Understanding the Function of Mammalian Sirtuins and Protein Lysine Acylation.

Protein lysine acetylation is an important posttranslational modification that regulates numerous biological processes. Targeting lysine acetylation regulatory factors, such as acetyltransferases, deacetylases, and acetyl-lysine recognition domains, has been shown to have potential for treating human diseases, including cancer and neurological diseases. Over the past decade, many other acyl-lysine modifications, such as succinylation, crotonylation, and long-chain fatty acylation, have also been investigated and shown to have interesting biological functions. Here, we provide an overview of the functions of different acyl-lysine modifications in mammals. We focus on lysine acetylation as it is well characterized, and principles learned from acetylation are useful for understanding the functions of other lysine acylations. We pay special attention to the sirtuins, given that the study of sirtuins has provided a great deal of information about the functions of lysine acylation. We emphasize the regulation of sirtuins to illustrate that their regulation enables cells to respond to various signals and stresses.

OXCT1 functions as a succinyltransferase, contributing to hepatocellular carcinoma via succinylating LACTB.

Metabolic reprogramming is an important feature of cancers that has been closely linked to post-translational protein modification (PTM). Lysine succinylation is a recently identified PTM involved in regulating protein functions, whereas its regulatory mechanism and possible roles in tumor progression remain unclear. Here, we show that OXCT1, an enzyme catalyzing ketone body oxidation, functions as a lysine succinyltransferase to contribute to tumor progression. Mechanistically, we find that OXCT1 functions as a succinyltransferase, with residue G424 essential for this activity. We also identified serine beta-lactamase-like protein (LACTB) as a main target of OXCT1-mediated succinylation. Extensive succinylation of LACTB K284 inhibits its proteolytic activity, resulting in increased mitochondrial membrane potential and respiration, ultimately leading to hepatocellular carcinoma (HCC) progression. In summary, this study establishes lysine succinyltransferase function of OXCT1 and highlights a link between HCC prognosis and LACTB K284 succinylation, suggesting a potentially valuable biomarker and therapeutic target for further development.

The Substrate Specificity of Sirtuins.

Sirtuins are NAD(+)-dependent enzymes universally present in all organisms, where they play central roles in regulating numerous biological processes. Although early studies showed that sirtuins deacetylated lysines in a reaction that consumes NAD(+), more recent studies have revealed that these enzymes can remove a variety of acyl-lysine modifications. The specificities for varied acyl modifications may thus underlie the distinct roles of the different sirtuins within a given organism. This review summarizes the structure, chemistry, and substrate specificity of sirtuins with a focus on how different sirtuins recognize distinct substrates and thus carry out specific functions.

SUCLA2-coupled regulation of GLS succinylation and activity counteracts oxidative stress in tumor cells.

Glutaminase regulates glutaminolysis to promote cancer cell proliferation. However, the mechanism underlying glutaminase activity regulation is largely unknown. Here, we demonstrate that kidney-type glutaminase (GLS) is highly expressed in human pancreatic ductal adenocarcinoma (PDAC) specimens with correspondingly upregulated glutamine dependence for PDAC cell proliferation. Upon oxidative stress, the succinyl-coenzyme A (CoA) synthetase ADP-forming subunit ß (SUCLA2) phosphorylated by p38 mitogen-activated protein kinase (MAPK) at S79 dissociates from GLS, resulting in enhanced GLS K311 succinylation, oligomerization, and activity. Activated GLS increases glutaminolysis and the production of nicotinamide adenine dinucleotide phosphate (NADPH) and glutathione, thereby counteracting oxidative stress and promoting tumor cell survival and tumor growth in mice. In addition, the levels of SUCLA2 pS79 and GLS K311 succinylation, which were mutually correlated, were positively associated with advanced stages of PDAC and poor prognosis for patients. Our findings reveal critical regulation of GLS by SUCLA2-coupled GLS succinylation regulation and underscore the regulatory role of metabolites in glutaminolysis and PDAC development.

SUCLG1 restricts POLRMT succinylation to enhance mitochondrial biogenesis and leukemia progression.

