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Monitoring mitochondrial translation by pulse SILAC

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Item Type:Article
Title:Monitoring mitochondrial translation by pulse SILAC
Creators Name:Imami, K. and Selbach, M. and Ishihama, Y.
Abstract:Mitochondrial ribosomes are specialized to translate the 13 membrane proteins encoded in the mitochondrial genome, which shapes the oxidative phosphorylation (OXPHOS) complexes essential for cellular energy metabolism. Despite the importance of mitochondrial translation control, it is challenging to identify and quantify the mitochondrial-encoded proteins due to their hydrophobic nature and low abundance. Here, we introduce a mass spectrometry-based proteomic method that combines biochemical isolation of mitochondria with pulse stable isotope labeling by amino acids in cell culture (pSILAC). Our method provides the highest protein identification rate with the shortest measurement time among currently available methods, enabling us to quantify 12 out of the 13 mitochondrial-encoded proteins. We applied this method to uncover the global picture of (post-)translational regulation of both mitochondrial- and nuclear-encoded subunits of OXPHOS complexes. We found that inhibition of mitochondrial translation led to degradation of orphan nuclear-encoded subunits that are considered to form subcomplexes with the mitochondrial-encoded subunits. This method should be readily applicable to study mitochondrial translation programs in many contexts, including oxidative stress and mitochondrial disease.
Keywords:Proteomics, Pulse SILAC, Translation, Mitochondria, OXPHOS, Protein Complex
Source:Journal of Biological Chemistry
ISSN:1083-351X
Publisher:American Society for Biochemistry and Molecular Biology
Volume:299
Number:2
Page Range:102865
Date:February 2023
Official Publication:https://doi.org/10.1016/j.jbc.2022.102865
PubMed:View item in PubMed
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https://edoc.mdc-berlin.de/19967/Final version

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