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In vitro acylation assay using fractionated HepG2 cell extracts. (a) Schematic diagram of the fractionation of HepG2 cell extracts and in vitro acylation assay. (b) Validation of HepG2 cells fractionated into cytosolic, nuclear, and chromatin extracts. 4 μg of each extract was assessed by western blot (left panel). IKKα, HDAC2, and AcH3 were used as cytosolic, nuclear, and chromatin fraction markers, respectively. CBB staining of loading proteins is shown in the right panel. Cyt: cytosolic extracts; NEs: nuclear extracts; Chr: chromatin extracts. (c) In vitro acylation assays using fractionated cell extracts. ¹⁴C-labeled coenzyme A incorporation was assessed by liquid scintillation counting. Reaction mixtures without cell extracts were used as negative controls (mock). To show that the incorporation was an enzymatic reaction, cell extracts were heat-inactivated at 96°C for 10 min. Ac-CoA: acetyl-coenzyme A; Mal-CoA: malonyl-coenzyme A; Suc-CoA: succinyl-coenzyme A.
Source publication
Posttranslational modification (PTM) of proteins is used to regulate protein activity and stability. Histone PTMs are regarded as some of the most important, as they can directly regulate gene expression through chromatin reorganization. Recently, histone proteins were found to undergo succinylation, adding to other well-known PTMs such as acetylat...
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Citations
... The transfer of the succinyl group is catalyzed by succinyltransferases or non-enzyme catalyzed. For the enzymes, they are a family of enzymes conserved across different species (9)(10)(11). The non-enzymatic catalysis of succinylation may regulate the succinylation of cytoplasmic protein lysine through the activity of cytoplasmic SIRT5 (7). ...
Modifications of protein post-translation are critical modulatory processes, which alters target protein biological activity，function and/or location, even involved in pathogenesis of some diseases. So far, there are at least 16 types of post-translation modifications identified, particularly through recent mass spectrometry analysis. Among them, succinylation (Ksuc) on protein lysine residues causes a variety of biological changes. Succinylation of proteins contributes to many cellular processes such as proliferation, growth, differentiation, metabolism and even tumorigenesis. Mechanically, Succinylation leads to conformation alteration of chromatin or remodeling. As a result, transcription/expression of target genes is changed accordingly. Recent research indicated that succinylation mainly contributes to metabolism modulations, from gene expression of metabolic enzymes to their activity modulation. In this review, we will conclude roles of succinylation in metabolic regulation of glucose, fat, amino acids and related metabolic disease launched by aberrant succinylation. Our goal is to stimulate extra attention to these still not well researched perhaps important succinylation modification on proteins and cell processes.
... It is possible though that the KAT2A succinyltransferase activity is physiologically less important than nonenzymatic succinylation as Anmangandla et al. [33] pointed out that the succinyltransferase activity of KAT2A is low compared to its acetyltransferase activity. Interestingly, using a pan-cancer dataset to study the expression of the succinylation regulators CPT1A, KAT2A, sirtuin 5 (SIRT 5), and SIRT7 showed that especially KAT2A was up-regulated in all types of tumours compared to healthy controls [34]. This indicates that KAT2A might be more important in cancer cells than under physiological conditions. ...