Mitochondria are cellular powerhouses that generate energy through the electron transport chain (ETC). The mitochondrial genome (mtDNA) encodes essential ETC proteins in a compartmentalized manner, however, the mechanism underlying metabolic regulation of mtDNA function remains unknown. Here, we report that expression of tricarboxylic acid cycle enzyme succinate-CoA ligase SUCLG1 strongly correlates with ETC genes across various TCGA cancer transcriptomes. Mechanistically, SUCLG1 restricts succinyl-CoA levels to suppress the succinylation of mitochondrial RNA polymerase (POLRMT). Lysine 622 succinylation disrupts the interaction of POLRMT with mtDNA and mitochondrial transcription factors. SUCLG1-mediated POLRMT hyposuccinylation maintains mtDNA transcription, mitochondrial biogenesis, and leukemia cell proliferation. Specifically, leukemia-promoting FMS-like tyrosine kinase 3 (FLT3) mutations modulate nuclear transcription and upregulate SUCLG1 expression to reduce succinyl-CoA and POLRMT succinylation, resulting in enhanced mitobiogenesis. In line, genetic depletion of POLRMT or SUCLG1 significantly delays disease progression in mouse and humanized leukemia models. Importantly, succinyl-CoA level and POLRMT succinylation are downregulated in FLT3-mutated clinical leukemia samples, linking enhanced mitobiogenesis to cancer progression. Together, SUCLG1 connects succinyl-CoA with POLRMT succinylation to modulate mitochondrial function and cancer development.

Regulation of UCP1 and Mitochondrial Metabolism in Brown Adipose Tissue by Reversible Succinylation.

Brown adipose tissue (BAT) is rich in mitochondria and plays important roles in energy expenditure, thermogenesis, and glucose homeostasis. We find that levels of mitochondrial protein succinylation and malonylation are high in BAT and subject to physiological and genetic regulation. BAT-specific deletion of Sirt5, a mitochondrial desuccinylase and demalonylase, results in dramatic increases in global protein succinylation and malonylation. Mass spectrometry-based quantification of succinylation reveals that Sirt5 regulates the key thermogenic protein in BAT, UCP1. Mutation of the two succinylated lysines in UCP1 to acyl-mimetic glutamine and glutamic acid significantly decreases its stability and activity. The reduced function of UCP1 and other proteins in Sirt5KO BAT results in impaired mitochondria respiration, defective mitophagy, and metabolic inflexibility. Thus, succinylation of UCP1 and other mitochondrial proteins plays an important role in BAT and in regulation of energy homeostasis.

Conjoint analysis of succinylome and phosphorylome reveals imbalanced HDAC phosphorylation-driven succinylayion dynamic contibutes to lung cancer.

Cancerous genetic mutations result in a complex and comprehensive post-translational modification (PTM) dynamics, in which protein succinylation is well known for its ability to reprogram cell metabolism and is involved in the malignant evolution. Little is known about the regulatory interactions between succinylation and other PTMs in the PTM network. Here, we developed a conjoint analysis and systematic clustering method to explore the intermodification communications between succinylome and phosphorylome from eight lung cancer patients. We found that the intermodification coorperation in both parallel and series. Besides directly participating in metabolism pathways, some phosphosites out of mitochondria were identified as an upstream regulatory modification directing succinylome dynamics in cancer metabolism reprogramming. Phosphorylated activation of histone deacetylase (HDAC) in lung cancer resulted in the removal of acetylation and favored the occurrence of succinylation modification of mitochondrial proteins. These results suggest a tandem regulation between succinylation and phosphorylation in the PTM network and provide HDAC-related targets for intervening mitochondrial succinylation and cancer metabolism reprogramming.

Succinylated lysine residue prediction revisited.

