Succinylation is a relatively common post-translational modification. It occurs in the cytoplasm, mitochondria, and the nucleus, where its essential precursor, succinyl-CoA, is present, allowing for the modification of non-histone and histone proteins. In normal cells, succinylation levels are carefully regulated to sustain a dynamic balance, necessitating the involvement of various regulatory mechanisms, including non-enzymatic reactions, succinyltransferases, and desuccinylases. Among these regulatory factors, sirtuin 5, the first identified desuccinylase, plays a significant role and has been extensively researched. The level of succinylation has a significant effect on multiple metabolic pathways, including the tricarboxylic acid cycle, redox balance, and fatty acid metabolism. Dysregulated succinylation can contribute to the progression or exacerbation of various urinary diseases. Succinylation predominantly affects disease progression by altering the expression of key genes and modulating the activity of enzymes involved in vital metabolic processes. Desuccinylases primarily affect enzymes associated with Warburg's effect, thereby affecting the energy supply of tumor cells, while succinyltransferases can regulate gene transcription to alter cell phenotype, thereby involving the development of urinary diseases. Considering these effects, targeting succinylation-related enzymes to regulate metabolic pathways or gene expression may offer a promising therapeutic strategy for treating urinary diseases.

Environmental factors may affect gene expression through epigenetic modifications of histones and transcription factors. Here, we report that cellular uptake of sorbate, a common food preservative, induces lysine sorbylation (Ksor) in mammalian cells and tissue mediated by the noncanonical activities of class I histone deacetylases (HDAC1-3). We demonstrated that HDAC1-3 catalyze sorbylation upon sorbate uptake and desorbylation in the absence of sorbate both in vitro and in cells. Sorbate uptake in mice livers significantly induced histone Ksor, correlating with decreased expressions of inflammation-response genes. Accordingly, sorbate treatment in macrophage RAW264.7 cells upon lipopolysaccharide (LPS) stimulation dose-dependently down-regulated proinflammatory gene expressions and nitric oxide production. Proteomic profiling identified RelA, a component of the NF-κB complex, and its interacting proteins as bona fide Ksor targets and sorbate treatment significantly decreased NF-κB transcriptional activities in response to LPS stimulation in RAW264.7 cells. Together, our study demonstrated a noncanonical mechanism of sorbate uptake in regulating epigenetic histone modifications and inflammatory gene expression.

Diabetic nephropathy (DN) remains difficult to treat due to its complex mechanisms. This study explores the ferroptosis mechanism in DN, focusing on the regulation of SAT2 expression by KAT2A-mediated H3K79 succinylation (H3K79succ).

A DN rat model was created using streptozotocin (STZ) and a high-fat diet (HFD). KAT2A expression in rat kidney tissue was analyzed by RT-qPCR, WB, and immunohistochemistry. Renal pathology and function were assessed, and ferroptosis markers (ROS, GSH, MDA, and iron content) were measured. A high-glucose-induced HPo cell model was used for in vitro validation. KAT2A knockdown and CUT&Tag/RNA-seq were used to identify potential targets, and the regulation of SAT2 by KAT2A was confirmed through RT-qPCR, WB, and ChIP-qPCR.

Elevated KAT2A and H3K79succ expression were observed in DN rat kidney tissues and HPo cells. KAT2A knockdown reversed kidney damage, improved renal function, and suppressed inflammation and ferroptosis. CUT&Tag and RNA-seq identified SAT2 as a KAT2A target, and we confirmed that KAT2A-mediated H3K79succ enhances SAT2 expression, promoting ferroptosis in DN.

This study uncovers the role of KAT2A in DN, demonstrating its promotion of inflammation and ferroptosis through H3K79succ and SAT2 upregulation, offering insights into DN pathogenesis and potential therapeutic strategies.

Porcine deltacoronavirus (PDCoV) is an emerging enteropathogenic coronavirus capable of infecting various animal species, including humans. In this study, we explored the roles of sirtuins (SIRTs), a conserved family of protein deacylases and mono-adenosine diphosphate-ribosyltransferases, in PDCoV replication. Surprisingly, we found that SIRT5-a unique member of SIRTs with distinct desuccinylation, demalonylation, and deglutarylation activities-is a proviral factor essential for PDCoV replication; its catalytic activities are crucial in this process. Mechanistically, SIRT5 interacts with and desuccinylates the PDCoV membrane (M) protein. This modification activates the ataxia-telangiectasia mutated (ATM) pathway, facilitates ubiquitination of peroxisomal biogenesis protein 5 (PEX5), and recruits sequestosome 1 (SQSTM1/p62) to initiate selective peroxisomal autophagy (pexophagy). The pexophagy process disrupts peroxisomal function, elevates reactive oxygen species (ROS) levels, and suppresses type I and III interferon production, thereby enhancing viral replication. We also identified lysine 207 (K207) as the primary succinylation site of the M protein. Mutations mimicking the desuccinylated or succinylated states of K207 substantially influence viral replication and the ability to induce pexophagy. These findings reveal a novel role for SIRT5 in regulating pexophagy during viral infection and suggest a therapeutic target for efforts to combat coronavirus infections.