Cells need to adapt their activities to extra- and intracellular signalling cues. To translate a received extracellular signal, cells have specific receptors that transmit the signal to downstream proteins so that it can reach the nucleus to initiate or repress gene transcription. Post-translational modifications (PTMs) of proteins are reversible or irreversible chemical modifications that help to further modulate protein activity. The most commonly observed PTMs are the phosphorylation of serine, threonine, and tyrosine residues, followed by acetylation, glycosylation, and amidation. In addition to PTMs that involve the modification of a certain amino acid (phosphorylation, hydrophobic groups for membrane localisation, or chemical groups like acylation), or the conjugation of peptides (SUMOylation, NEDDylation), structural changes such as the formation of disulphide bridge, protein cleavage or splicing can also be classified as PTMs. Recently, it was discovered that metabolites from the tricarboxylic acid (TCA) cycle are not only intermediates that support cellular metabolism but can also modify lysine residues. This has been shown for acetate, succinate, and lactate, among others. Due to the importance of mitochondria for the overall fitness of organisms, the regulatory function of such PTMs is critical for protection from aging, neurodegeneration, or cardiovascular disease. Cancer cells and activated immune cells display a phenotype of accelerated metabolic activity known as the Warburg effect. This metabolic state is characterised by enhanced glycolysis, the use of the pentose phosphate pathway as well as a disruption of the TCA cycle, ultimately causing the accumulation of metabolites like citrate, succinate, and malate. Succinate can then serve as a signalling molecule by directly interacting with proteins, by binding to its G protein-coupled receptor 91 (GPR91) and by post-translationally modifying proteins through succinylation of lysine residues, respectively. This review is focus on the process of protein succinylation and its importance in health and disease.
... The importance of succinylation ranges from coupling metabolism with protein function in the nervous system [58,59], association with energy regulation and several cellular metabolic processes including glucose metabolism [56,60,61]. Histone proteins that may directly regulate gene expression via chromatin reorganization are subject to acetylation and succinylation as a major post translational modification process [62]. Thus, alterations in succinate that we describe in our clinical groups suggest that these important succinylation pathways may be compromised in AD. ...
Background
We previously identified urine dicarboxylic acids (DCA) of carbon length 4‐9 as candidate biomarkers of probable Alzheimer’s disease (AD) and energy dysregulation, and found their levels correlated with hippocampal volume. Our aim was to evaluate these initial findings in a second cohort.
Method
Study participants (> 65 years) were classified clinically as cognitively healthy (CH, n=188) or with probable AD (n=66) based on Uniform Data System‐3 of the National Alzheimer’s Coordinating Center criteria. Cerebrospinal fluid (CSF) amyloid ß 42 (A) and phospho‐ tau (T) levels were measured by validated immunoassay. Urine was collected in a “spot” sample after a 12 h overnight fast; target DCAs were detected using gas chromatography coupled with negative ion chemical ionization mass spectrometry and each DCA was normalized to deuterated internal standards and to a percent of total fatty acids.
Result
The mean of short‐chain DCA’s (C4‐C5) were higher in CH (44.38% ±17.79) compared to AD individuals (34.79% ±12.39) (p=0.0002), while the long‐chain DCA’s (C7‐C9) were lower for CH (34.70% ±16.88) when compared to AD individuals (45.22% ±15.11) (p<0.0001). DCA levels in the subset of CH participants with A ⁺ /T ⁺ were in a transition to the levels of the AD group and C4‐C5 and C7‐C9 differed from those with A ‐ /T ‐ biomarkers (p=0.0001). Combining results into a ratio of short‐ to long‐chain DCAs distinguished the A ‐ /T ‐ (2.330 ±2.144) from the A ⁺ /T ⁺ group (1.178 ±0.946); p<0.0001. ROC curve of the DCA ratio between CH‐NAT and AD is 82%. Study participant clinical and DCA groups were not significantly correlated with the common confounders of age, sex, education or ApoE genotype.
Conclusion
Our data extends our initial finding that urine DCA measures have potential as biomarkers of AD pathology, including for cognitively healthy individuals who are positive for CSF AT biomarkers. We propose that measurement of urine DCAs provides a new approach to screen for metabolic fluctuations in early AD and to monitor the effect of novel therapies in clinical trials. While studies are needed, including with other populations, varied clinical classifications, simplified chemistry, this data supports DCA biomarkers for widespread screening since the necessary urine sample is available from everyone.
... Some studies have shown that abnormalities and variations of succinylation associated with the pathogenesis of many diseases, including tumors [6][7][8][9][10], cardiac metabolic diseases [11,12], liver metabolic diseases [13], and nervous system diseases [7,14,15]. Therefore, understanding succinylation and identifying the site of succinylation will help determine the pathogenesis of related diseases and develop targeted drugs [16]. ...