Lysine succinylation is a kind of post-translational modification (PTM) that plays a crucial role in regulating the cellular processes. Aberrant succinylation may cause inflammation, cancers, metabolism diseases and nervous system diseases. The experimental methods to detect succinylation sites are time-consuming and costly. This thus calls for computational models with high efficacy, and attention has been given in the literature to develop such models, albeit with only moderate success in the context of different evaluation metrics. One crucial aspect in this context is the biochemical and physicochemical properties of amino acids, which appear to be useful as features for such computational predictors. However, some of the existing computational models did not use the biochemical and physicochemical properties of amino acids. In contrast, some others used them without considering the inter-dependency among the properties. The combinations of biochemical and physicochemical properties derived through our optimization process achieve better results than the results achieved by combining all the properties. We propose three deep learning architectures: CNN+Bi-LSTM (CBL), Bi-LSTM+CNN (BLC) and their combination (CBL_BLC). We find that CBL_BLC outperforms the other two. Ensembling of different models successfully improves the results. Notably, tuning the threshold of the ensemble classifiers further improves the results. Upon comparing our work with other existing works on two datasets, we successfully achieve better sensitivity and specificity by varying the threshold value.

RMTLysPTM: recognizing multiple types of lysine PTM sites by deep analysis on sequences.

Post-translational modification (PTM) occurs after a protein is translated from ribonucleic acid. It is an important living creature life phenomenon because it is implicated in almost all cellular processes. Identification of PTM sites from a given protein sequence is a hot topic in bioinformatics. Lots of computational methods have been proposed, and they provide good performance. However, most previous methods can only tackle one PTM type. Few methods consider multiple PTM types. In this study, a multi-label classification model, named RMTLysPTM, was developed to recognize four types of lysine (K) PTM sites, including acetylation, crotonylation, methylation and succinylation. The surrounding sites of a lysine site were selected to constitute a peptide segment, representing the lysine at the center. Deep analysis was conducted to count the distribution of 2-residues with fixed location across the four types of lysine PTM sites. By aggregating the distribution information of 2-residues in one peptide segment, the peptide segment was encoded by informative features. Furthermore, a prediction engine that can precisely capture the traits of the above representations was designed to recognize the types of lysine PTM sites. The cross-validation results on two datasets (Qiu and CPLM training datasets) suggested that the model had extremely high performance and RMTLysPTM had strong generalization ability by testing it on protein Q16778 and CPLM testing datasets. The model was found to be generally superior to all previous models and those using popular methods and features. A web server was set up for RMTLysPTM, and it can be accessed at http://119.3.127.138/.

12,13-diHOME attenuates high glucose-induced calcification of vascular smooth muscle cells through repressing CPT1A-mediated HMGB1 succinylation.

Diabetes is closely associated with vascular calcification (VC). Exorbitant glucose concentration activates pro-calcific effects in vascular smooth muscle cells (VSMCs). This study enrolled 159 elderly patients with type 2 diabetes and divided them into three groups, T1, T2 and T3, according to brachial-ankle pulse wave velocity(BaPWV). There were statistically significant differences in the waist circumference, waist hip ratio, systolic blood pressure, 12,13-diHOME (a lipokin) concentration among T1, T2 and T3. 12,13-diHOME levels were positively correlated to high density lipoprotein cholesterol and total cholesterol, but negatively correlated to with waist circumference, waist hip ratio, systolic blood pressure and baPWV. Studies in vitro showed that 12,13-diHOME effectively inhibits calcification in VSMCs under high glucose conditions. Notably, 12,13-diHOME suppressed the up-regulation of carnitine O-palmitoyltransferase 1 (CPT1A) and CPT1A-induced succinylation of HMGB1. The succinylation of HMGB1 at the K90 promoted the protein stability and induced the enrichment of HMGB1 in cytoplasm, which induced the calcification in VSMCs. Together, 12,13-diHOME attenuates high glucose-induced calcification in VSMCs through repressing CPT1A-mediated HMGB1 succinylation.

Lysine Succinylation of VBS Contributes to Sclerotia Development and Aflatoxin Biosynthesis in Aspergillus flavus.