Deregulation of amino acid (AA) metabolism has been reported in several pathological conditions, including metabolic diseases (e.g., obesity and diabetes), cardiovascular diseases, and cancer. However, the role of alterations in AA levels in chronic liver disorders such as metabolic dysfunction-associated steatotic liver disease (MASLD) remains largely unexplored. In this study we aimed to evaluate the hepatic AA composition in patients with different stages of MASLD, and their relationship with MASLD-related risk factors. A case-control study was conducted in 40 patients with obesity undergoing bariatric surgery at Virgen de la Arrixaca University Hospital (Murcia, Spain), where MASLD diagnosis was confirmed by histological analysis of liver biopsies, and hepatic AA levels were measured using ultra-performance liquid chromatography high-resolution time-of-flight mass spectrometry. Our results revealed that the hepatic AA profile was significantly altered in patients with MASLD. More specifically, comparison between MASLD patients revealed a significant increase in hepatic levels of arginine, glycine and cystine in MASH samples compared to steatotic livers. In addition, hepatic concentrations of arginine, lysine and cystine positively correlated with histopathological diagnosis and other MASLD-related parameters, including transaminases and CK-18 levels. These findings suggest that alterations in certain hepatic AA levels such as arginine, lysine, glycine and cystine in MASLD patients could have translational relevance in understanding the onset of this disease.

Allergic airway inflammation (AAI) is a prevalent respiratory disorder that affects a vast number of individuals globally. There exists a complex interplay among inflammation, immune responses, and metabolic processes, which is of paramount importance in the pathogenesis of AAI. Metabolic dysregulation and protein translational modification (PTM) are well-recognized hallmarks of diseases, playing pivotal roles in the onset and progression of numerous ailments. However, the role of gut microbiota metabolites in the development of AAI, as well as their influence on PTM modifications within this disease context, have not been thoroughly explored and investigated thus far. In AAI patients, succinate was identified as a key metabolite, positively correlated with certain immune parameters and IgE levels, and having good diagnostic value. In AAI mice, gut bacteria were the main source of high succinate levels. Mendelian randomization showed succinate as a risk factor for asthma. Exogenous succinate worsened AAI in mice, increasing airway resistance and inflammatory factor levels. Protein succinylation in AAI mice lungs differed significantly from normal mice, with up-regulated proteins in metabolic pathways. FMT alleviated AAI symptoms by reducing succinate and protein succinylation levels. In vitro, succinate promoted protein succinylation in BEAS-2B cells, and SOD2 was identified as a key succinylated protein, with the K68 site crucial for its modification and enzyme activity regulation. Gut flora-derived succinate exacerbates AAI in mice by increasing lung protein succinylation, and FMT can reverse this. These findings offer new insights into AAI mechanisms and potential therapeutic targets.

Succinate dehydrogenase (SDH), considered as the linkage between tricarboxylic acid cycle (TCA cycle) and electron transport chain, plays a vital role in adenosine triphosphate (ATP) production and cell physiology. SDH deficiency is a notable characteristic in many cancers. Recent studies have pinpointed the dysregulation of SDH can directly result its decreased catalytic activity and the accumulation of oncometabolite succinate, promoting tumor progression in different perspectives. This article expounds the various types of SDH deficiency in tumors and the corresponding pathological features. In addition, we discuss the mechanisms through which defective SDH fosters carcinogenesis, pioneering a categorization of these mechanisms as being either succinate-dependent or independent. Since SDH-deficient and cumulative succinate are regarded as the typical features of some cancers, like gastrointestinal stromal tumors, pheochromocytomas and paragangliomas, we summarize the presented medical management of SDH-deficient tumor patients in clinical and preclinical, identifying the potential strategies for future cancer therapeutics.

Intramembranous ossification, a major bone development process, begins with the condensation of precursor cells through the timely structural adaption of extracellular matrix (ECM) catering to rapid cellular morphological changes. Inspired by this, we design a highly cell-adaptable hydrogel to recapitulate an ECM-dependent mechanotransduction-metabolism-epitranscriptomics axis in mesenchymal stromal cells (MSCs). This hydrogel significantly enhances the E-cadherin-mediated cell-cell interactions of MSCs and promotes glucose uptake and tricarboxylic acid (TCA) cycle activities. We further show that elevated succinate inhibits fat mass and obesity-associated protein (FTO), a N6-methyladenosine (m6A) demethylase, thereby enhancing methyltransferase-like 3 (METTL3)-driven m6A methylation. Methylated RNA immunoprecipitation sequencing (MeRIP-seq) indicates increased m6A methylation of runt-related transcription 2 (Runx2), a key osteogenic signaling factor, promoting osteogenesis of hydrogel-delivered MSCs and bone regeneration in critical-sized bone defects. Our findings reveal the mechanism underlying the critical impact of adaptable ECM structures on tissue development and provide valuable guidance for the design of ECM-mimetic cell carriers to enhance the therapeutic outcomes of regenerative medicine.

A new pathogenic role for mitochondrial dysfunction has been associated with the development of idiopathic pulmonary fibrosis (IPF). Lysine succinylation (Ksucc) is involved in many energy metabolism pathways in mitochondria, making Ksucc highly valuable for studying IPF. We used liquid chromatography with tandem mass spectrometry (LC-MS/MS) to perform the first global profiling of Ksucc in fibrotic lung tissues from IPF patients, providing a proof of concept for the alteration of Ksucc in IPF and highlighting its potential as a therapeutic target. Selected candidate proteins were further verified by targeted proteomics using parallel reaction monitoring (PRM). In this study, we identified 1964 Ksucc sites on 628 modified proteins, with675 of these Ksucc sites on 124 modified proteins closely related to mitochondrial metabolism. 117 succinylated proteins were associated with energy metabolism in mitochondria by comparing these proteins with those previously reported in normal lung tissues. The Ksucc levels in KYAT3, HSD17B8, GRHPR, and IDH2 were different between control and IPF groups by Using PRM. This study provides insight into Ksucc profile alterations in IPF pathogenesis and Ksucc sites in proteins associated with mitochondrial energy metabolism can also serve as candidate molecules for future mechanism exploration and drug target selection in IPF.