Succinylation is an important posttranslational modification of proteins, which plays a key role in protein conformation regulation and cellular function control. Many studies have shown that succinylation modification on protein lysine residue is closely related to the occurrence of many diseases. To understand the mechanism of succinylation profoundly, it is necessary to identify succinylation sites in proteins accurately. In this study, we develop a new model, IFS-LightGBM (BO), which utilizes the incremental feature selection (IFS) method, the LightGBM feature selection method, the Bayesian optimization algorithm, and the LightGBM classifier, to predict succinylation sites in proteins. Specifically, pseudo amino acid composition (PseAAC), position-specific scoring matrix (PSSM), disorder status, and Composition of k-spaced Amino Acid Pairs (CKSAAP) are firstly employed to extract feature information. Then, utilizing the combination of the LightGBM feature selection method and the incremental feature selection (IFS) method selects the optimal feature subset for the LightGBM classifier. Finally, to increase prediction accuracy and reduce the computation load, the Bayesian optimization algorithm is used to optimize the parameters of the LightGBM classifier. The results reveal that the IFS-LightGBM (BO)-based prediction model performs better when it is evaluated by some common metrics, such as accuracy, recall, precision, Matthews Correlation Coefficient (MCC), and F-measure.
... They make up the chromatin within the nucleus and play a key role in a variety of processes including transcription, DNA replication, DNA repair and recombination, chromosome stability, germline DNA packaging, PTM through ADP-ribosylation by PARP1 and functions related to the acrosome during spermiogenesis (Mariño-Ramírez et al., 2005;Ooi and Henikoff, 2007;Messner and Hottiger, 2011;De Vries et al., 2012;Venkatesh and Workman, 2015). Although well known for PTM through acetylation and deacetylation, histones can also undergo PTM by phosphorylation, methylation, ubiquitination, formylation, succinylation and citrullination (Cheung, Allis and Sassone-Corsi, 2000;Nakashima, Hagiwara and Yamada, 2002;Cuthbert et al., 2004;Jiang et al., 2007;Shahbazian and Grunstein, 2007;Wiśniewski, Zougman and Mann, 2008;Wang et al., 2009;Zhang et al., 2011;Cao and Yan, 2012;Xie et al., 2012;Greer and Shi, 2012;Rossetto, Avvakumov and Côté, 2012;Yokoyama, Katsura and Sugawara, 2017). This can disrupt the interactions between the histone proteins and DNA as well as between the core histone components, affecting histone-histone interactions. ...
... The importance of succinylation ranges from coupling metabolism with protein function in the nervous system [58,59], association with energy regulation and several cellular metabolic processes including glucose metabolism [56,60,61]. Histone proteins that may directly regulate gene expression via chromatin reorganization are subject to acetylation and succinylation as a major post translational modification process [62]. Thus, alterations in succinate that we describe in our clinical groups suggest that these important succinylation pathways may be compromised in AD. ...
Non-invasive biomarkers will enable widespread screening and early diagnosis of Alzheimer’s disease (AD). We hypothesized that the considerable loss of brain tissue in AD will result in detection of brain lipid components in urine, and that these will change in concert with CSF and brain biomarkers of AD. We examined urine dicarboxylic acids (DCA) of carbon length 3–10 to reflect products of oxidative damage and energy generation or balance that may account for changes in brain function in AD. Mean C4-C5 DCAs were lower and mean C7-C10 DCAs were higher in the urine from AD compared to cognitively healthy (CH) individuals. Moreover, mean C4-C5 DCAs were lower and mean C7-C9 were higher in urine from CH individuals with abnormal compared to normal CSF amyloid and Tau levels; i.e., the apparent urine changes in AD also appeared to be present in CH individuals that have CSF risk factors of early AD pathology. In examining the relationship between urine DCAs and AD biomarkers, we found short chain DCAs positively correlated with CSF Aβ42, while C7-C10 DCAs negatively correlated with CSF Aβ42 and positively correlated with CSF Tau levels. Furthermore, we found a negative correlation of C7-C10 DCAs with hippocampal volume (p < 0.01), which was not found in the occipital volume. Urine measures of DCAs have an 82% ability to predict cognitively healthy participants with normal CSF amyloid/Tau. These data suggest that urine measures of increased lipoxidation and dysfunctional energy balance reflect early AD pathology from brain and CSF biomarkers. Measures of urine DCAs may contribute to personalized healthcare by indicating AD pathology and may be utilized to explore population wellness or monitor the efficacy of therapies in clinical trials.