Aspergillus flavus is a common saprophytic and pathogenic fungus, and its secondary metabolic pathways are one of the most highly characterized owing to its aflatoxin (AF) metabolite affecting global economic crops and human health. Different natural environments can cause significant variations in AF synthesis. Succinylation was recently identified as one of the most critical regulatory post-translational modifications affecting metabolic pathways. It is primarily reported in human cells and bacteria with few studies on fungi. Proteomic quantification of lysine succinylation (Ksuc) exploring its potential involvement in secondary metabolism regulation (including AF production) has not been performed under natural conditions in A. flavus. In this study, a quantification method was performed based on tandem mass tag labeling and antibody-based affinity enrichment of succinylated peptides via high accuracy nano-liquid chromatography with tandem mass spectrometry to explore the succinylation mechanism affecting the pathogenicity of naturally isolated A. flavus strains with varying toxin production. Altogether, 1240 Ksuc sites in 768 proteins were identified with 1103 sites in 685 proteins quantified. Comparing succinylated protein levels between high and low AF-producing A. flavus strains, bioinformatics analysis indicated that most succinylated proteins located in the AF biosynthetic pathway were downregulated, which directly affected AF synthesis. Versicolorin B synthase is a key catalytic enzyme for heterochrome B synthesis during AF synthesis. Site-directed mutagenesis and biochemical studies revealed that versicolorin B synthase succinylation is an important regulatory mechanism affecting sclerotia development and AF biosynthesis in A. flavus. In summary, our quantitative study of the lysine succinylome in high/low AF-producing strains revealed the role of Ksuc in regulating AF biosynthesis. We revealed novel insights into the metabolism of AF biosynthesis using naturally isolated A. flavus strains and identified a rich source of metabolism-related enzymes regulated by succinylation.

Recent Contributions of Proteomics to Our Understanding of Reversible Nε-Lysine Acylation in Bacteria.

Post-translational modifications (PTMs) have been extensively studied in both eukaryotes and prokaryotes. Lysine acetylation, originally thought to be a rare occurrence in bacteria, is now recognized as a prevalent and important PTM in more than 50 species. This expansion in interest in bacterial PTMs became possible with the advancement of mass spectrometry technology and improved reagents such as acyl-modification specific antibodies. In this Review, we discuss how mass spectrometry-based proteomic studies of lysine acetylation and other acyl modifications have contributed to our understanding of bacterial physiology, focusing on recently published studies from 2018 to 2023. We begin with a discussion of approaches used to study bacterial PTMs. Next, we discuss newly characterized acylomes, including acetylomes, succinylomes, and malonylomes, in different bacterial species. In addition, we examine proteomic contributions to our understanding of bacterial virulence and biofilm formation. Finally, we discuss the contributions of mass spectrometry to our understanding of the mechanisms of acetylation, both enzymatic and nonenzymatic. We end with a discussion of the current state of the field and possible future research avenues to explore.

Vibrio alginolyticus PEPCK Mediates Florfenicol Resistance through Lysine Succinylation Modification.

Protein succinylation modification is a common post-translational modification (PTM) that plays an important role in bacterial metabolic regulation. In this study, quantitative analysis was conducted on the succinylated proteome of wild-type and florfenicol-resistant Vibrio alginolyticus to investigate the mechanism of succinylation regulating antibiotic resistance. Bioinformatic analysis showed that the differentially succinylated proteins were mainly enriched in energy metabolism, and it was found that the succinylation level of phosphoenolpyruvate carboxyl kinase (PEPCK) was highly expressed in the florfenicol-resistant strain. Site-directed mutagenesis was used to mutate the lysine (K) at the succinylation site of PEPCK to glutamic acid (E) and arginine (R), respectively, to investigate the function of lysine succinylation of PEPCK in the florfenicol resistance of V. alginolyticus. The detection of site-directed mutagenesis strain viability under florfenicol revealed that the survival rate of the E mutant was significantly higher than that of the R mutant and wild type, indicating that succinylation modification of PEPCK protein may affect the resistance of V. alginolyticus to florfenicol. This study indicates the important role of PEPCK during V. alginolyticus antibiotic-resistance evolution and provides a theoretical basis for the prevention and control of vibriosis and the development of new antibiotics.

Role of novel protein acylation modifications in immunity and its related diseases.

