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Label-Free Quantitative Proteomics in a Methylmalonyl-CoA Mutase-Silenced Neuroblastoma Cell Line
- Source :
- International Journal of Molecular Sciences, Vol 19, Iss 11, p 3580 (2018)
- Publication Year :
- 2018
- Publisher :
- MDPI AG, 2018.
-
Abstract
- Methylmalonic acidemias (MMAs) are inborn errors of metabolism due to the deficient activity of methylmalonyl-CoA mutase (MUT). MUT catalyzes the formation of succinyl-CoA from methylmalonyl-CoA, produced from propionyl-CoA catabolism and derived from odd chain fatty acids β-oxidation, cholesterol, and branched-chain amino acids degradation. Increased methylmalonyl-CoA levels allow for the presymptomatic diagnosis of the disease, even though no approved therapies exist. MMA patients show hyperammonemia, ketoacidosis, lethargy, respiratory distress, cognitive impairment, and hepatomegaly. The long-term consequences concern neurologic damage and terminal kidney failure, with little chance of survival. The cellular pathways affected by MUT deficiency were investigated using a quantitative proteomics approach on a cellular model of MUT knockdown. Currently, a consistent reduction of the MUT protein expression was obtained in the neuroblastoma cell line (SH-SY5Y) by using small-interfering RNA (siRNA) directed against an MUT transcript (MUT siRNA). The MUT absence did not affect the cell viability and apoptotic process in SH-SY5Y. In the present study, we evaluate and quantify the alterations in the protein expression profile as a consequence of MUT-silencing by a mass spectrometry-based label-free quantitative analysis, using two different quantitative strategies. Both quantitative methods allowed us to observe that the expression of the proteins involved in mitochondrial oxido-reductive homeostasis balance was affected by MUT deficiency. The alterated functional mitochondrial activity was observed in siRNA_MUT cells cultured with a propionate-supplemented medium. Finally, alterations in the levels of proteins involved in the metabolic pathways, like carbohydrate metabolism and lipid metabolism, were found.
Details
- Language :
- English
- ISSN :
- 14220067
- Volume :
- 19
- Issue :
- 11
- Database :
- Directory of Open Access Journals
- Journal :
- International Journal of Molecular Sciences
- Publication Type :
- Academic Journal
- Accession number :
- edsdoj.35a4f0704bbb44aa9a3fbc619000fc32
- Document Type :
- article
- Full Text :
- https://doi.org/10.3390/ijms19113580