... Protein folding and final structure including biochemical activity, half-life and stability are determined by the primary sequence of a protein (Marks et al. 2012). However, during the life span, the proteome of a protein would be two or three folds more complex rather than encoding genomes (Yokoyama et al. 2017). PTM, a proteome expansion route, is present in eukaryotes and prokaryotes but the exposure of PTMs is more common in eukaryotic cells. ...
Post-translational modifications (PTMs) increase proteome activity for controlling every feature of normal cell biology. PTMs such as phosphorylation, acetylation, glycosylation, fatty acylation, palmitoylation, myristoylation, ubiquitination, SUMOylation (small ubiquitin-like modifiers), methylation, deamidation, nitrosylation, etc. of proteins can regulate the properties of protein including intracellular distribution , functionality, stability, accumulation, as well as interactions. PTMs take place at any stage of the protein life cycle, regulating protein folding and activity in time and space, subcellular localization of the protein, and their activity. Hence, PTMs play a pivotal role in the regulation of numerous cellular processes. Abnormal PTMs of one or more culprit proteins might contribute to neurodegeneration, which is shown in some
... This indicates that H3K79Suc may act as an activating mark for gene transcription [34]. In independent support of enzymatic succinylation, nuclear protein extracts from HepG2 cells separated by strong cation exchange chromatography showed that a specific fraction (not those containing the HAT p300/ CBP) of purified proteins could catalyse succinylation of purified histone proteins in a manner that was sensitive to heat inactivation and was decreased by competition with acetyl-CoA, supporting the idea of protein dependent catalysis of histone succinylation [72]. Lysine succinyletransferase activity has also been proposed for OGDH [73] in the mitochondria and CPT1 [30] on the outer mitochondrial outer membrane, although the physiological relevance of these activities is not clear. ...
Background
Many metabolites serve as important signalling molecules to adjust cellular activities and functions based on nutrient availability. Links between acetyl-CoA metabolism, histone lysine acetylation, and gene expression have been documented and studied over the past decade. In recent years, several additional acyl modifications to histone lysine residues have been identified, which depend on acyl-coenzyme A thioesters (acyl-CoAs) as acyl donors. Acyl-CoAs are intermediates of multiple distinct metabolic pathways, and substantial evidence has emerged that histone acylation is metabolically sensitive. Nevertheless, the metabolic sources of acyl-CoAs used for chromatin modification in most cases remain poorly understood. Elucidating how these modifications are coupled to and regulated by cellular metabolism is important in deciphering the functional and contextual significance of these diverse chemical modifications.
Scope of review
In this article, we review the metabolic pathways that produce acyl-CoAs, as well as emerging evidence for functional roles of diverse acyl-CoAs in chromatin regulation. Because acetyl-CoA has been extensively reviewed elsewhere, we will focus on four other acyl-CoA metabolites integral to major metabolic pathways that are also known to modify histones: succinyl-CoA, propionyl-CoA, crotonoyl-CoA, and butyryl-CoA. We also briefly mention several other acyl-CoA species, which present opportunities for further research; malonyl-CoA, glutaryl-CoA, 3-hydroxybutyryl-CoA, 2-hydroxyisobutyryl-CoA, and lactyl-CoA. Each acyl-CoA species has distinct roles in metabolism, indicating the potential to report shifts in the metabolic status of the cell. For each metabolite, we consider the metabolic pathways in which it participates and the nutrient sources from which it is derived, the compartmentalization of its metabolism, and the factors reported to influence its abundance and potential nuclear availability. We also briefly highlight reported mechanisms of regulation and the biological functions of these metabolically-linked acylation marks. Finally, we aim to illuminate key questions in acyl-CoA metabolism as they relate to the control of chromatin modification.