The tumor microenvironment predominantly polarizes tumor-associated macrophages (TAMs) toward an M2-like phenotype, thereby inhibiting antitumor immune responses. This process is substantially affected by metabolic reprogramming; however, reeducating TAMs to enhance their antitumor capabilities through metabolic remodeling remains a challenge. Here, we show that tumor-derived microparticles loaded with succinate (SMPs) can remodel the metabolic state of TAMs. SMPs promote classical M1-like polarization of macrophages by enhancing glycolysis and attenuating the tricarboxylic acid (TCA) cycle in a protein succinylation-dependent manner. Mechanistically, succinate is delivered into the mitochondria and nucleus by SMPs, leading to succinylation of isocitrate dehydrogenase 2 (IDH2) and histone H3K122 within the lactate dehydrogenase A (Ldha) promoter region. Our findings provide a distinct approach for TAM polarization using cell membrane-derived microparticles loaded with endogenous metabolites, a platform that may be used more broadly for posttranslational modification-based tumor immunotherapy.

Tumor cells evolve strong antioxidant capacities to counteract the abnormal high level of reactive oxygen species (ROS) in the tumor microenvironment. Glutamate-cysteine ligase catalyzing subunit (GCLC) for synthesis of antioxidant glutathione (GSH) represents the key enzyme to maintain redox homeostasis of tumor cells, however, whether its activity is regulated by posttranslational modifications, such as succinylation, remains to be clarified. Here, we demonstrate the existence of succinylation modification on GCLC by in vitro and in vivo assays. NAD-dependent deacetylase Sirtuin-2 (SIRT2) serves as the desuccinylase and catalyzes GCLC desuccinylation at sites of K38, K126, and K326. Specifically, GCLC directly interacts with SIRT2, which can be substantially enhanced upon ROS treatment. This strengthened association results in GCLC desuccinylation and activation, consequently promoting GSH synthesis and rendering cancer cells resistant to ferroptosis induction. Depletion of SIRT2 decreases total GSH level and meanwhile increases the cellular susceptibility to ferroptosis, which can mostly be rescued by introducing wild-type GCLC, but not its 3K-E mutant. We further demonstrated that histone acetyltransferase P300 serves as the succinyltransferase of GCLC, and their association is remarkably decreased after ROS treatment. Thus, SIRT2-regulated GCLC succinylation represents an essential signaling axis for cancer cells to maintain their redox balance in coping with oxidative stress-induced ferroptosis.

Proteoforms are distinct molecular forms of proteins that act as building blocks of organisms, with post-translational modifications (PTMs) being one of the key changes that generate these variations. Mass spectrometry (MS)-based top-down proteomics (TDP) is the leading technology for proteoform identification due to its preservation of intact proteoforms for analysis, making it well-suited for comprehensive PTM characterization. A crucial step in TDP is searching MS data against a database of candidate proteoforms. To extend the reach of TDP to organisms with limited PTM annotations, we developed Proteoform-predictor, an open-source tool that integrates homology-based PTM site prediction into proteoform database creation. The new tool creates databases of proteoform candidates after registration of homologous sequences, transferring PTM sites from well-characterized species to those with less comprehensive proteomic data. Our tool features a user-friendly interface and intuitive workflow, making it accessible to a wide range of researchers. We demonstrate that Proteoform-predictor expands proteoform databases with tens of thousands of proteoforms for three bacterial strains by comparing them to the reference proteome of Escherichia coli (E. coli) K12. Subsequent TDP analysis for Serratia marcescens (S. marcescens) and Salmonella typhimurium (S. typhimurium) demonstrated significant improvement in protein and proteoform identification, even for proteins with variant sequences. As TDP technology advances, Proteoform-predictor will become an important tool for expanding the applicability of proteoform identification and PTM biology to more diverse species across the phylogenetic tree of life.

Vitamin C (vitC) is essential for health and shows promise in treating diseases like cancer, yet its mechanisms remain elusive. Here, we report that vitC directly modifies lysine residues to form "vitcyl-lysine"-a process termed vitcylation. Vitcylation occurs in a dose-, pH-, and sequence-dependent manner in both cell-free systems and living cells. Mechanistically, vitC vitcylates signal transducer and activator of transcription-1 (STAT1)- lysine-298 (K298), impairing its interaction with T cell protein-tyrosine phosphatase (TCPTP) and preventing STAT1-Y701 dephosphorylation. This leads to enhanced STAT1-mediated interferon (IFN) signaling in tumor cells, increased major histocompatibility complex (MHC)/human leukocyte antigen (HLA) class I expression, and activation of anti-tumor immunity in vitro and in vivo. The discovery of vitcylation as a distinctive post-translational modification provides significant insights into vitC's cellular function and therapeutic potential, opening avenues for understanding its biological effects and applications in disease treatment.

