cell line with a lentiviral-based shRNA-library, and screened the cell population for growth impairment in galactose-based medium, which requires an intact OxPhos system. Candidate genes were ranked according to their co-expression with known genes encoding OxPhos mitochondria-located proteins. For the top ranking candidates the cellular process in which they are involved was evaluated. Our results show that the use of genome-wide RNAi together with screening for deficient growth in galactose medium is a suitable approach to identifying OxPhos-related and cellular energy metabolism-related genes. Interestingly also ubiquitin-proteasome related genes were selected.