Major conclusions
A majority of acyl-CoA species are annotated to mitochondrial metabolic processes. Since acyl-CoAs are not known to be directly transported across mitochondrial membranes, they must be synthesized outside of mitochondria and potentially within the nucleus to participate in chromatin regulation. Thus, subcellular metabolic compartmentalization likely plays a key role in the regulation of histone acylation. Metabolite tracing in combination with targeting of relevant enzymes and transporters will help to elucidate the metabolic pathways that connect acyl-CoA metabolism to chromatin modification. The specific function of each acyl-CoA may be determined in part by biochemical properties that affect their propensity for enzymatic versus non-enzymatic protein modification, as well as the various enzymes that can add, remove and bind each modification. Further, competitive and inhibitory effects of different acyl-CoA species on these enzymes make determining the relative abundance of acyl-CoA species in specific contexts important to understand the regulation of chromatin acylation. An improved and more nuanced understanding of metabolic regulation of chromatin and its roles in physiological and disease-related processes will emerge as these questions are answered.
... Lysine succinylation is an evolutionarily conserved posttranslational modification (PTM) known to be involved in the regulation of diverse cellular process [1][2][3][4][5][6][7]. The succinylation process modifies a target protein with a succinyl group (-CO-CH2-CH2-CO2H), which is transmitted from succinyl-CoA to the specific α-amino group of a lysine residue [8][9][10][11][12]. The succinylation firstly was discovered in histone protein [13], and its regulatory role has been examined through the gene expression regarding chromatin reorganization [14][15][16]. ...
... Nevertheless, the published studies have provided little information regarding the enzyme which catalyzes histone lysine succinylation [17][18][19]. In fact, it is unclear whether this reaction is enzymatic or not [8,9,20]. In addition to histones, the succinylated proteins were found in the cytoplasm, nucleus, and mitochondria [7,[21][22][23][24], indicating that lysine succinylation controls a variety of biological functions [14,18,25,26]. ...
Lysine succinylation is a form of posttranslational modification of the proteins that play an essential functional role in every aspect of cell metabolism in both prokaryotes and eukaryotes. Aside from experimental identification of succinylation sites, there has been an intense effort geared towards the development of sequence-based prediction through machine learning, due to its promising and essential properties of being highly accurate, robust and cost-effective. In spite of these advantages, there are several problems that are in need of attention in the design and development of succinylation site predictors. Notwithstanding of many studies on the employment of machine learning approaches, few articles have examined this bioinformatics field in a systematic manner. Thus, we review the advancements regarding the current state-of-the-art prediction models, datasets, and online resources and illustrate the challenges and limitations to present a useful guideline for developing powerful succinylation site prediction tools.
Lysine succinylation is a post-translational modification (PTM) of protein in which a succinyl group (-CO-CH2-CH2-CO2H) is added to a lysine residue of protein that reverses lysine's positive charge to a negative charge and leads to the significant changes in protein structure and function. It occurs on a wide range of proteins and plays an important role in various cellular and biological processes in both eukaryotes and prokaryotes. Beyond experimentally identified succinylation sites, there have been a lot of studies for developing sequence-based prediction using machine learning approaches, because it has the promise of being extremely time-saving, accurate, robust, and cost-effective. Despite of these benefits on computational prediction of lysine succinylation sites for different species, there are a number of issues that need to be addressed in the design and development of succinylation site predictors. In spite the fact that many studies used different statistical and machine learning computational tools, only a few studies have focused on these bioinformatics issues in depth. Therefore, in this comprehensive comparative review, an attempt is made to present the latest advances in the prediction models, datasets, and online resources, as well as the obstacles and limits, to provide an advantageous guideline for developing more suitable and effective succinylation site prediction tools.