Gemcitabine combined with cisplatin is the first-line chemotherapy for advanced cholangiocarcinoma, but drug resistance remains a challenge, leading to unsatisfactory therapeutic effect. Here, we elucidate the possibility of chemotherapy regimens sensitized by inhibiting succinylation in patients with cholangiocarcinoma from the perspective of post-translational modification. Our omics analysis reveals that succinylation of PDHA1 lysine 83, a key enzyme in the tricarboxylic acid cycle, alters PDH enzyme activity, modulates metabolic flux, and leads to alpha-ketoglutaric acid accumulation in the tumor microenvironment. This process activates the OXGR1 receptor on macrophages, triggering MAPK signaling and inhibiting MHC-II antigen presentation, which promotes immune escape and tumor progression. Moreover, we show that inhibiting PDHA1 succinylation with CPI-613 enhances the efficacy of gemcitabine and cisplatin. Targeting PDHA1 succinylation may be a promising strategy to improve treatment outcomes in cholangiocarcinoma and warrants further clinical exploration.

Tertiary lymphoid structures (TLS) in the tumor microenvironment (TME) are emerging solid-tumor indicators of prognosis and response to immunotherapy. Considering that tumorigenesis requires metabolic reprogramming and subsequent TME remodeling, the discovery of TLS metabolic regulators is expected to produce immunotherapeutic targets. To identify such metabolic regulators, we constructed a metabolism-focused sgRNA library and performed an in vivo CRISPR screening in an orthotopic lung tumor mouse model. Combined with The Cancer Genome Atlas database analysis of TLS-related metabolic hub genes, we found that the loss of Acat1 in tumor cells sensitized tumors to anti-PD1 treatment, accompanied by increased TLS in the TME. Mechanistic studies revealed that ACAT1 resulted in mitochondrial protein hypersuccinylation in lung tumor cells and subsequently enhanced mitochondrial oxidative metabolism, which impeded TLS formation. Elimination of ROS by NAC or Acat1 knockdown promoted B cell aggregation and TLS construction. Consistently, data from tissue microassays of 305 patients with lung cancer showed that TLS were more abundant in non-small cell lung cancer (NSCLC) tissues with lower ACAT1 expression. Intratumoral ACAT1 expression was associated with poor immunotherapy outcomes in patients with NSCLC. In conclusion, our results identified ACAT1 as a metabolic regulator of TLS and a promising immunotherapeutic target in NSCLC.

Aging changes the protein activity status to affect the body's functions. However, how aging regulates protein posttranslational modifications (PTMs) to modulate the antiviral defense ability of the body remains unclear. Here, we found that aging promotes STAT1 β-hydroxybutyrylation (Kbhb) at Lys592, which inhibits the interaction between STAT1 and type-I interferon (IFN-I) receptor 2 (IFNAR2), thereby attenuating IFN-I-mediated antiviral defense activity. Additionally, we discovered that a small molecule from a plant source, hydroxy camptothecine, can effectively reduce the level of STAT1 Kbhb, thus increasing antiviral defense ability in vivo. Further studies revealed that STAT1 O-GlcNAc modifications at Thr699 block CBP-induced STAT1 Kbhb. Importantly, fructose can improve IFN-I antiviral defense activity by orchestrating STAT1 O-GlcNAc and Kbhb modifications. This study reveals the significance of the switch between STAT1 Kbhb and O-GlcNAc modifications in regulating IFN-I antiviral immunity during aging and provides potential strategies to improve the body's antiviral defense ability in elderly individuals.

Intracellular Ca2+ cycling governs effective myocardial systolic contraction and diastolic relaxation. SERCA2a (sarco/endoplasmic reticulum Ca2+ ATPase type 2a), which plays a crucial role in controlling intracellular Ca2+ signaling and myocardial cell function, is downregulated and inactivated during sepsis-induced heart dysfunction. However, the cause of this dysregulation remains unclear. In this study, we investigated the effect of lysine succinylation in lipopolysaccharide-induced septic heart dysfunction through global succinylome analysis of myocardial tissues from septic rats.

We conducted a succinylome profiling and developed a protein language model-based framework to prioritize succinylation at a functionally important site, and further analysis revealed crosstalk between ubiquitination and succinylation of SERCA2a. The succinylation of SERCA2a in septic rats or lipopolysaccharide-treated cells were detected by co-immunoprecipitation. Thereafter, a desuccinylated SERCA2aK352R was introduced and its function and stability were determined by Ca2+ transient and Western blot, respectively. Meanwhile, the effect on SERCA2aK352R on heart function was assessed in vivo by echocardiography and hemodynamics.

We identified 10 324 succinylated lysine sites in heart tissues, including 1042 differentially succinylated lysine sites, in response to lipopolysaccharide. SERCA2a was hypersuccinylated in the myocardial tissues of septic rats and lipopolysaccharide-treated cardiomyocytes. Increased ubiquitination level, reduced protein level, and activity of SERCA2a were observed, along with increased succinylation of SERCA2a in vivo and in vitro. K352 was essential for SERCA2a succinylation, which reduced SERCA2a protein level by promoting formation of the K48 ubiquitin chain on SERCA2a and its degradation by proteasomes. Co-immunoprecipitation combined with liquid chromatography-tandem mass spectrometry identified that SIRT2 (sirtuin2), a deacylase, exhibited interaction with SERCA2a. Furthermore, SIRT2 decreased K352 succinylation of SERCA2a, suggesting that SIRT2 may function as a desuccinylase for SERCA2a.

Succinylation of SERCA2a at K352, which was controlled by SIRT2, promotes its ubiquitinoylation and degradation by proteasomes in sepsis-induced heart dysfunction.

