Proteomic Analysis of Lysine Acetylation Sites in Rat Tissues Reveals Organ Specificity and Subcellular Patterns

Novo Nordisk Foundation Center for Protein Research, Department for Proteomics, Faculty of Health Sciences, University of Copenhagen, Denmark.
Cell Reports (Impact Factor: 8.36). 08/2012; 2(2):419-31. DOI: 10.1016/j.celrep.2012.07.006
Source: PubMed


Lysine acetylation is a major posttranslational modification involved in a broad array of physiological functions. Here, we provide an organ-wide map of lysine acetylation sites from 16 rat tissues analyzed by high-resolution tandem mass spectrometry. We quantify 15,474 modification sites on 4,541 proteins and provide the data set as a web-based database. We demonstrate that lysine acetylation displays site-specific sequence motifs that diverge between cellular compartments, with a significant fraction of nuclear sites conforming to the consensus motifs G-AcK and AcK-P. Our data set reveals that the subcellular acetylation distribution is tissue-type dependent and that acetylation targets tissue-specific pathways involved in fundamental physiological processes. We compare lysine acetylation patterns for rat as well as human skeletal muscle biopsies and demonstrate its general involvement in muscle contraction. Furthermore, we illustrate that acetylation of fructose-bisphosphate aldolase and glycerol-3-phosphate dehydrogenase serves as a cellular mechanism to switch off enzymatic activity.

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Available from: Alicia Lundby
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    • "The protein extraction of the rat liver tissues was performed according to previous report [32] and digested by trypsin (Promega) with the second digestion method to ensure thorough digestion [33]. Briefly, the liver tissues were first grinded by liquid nitrogen and the powder was transferred to 5 mL centrifuge tube and precipitated with cold 10 % TCA/acetone supplemented with 50 mM DTT, 0.1 % Protease Inhibitor Cocktail Set IV and HDACinhibitor (50 mM sodium butyrate, 30 mM nicotinamide, and 3 μM Trichostatin A) for 2 h at −20 °C. "
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    ABSTRACT: Non-alcoholic fatty liver disease (NAFLD) is a clinical frequent disease. However, its pathogenesis still needs further study, especially the mechanism at the molecular level. The recent identified novel protein post-translational modification, lysine succinylation was reported involved in diverse metabolism and cellular processes. In this study, we performed the quantitative succinylome analysis in the liver of NAFLD model to elucidate the regulatory role of lysine succinylation in NAFLD progression. Firstly, experimental model of NAFLD was induced by carbon tetrachloride injection and supplementary high-lipid and low-protein diet. Then series histochemical and biochemical variables were determined. For the quantitative succinylome analysis, tandem mass tags (TMT)-labeling, highly sensitive immune-affinity purification, liquid chromatography-tandem mass spectrometry techniques were applied. Bioinformatics analysis including gene ontology annotation based classification; Wolfpsort based subcellular prediction; function enrichment; protein-protein interaction network construction and conserved succinylation site motifs extraction were performed to decipher the differentially changed succinylated proteins and sites and p-value < 0.05 was selected as threshold. Totally, 815 succinylation sites on 407 proteins were identified, of which 243 succinylation acetylation sites on 178 proteins showed changed succinylation level with the threshold fold change > 1.5. Theses differentially changed succinylated proteins were involved in diverse metabolism pathways and cellular processes including carbon metabolism, amino acid metabolism, fat acid metabolism, binding and catalyzing, anti-oxidation and xenobiotics metabolism. Besides, these differentially changed succinylated proteins were prominently localized to cytoplasm and mitochondria. Moreover, 8 conserved succinylation site motifs were extracted around the succinylation sites. Protein succinylation was an extensive post-translation modification in rat. The changed succinylation level in diverse proteins may disturb multiple metabolism pathways and promote non-alcoholic fatty liver disease development. This study provided a basis for further characterization of the pathophysiological role of lysine succinylation in NAFLD progression, which laid a foundation for the innovation of novel NAFLD drugs and therapies.
    Preview · Article · Dec 2016 · Proteome Science
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    • "The role of acetylation in regulating chromatin structure and gene transcription is well studied with both lysine acetyltransferases and deacetylases taking part (Shahbazian & Grunstein, 2007). Mass spectrometry (MS), combined with acetylated peptide enrichment, has enabled the identification of thousands of acetylation sites on diverse proteins (Kim et al, 2006; Choudhary et al, 2009; Lundby et al, 2012), implicating acetylation in regulating various cellular processes. Metabolic proteins, particularly in mitochondria, were found to be frequently acetylated, and a multitude of studies have investigated the role of acetylation in regulating metabolism (Wang et al, 2010; Zhao et al, 2010; Newman et al, 2012). "
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    ABSTRACT: Acetylation is frequently detected on mitochondrial enzymes, and the sirtuin deacetylase SIRT3 is thought to regulate metabolism by deacetylating mitochondrial proteins. However, the stoichiometry of acetylation has not been studied and is important for understanding whether SIRT3 regulates or suppresses acetylation. Using quantitative mass spectrometry, we measured acetylation stoichiometry in mouse liver tissue and found that SIRT3 suppressed acetylation to a very low stoichiometry at its target sites. By examining acetylation changes in the liver, heart, brain, and brown adipose tissue of fasted mice, we found that SIRT3-targeted sites were mostly unaffected by fasting, a dietary manipulation that is thought to regulate metabolism through SIRT3-dependent deacetylation. Globally increased mitochondrial acetylation in fasted liver tissue, higher stoichiometry at mitochondrial acetylation sites, and greater sensitivity of SIRT3-targeted sites to chemical acetylation in vitro and fasting-induced acetylation in vivo, suggest a nonenzymatic mechanism of acetylation. Our data indicate that most mitochondrial acetylation occurs as a low-level nonenzymatic protein lesion and that SIRT3 functions as a protein repair factor that removes acetylation lesions from lysine residues.
    Full-text · Article · Sep 2015 · The EMBO Journal
    • "This may contribute to the elevated variability in lysine acetylation patterns encountered in organs and subcellular fractions in distinct functional states (Lundby et al., 2012). Here, we discuss the ability of acetyl-CoA to regulate adaptive responses to homeostatic perturbations by controlling the equilibrium between catabolic and anabolic reactions as well as by influencing major cellular processes such as cell cycle progression , mitosis, autophagy, and regulated cell death (RCD). "
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    ABSTRACT: Acetyl-coenzyme A (acetyl-CoA) is a central metabolic intermediate. The abundance of acetyl-CoA in distinct subcellular compartments reflects the general energetic state of the cell. Moreover, acetyl-CoA concentrations influence the activity or specificity of multiple enzymes, either in an allosteric manner or by altering substrate availability. Finally, by influencing the acetylation profile of several proteins, including histones, acetyl-CoA controls key cellular processes, including energy metabolism, mitosis, and autophagy, both directly and via the epigenetic regulation of gene expression. Thus, acetyl-CoA determines the balance between cellular catabolism and anabolism by simultaneously operating as a metabolic intermediate and as a second messenger. Copyright © 2015 Elsevier Inc. All rights reserved.
    No preview · Article · Jun 2015 · Cell metabolism
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