Cellular stress response is a pivotal process in tumor development and therapy. Activating transcription factor 3 (ATF3), a representative stress-responsive protein, plays pleiotropic roles in various biological processes. Over the past decade, studies have described not only the general role of ATF3 in tumor metabolism but also the complexity of ATF3 expression regulation and its associated modifications, including phosphorylation, ubiquitination, SUMOylation, and NEDDylation. Interestingly, beyond being a transcription factor, ATF3 can act as a modifier to control the ubiquitination of target molecules, such as p53, to exert its function in tumors. These advances in uncovering ATF3 biological function have yielded new insights into the cellular stress response during tumor development and will be instrumental in developing novel interventions. In this review, we update the role of ATF3 as a nexus in amino acid metabolism, lipid metabolism, glycometabolism, and other metabolic pathways in tumors; delineate the underlying mechanisms involving DNA level regulation, epigenetic regulation, and post-translational modifications of ATF3; and summarize the progression of tumor mono/combination therapies related to ATF3. In particular, we discuss the challenges that need to be addressed to provide a new conceptual framework for further understanding the potential therapeutic value of ATF3 in ongoing clinical trials.

Non-small cell lung cancer (NSCLC), the most common type of lung cancer, remains a leading cause of cancer-related mortality worldwide. Immune checkpoint inhibitors (ICIs) have emerged as a promising therapy for NSCLC but only benefit a subset of patients. In this issue of the JCI, Jiao et al. revealed that acetyl-CoA acetyltransferase 1 (ACAT1) limited the efficacy of ICIs in NSCLC by impeding tertiary lymphoid structures (TLS) in the tumor microenvironment (TME). Targeting ACAT1 in tumor cells reduced mitochondrial hypersuccinylation and oxidative stress, enhancing TLS abundance and improving the efficacy of ICIs in preclinical murine models of NSCLC.

The nonenergy - producing functions of metabolism are attracting increasing attention, as metabolic changes are involved in discrete pathways modulating enzyme activity and gene expression. Substantial evidence suggests that myocardial metabolic remodeling occurring during diabetic cardiomyopathy, heart failure, and cardiac pathological stress (e.g., myocardial ischemia, pressure overload) contributes to the progression of pathology. Within the rewired metabolic network, metabolic intermediates and end-products can directly alter protein function and/or regulate epigenetic modifications by providing acyl groups for posttranslational modifications, thereby affecting the overall cardiac stress response and providing a direct link between cellular metabolism and cardiac pathology. This review provides a comprehensive overview of the functional diversity and mechanistic roles of several types of metabolite-mediated histone and nonhistone acylation, namely O-GlcNAcylation, lactylation, crotonylation, β-hydroxybutyrylation, and succinylation, as well as fatty acid-mediated modifications, in regulating physiological processes and contributing to the progression of heart disease. Furthermore, it explores the potential of these modifications as therapeutic targets for disease intervention.

SIRT7, one of the least studied members of the Sirtuins family, is an NAD+-dependent lysine deacetylase and desuccinylase. While previous studies using affinity enrichment and quantitative proteomics identified numerous lysine-deacetylated substrates of SIRT7, its lysine-desuccinylated substrates remain underexplored, limiting our understanding of its role in cellular homeostasis. Here, we demonstrated that SIRT7 is predominantly expressed in immune tissues, especially in adaptive immune cells, including T cells. Through proteomics, lysine succinylome, and acetylome analysis of spleen from wild-type (WT) and Sirt7-/- mice, we identified significant succinylation of proteins involved in the branched-chain amino acid (BCAA) catabolism pathway in Sirt7-/- mice. We further found that SIRT7 partially localizes to mitochondria, interacting with key enzymes of the BCAA catabolism pathway and promoting their desuccinylation. Sirt7 deficiency leads to enhanced BCAA catabolism, accumulation of acyl-CoA, and increased fatty acid (FA) synthesis. As T cells rely heavily on amino acid metabolism for activation, differentiation, and function, we investigated the impact of SIRT7 using a T cell-specific Sirt7 knockout mouse model (Sirt7fl/flCd4-Cre). Our results show that SIRT7 is crucial for T cell proliferation, activation, and antitumor function. Sirt7 deficiency in T cells results in the accumulation of BCAA metabolites and FAs, reduced cytotoxic cytokines secretion such as IFN-γ, and T cell exhaustion. Reducing BCAA levels with BT2, a BCKDK inhibitor, or BCAA-free treatment alleviated these effects, while FA treatment exacerbates them. Overall, our findings identify SIRT7 as a critical regulator linking BCAA and FA metabolism to T cell antitumor immunity, providing new insights into its potential as a therapeutic target.

Background Posttranslational modifications of histone lysine (K) have integral connections with cell metabolism, and participate in the carcinogenesis of various cancers. This study focuses on evaluating the expression of histone H4 lys 5 lactylation (H4K5lac) and its clinical role in breast cancer (BC). Methods During this research, immunohistochemistry (IHC) and immunoblotting, utilizing a specific primary anti-L-lactyl-histone H4 (Lys 5) rabbit monoclonal antibody, were employed to assess H4K5lac expression in BC tissue chips. H4K5lac expression in the peripheral blood mononuclear cells (PBMCs) of BC patients was investigated through immunoblotting. Results IHC revealed upregulation of histone H4K5lac in both triple-negative breast cancer (TNBC) and non-TNBC tissues, with positive rate of 91.40% [170/(150 + 19 + 17)] and 93.64% (103/110) in TNBC and non-TNBC tissues, respectively. The expression of H4K5lac demonstrated positive correlations with lymph nodes (%), and Ki-67 expression. Survival analysis indicated a negative correlation between H4K5lac expression and overall survival (OS) time in both TNBC (HR [hazard ratio] = 2.773, 95%CI [confidence interval]: 1.128-6.851, P = 0.0384) and non-TNBC cases (HR = 2.156, 95%CI: 1.011-4.599, P = 0.0275). Furthermore, elevated levels of H4K5lac were observed in the PBMCs of BC cases, and H4K5lac expression is positively correlated with serum lactate and carcinoma embryonic antigen (CEA) levels. Conclusions Histone H4K5lac exhibits increased levels in both BC tissues and PBMCs, suggesting its potential as a promising biomarker for BC. This study might pave the way toward novel lactylation treatment strategies in BC.

Papillary thyroid cancer (PTC) is one of the malignant tumors with rapidly increasing morbidity and mortality. Sirtuin 7 (SIRT7) is a desuccinylase that is involved in tumorigenesis. The activation of large tumor suppressor 1 (LATS1) can effectively suppress tumorigenesis in multiple tumors and can be affected by SIRT7. This study aimed to explore the role and mechanism of SIRT7 in PTC progression.

The RNA and protein levels were detected by quantitative real-time PCR (qPCR) and western blot, respectively. Cell proliferation was measured by cell counting kit-8 and colony formation. The apoptosis of PTC cells was analyzed by flow cytometry and Live/dead cell staining. The interaction between proteins was detected by co-immunoprecipitation.

The results showed that SIRT7 was highly expressed in PTC tissues and cells. Functional studies showed that knockdown of SIRT7 inhibited the proliferation and induced apoptosis of PTC cells. Mechanistically, SIRT7 could directly interact with LATS1 and reduce the stability of the LATS1 protein. Later, rescue experiments suggested that LATS1 silencing reversed the effect of SIRT7 knockdown on PTC cell growth and apoptosis. In addition, SIRT7 promoted tumor growth in vivo.

Taken together, silencing of SIRT7 promotes the succinylation of LATS1 to enhance LATS1 stability, thus inhibiting the progression of PTC. Therefore, SIRT7 and LATS1 may become novel and potential therapeutic targets for PTC.

Atrial fibrillation (AF) is the most common form of arrhythmia and is a growing clinical problem. Post-translational modifications (PTMs) constitute crucial epigenetic mechanisms but modification of lysine 2-hydroxyisobutyrylation (Khib) in AF is still unknown. This study aimed to investigate the role and mechanism of Khib in AF.

PTM proteomics was applied in the human atrial tissue from AF and sinus rhythm patients with heart valve disease during cardiac surgery to identify the Khib sites. The functional changes of differential modification sites were further validated at the cellular level. Cellular electrophysiology was performed to record the ion channel current and action potential duration (APD).

The modification of 124 Khib sites in 35 proteins and 67 sites in 48 proteins exhibited significant increase or decrease in AF compared to sinus rhythm. Ten Khib sites were included in energy metabolism-related signaling pathways (HXK1, TPIS, PGM1, and ODPX in glycolysis; MDHC and IDH3A in tricarboxylic acid cycle; NDUS2, ETFB, ADT3, and ATPB in oxidative respiratory chain). Importantly, decreased HXK1 K418hib regulated by HDAC2 attenuated the original chemical binding domain between HXK1 and glucose, inhibited the binding ability between HXK1 and glucose, and reduced catalytic ability of the enzyme, resulting in low production of glucose-6-phosphate and ATP. Further, it also increased Kir6.2 protein and the current of KATP channel, and decreased APD.

This study demonstrates the importance of Khib to catalysis of HXK1 and reveals molecular mechanisms of HXK1 K418hib in AF, providing new insight into strategies of AF.

Tumors undergo metabolic reprogramming to meet the energetic, synthetic and redox demands essential for malignancy, often characterized by increased glycolysis and lactate production. However, the role of mitochondrial metabolism in tumor immunity remains unclear. The present study integrates spatial transcriptomics, bulk transcriptomics and proteomics, revealing a strong link between the metabolite succinyl-CoA and tumor immunity as well as the efficacy of anti-programmed cell death protein-1 (PD-1) therapy in patients with melanoma. Elevated succinyl-CoA levels, through α-ketoglutarate or succinate supplementation, enhanced T cell-mediated tumor elimination, both in vitro and in vivo. Mechanistically, succinylation of the ligand of PD-1 (PD-L1) at lysine 129 led to its degradation. Increased carnitine palmitoyltransferase 1A (CPT1A), identified as a succinyltransferase for PD-L1, boosted anti-tumor activity. Preclinically, bezafibrate, a hyperlipidemia drug, upregulated CPT1A and synergized with CTLA-4 monoclonal antibody to inhibit tumor growth. Clinically, higher PD-L1 and lower CPT1A levels in tumors correlated with better anti-PD-1 therapy responses, suggesting potential biomarkers for prediction of treatment efficacy.

Succinate upregulates enterohaemorrhagic Escherichia coli (EHEC) virulence. Lysine succinylation, a post-translational modification, regulates cellular function in eukaryotes but is less characterized in bacteria. We hypothesized that lysine succinylation regulates EHEC virulence. Here we used SILAC-based proteomics and characterized the EHEC succinylome to show that the transcription factor, PurR, is succinylated at K24 and K55. Succinylation of PurR inhibited its ability to directly bind DNA and repress expression of a major virulence factor, the Type 3 Secretion System (T3SS), thus increasing T3SS expression. Deletion of purR, or K24E or K55E mutation, increased EHEC adherence to cells and colonization of infant rabbits. Using mice treated with streptomycin to deplete succinate, or colonized with succinate-producing Prevotella copri to increase succinate levels, we showed that microbiota-derived succinate increased succinylation of PurR to promote virulence of Citrobacter rodentium, a model for EHEC, in mice. Lastly, we identified CitC as the succinyltransferase required for PurR modification.

Acute pancreatitis (AP) is a severe inflammatory disorder associated with metabolic reprogramming and mitochondrial dysfunction. This study investigated central carbon metabolism alterations in pancreatic acinar cells during AP, elucidated the molecular mechanisms of tricarboxylic acid (TCA) cycle disorders, and explored the role of protein hypersuccinylation in AP pathogenesis. Using in vitro and in vivo AP models, targeted metabolomics and bioinformatics analyses revealed TCA cycle dysregulation characterized by elevated succinyl-CoA and decreased succinate levels. Colorimetric assays, mass spectrometry, and site-directed mutagenesis demonstrated that SIRT5 downregulation led to SUCLA2 hypersuccinylation at K118, inhibiting succinyl-CoA synthetase activity and triggering a vicious cycle of succinyl-CoA accumulation and SUCLA2 succinylation. Adenovirus-mediated SIRT5 overexpression and SUCLA2 knockdown clarified the SIRT5-SUCLA2 pathway's role in regulating TCA cycle disorders. Protein succinylation levels positively correlated with pancreatic tissue damage and mitochondrial injury severity. Succinylome analysis identified cytochrome c1 (CYC1) as a key hypersuccinylated protein, and the SIRT5-SUCLA2 pathway regulated its succinylation level and electron transport chain complex III activity. Hypersuccinylation induced mitochondrial DNA release, activating the cGAS-STING pathway, contributing to multiple organ dysfunction syndrome. Modulating the SIRT5-SUCLA2 axis attenuated TCA cycle dysregulation, protein hypersuccinylation, mitochondrial damage, and inflammatory responses in AP. These findings reveal novel mechanisms linking the SIRT5-SUCLA2 axis, TCA cycle dysfunction, and protein hypersuccinylation in AP pathogenesis, providing potential therapeutic targets for AP treatment.

Protein succinylation, a post-translational modification wherein a succinyl group (-CO-CH₂-CH₂-CO-) attaches to lysine residues, plays a critical regulatory role in cellular processes. Dysregulated succinylation has been implicated in the onset and progression of various diseases, including liver, cardiac, pulmonary, and neurological disorders. However, identifying succinylation sites through experimental methods is often labor-intensive, costly, and technically challenging. To address this, we introduce an approach called CbiLSuccSite, that integrates Convolutional Neural Networks (CNN) with Bidirectional Long Short-Term Memory (Bi-LSTM) networks for the accurate prediction of protein succinylation sites. Our approach employs a word embedding layer to encode protein sequences, enabling the automatic learning of intricate patterns and dependencies without manual feature extraction. In 10-fold cross-validation, CBiLSuccSite achieved superior predictive performance, with an Area Under the Curve (AUC) of 0.826 and a Matthews Correlation Coefficient (MCC) of 0.502. Independent testing further validated its robustness, yielding an AUC of 0.818 and an MCC of 0.53. The integration of CNN and Bi-LSTM leverages the strengths of both architectures, establishing CBiLSuccSite as an effective tool for protein language processing and succinylation site prediction. Our model and code are publicly accessible at: https://github.com/nuinvtnu/CBiLSuccSite.

Histone lysine succinylation, an emerging epigenetic marker, has been implicated in diverse cellular functions, yet its role in cancer drug resistance is not well understood. Here we investigated the genome-wide alterations in histone 3 lysine 23 succinylation (H3K23su) and its impact on gene expression in 5-fluorouracil (5-FU)-resistant HCT15 colon cancer cells. We utilized CUT&Tag assays to identify differentially enriched regions (DERs) of H3K23su in 5-FU-resistant HCT15 cells via integration with ATAC-seq and RNA sequencing data. The regulatory network involving transcription factors (TFs), notably FOSL2 and KLF6, and their downstream target genes was dissected using motif enrichment analysis and chromatin immunoprecipitation assays. Our results revealed a strong positive correlation between H3K23su DERs, differentially expressed genes (DEGs) and H3K27ac, indicating that H3K23su enrichment is closely related to gene activation. The DEGs associated with the H3K23su GAIN regions were significantly enriched in pathways related to colorectal cancer, including the Wnt, MAPK and p53 signaling pathways. FOSL2 and KLF6 emerged as pivotal TFs potentially modulating DEGs associated with H3K23su DERs and were found to be essential for sustaining 5-FU resistance. Notably, we discovered that FOSL2 and KLF6 recruit the PCAF-p300/CBP complex to synergistically regulate SEMA3C expression, which subsequently modulates the canonical Wnt-β-catenin signaling pathway, leading to the upregulation of MYC and FOSL2. This study demonstrated that H3K23su is a critical epigenetic determinant of 5-FU resistance in colon cancer cells, exerting its effects through the modulation of critical genes and TFs. These findings indicate that interventions aimed at targeting TFs or enzymes involved in H3K23su modification could represent potential therapeutic strategies for treating colorectal cancers that are resistant to 5-FU treatment.

Ageing-induced skeletal muscle deterioration contributes to sarcopenia and frailty, adversely impacting the quality of life in the elderly. However, the molecular mechanisms behind primate skeletal muscle ageing remain largely unexplored. Here, we show that SIRT5 expression is reduced in aged primate skeletal muscles from both genders. SIRT5 deficiency in human myotubes hastens cellular senescence and intensifies inflammation. Mechanistically, we demonstrate that TBK1 is a natural substrate for SIRT5. SIRT5 desuccinylates TBK1 at lysine 137, which leads to TBK1 dephosphorylation and the suppression of the downstream inflammatory pathway. Using SIRT5 lentiviral vectors for skeletal muscle gene therapy in male mice enhances physical performance and alleviates age-related muscle dysfunction. This study sheds light on the molecular underpinnings of skeletal muscle ageing and presents the SIRT5-TBK1 pathway as a promising target for combating age-related skeletal muscle degeneration.

Hepatocellular carcinoma (HCC) is one of the leading causes of cancer-related death worldwide, and its onset and progression are closely associated with epigenetic modifications, particularly post-translational modifications of histones (HPTMs). In recent years, advances in mass spectrometry (MS) have revealed a series of novel HPTMs, including succinylation (Ksuc), citrullination (Kcit), butyrylation (Kbhb), lactylation (Kla), crotonylation (Kcr), and 2-hydroxyisobutyrylation (Khib). These modifications not only expand the histone code but also play significant roles in key carcinogenic processes such as tumor proliferation, metastasis, and metabolic reprogramming in HCC. This review provides the first comprehensive analysis of the impact of novel HPTMs on gene expression, cellular metabolism, immune evasion, and the tumor microenvironment. It specifically focuses on their roles in promoting tumor stem cell characteristics, epithelial-mesenchymal transition (EMT), and therapeutic resistance. Additionally, the review highlights the dynamic regulation of these modifications by specific enzymes, including "writers," "readers," and "erasers."

The innate immune system relies on a network of signaling proteins classified by shared domains, which serve as functional units that orchestrate inflammatory and host defensive activities. Within type I interferon (IFN) responses, the stimulator of interferon genes protein (STING), mitochondrial antiviral-signaling protein (MAVS), Toll-IL-1 receptor-resistance protein domain-containing adapter-inducing interferon-β (TRIF), Toll-like receptor adapter interacting with SLC15A4 on the lysosome (TASL), insulin receptor tyrosine kinase substrate protein of 53 kDa (IRSp53), and GEM interacting protein (GMIP) utilize a conserved pLxIS motif to recruit IRF family transcription factors. Notably, the pLxIS motif functions within a larger signaling unit, which is referred to here as an Activator of Interferon Expression via a pLxIS motif (ARIES) domain. ARIES domains consist of the pLxIS motif and adjacent kinase activation motifs that together drive IFN responses. This review explores how ARIES domains promote immune responses via shared and distinct signaling mechanisms, protein localization, and regulation of metabolic shifts, underscoring their evolutionary conservation and critical role in host defense.

Metamorphosis is an important way for insects to adapt to the environment. In this process, larval tissue destruction regulated by 20-hydroxyecdysone (20E) and adult tissue reconstruction regulated by insulin-like peptides (ILPs) occur simultaneously, but the detailed mechanism is still unclear. Here, the results of succinylome, subcellular localization, and protein interaction analysis show that non-succinylated insulin-degrading enzyme (IDE) localizes in the cytoplasm, binds to insulin-like growth factor 2 (IGF-2-like), and degrades it. When the metamorphosis is initiated, 20E up-regulated carnitine palmitoyltransferase 1A (Cpt1a) through transcription factor Krüppel-like factor 15 (KLF15), thus increasing the level of IDE succinylation on K179. Succinylated IDE translocated from cytoplasm to nucleus, combined with ecdysone receptor to promote 20E signaling pathway, causing larval tissue destruction, while IGF-2-like was released to promote adult tissue proliferation. That is, succinylation alters subcellular localization of IDE so that it can bind to different target proteins and act as a hub of metamorphosis.

Salicylic acid, as a plant hormone, significantly affects the physiological and biochemical indexes of soluble sugar, malondialdehyde content, peroxidase, and superoxide dismutase enzyme activity in Platycodon grandiflorus. Lysine malonylation is a post-translational modification that involves various cellular functions in plants, though it is rarely studied, especially in medicinal plants. In this study, the aim was to perform a comparative quantitative proteomic study of malonylation modification on P. grandiflorus root proteins after salicylic acid treatment using Western blot with specific antibodies, affinity enrichment and LC-MS/MS analysis methods. The analysis identified 1907 malonyl sites for 809 proteins, with 414 proteins and 798 modification sites quantified with high confidence. Post-treatment, 361 proteins were upregulated, and 310 were downregulated. Bioinformatics analysis revealed that malonylation in P. grandiflorus is primarily involved in photosynthesis and carbon metabolism. Physiological and biochemical analysis showed that salicylic acid treatment increased the malondialdehyde levels, soluble protein, superoxide dismutase, and peroxidase activity but did not significantly affect the total saponins content in P. grandiflorus. These findings provide an important basis for exploring the molecular mechanisms of P. grandiflorus following salicylic acid treatment and enhance understanding of the biological function of protein lysine malonylation in plants.