The cross-regulation of immunity and metabolism is currently a research hotspot in life sciences and immunology. Metabolic immunology plays an important role in cutting-edge fields such as metabolic regulatory mechanisms in immune cell development and function, and metabolic targets and immune-related disease pathways. Protein post-translational modification (PTM) is a key epigenetic mechanism that regulates various biological processes and highlights metabolite functions. Currently, more than 400 PTM types have been identified to affect the functions of several proteins. Among these, metabolic PTMs, particularly various newly identified histone or non-histone acylation modifications, can effectively regulate various functions, processes and diseases of the immune system, as well as immune-related diseases. Thus, drugs aimed at targeted acylation modification can have substantial therapeutic potential in regulating immunity, indicating a new direction for further clinical translational research. This review summarises the characteristics and functions of seven novel lysine acylation modifications, including succinylation, S-palmitoylation, lactylation, crotonylation, 2-hydroxyisobutyrylation, ß-hydroxybutyrylation and malonylation, and their association with immunity, thereby providing valuable references for the diagnosis and treatment of immune disorders associated with new acylation modifications.

Succinylation of Park7 activates a protective metabolic response to acute kidney injury.

Acute kidney injury (AKI) is extremely prevalent among hospitalizations and presents a significant risk for the development of chronic kidney disease and increased mortality. Ischemia caused by shock, trauma, and transplant are common causes of AKI. To attenuate ischemic AKI therapeutically, we need a better understanding of the physiological and cellular mechanisms underlying damage. Instances of ischemia are most damaging in proximal tubule epithelial cells (PTECs) where hypoxic signaling cascades, and perhaps more rapidly, posttranslational modifications (PTMs), act in concert to change cellular metabolism. Here, we focus on the effects of the understudied PTM, lysine succinylation. We have previously shown a protective effect of protein hypersuccinylation on PTECs after depletion of the desuccinylase sirtuin5. General trends in the results suggested that hypersuccinylation led to upregulation of peroxisomal activity and was protective against kidney injury. Included in the list of changes was the Parkinson's-related deglycase Park7. There is little known about any links between peroxisome activity and Park7. In this study, we show in vitro and in vivo that Park7 has a crucial role in protection from AKI and upregulated peroxisome activity. These data in combination with published results of Park7's protective role in cardiovascular damage and chronic kidney disease lead us to hypothesize that succinylation of Park7 may ameliorate oxidative damage resulting from AKI and prevent disease progression. This novel mechanism provides a potential therapeutic mechanism that can be targeted.NEW & NOTEWORTHY Succinylation is an understudied posttranslational modification that has been shown to increase peroxisomal activity. Furthermore, increased peroxisomal activity has been shown to reduce oxidative stress and protect proximal tubules after acute kidney injury. Analysis of mass spectrometry succinylomic and proteomic data reveals a novel role for Parkinson's related Park7 in mediating Nrf2 antioxidant response after kidney injury. This novel protection pathway provides new insights for kidney injury prevention and development of novel therapeutics.

SIRT7 promotes the proliferation and migration of anaplastic thyroid cancer cells by regulating the desuccinylation of KIF23.

OBJECTIVE: This study was designed to investigate the regulatory effects of kinesin family member (KIF) 23 on anaplastic thyroid cancer (ATC) cell viability and migration and the underlying mechanism. METHODS: Reverse transcription-quantitative polymerase chain reaction (RT-qPCR) was used to analyze the levels of KIF23 in ATC cells. Besides, the effects of KIF23 and sirtuin (SIRT) 7 on the viability and migration of ATC cells were detected using cell counting kit-8, transwell and wound healing assays. The interaction between SIRT7 and KIF23 was evaluated by co-immunoprecipitation (Co-IP) assay. The succinylation (succ) of KIF23 was analyzed by western blot. RESULTS: The KIF23 expression was upregulated in ATC cells. Silencing of KIF23 suppressed the viability and migration of 8505C and BCPAP cells. The KIF23-succ level was decreased in ATC cells. SIRT7 interacted with KIF23 to inhibit the succinylation of KIF23 at K537 site in human embryonic kidney (HEK)-293T cells. Overexpression of SIRT7 enhanced the protein stability of KIF23 in HEK-293T cells. Besides, overexpression of KIF23 promoted the viability and migration of 8505C and BCPAP cells, which was partly blocked by silenced SIRT7. CONCLUSIONS: SIRT7 promoted the proliferation and migration of ATC cells by regulating the desuccinylation of KIF23.

LncRNA SH3PXD2A-AS1 facilitates cisplatin resistance in non-small cell lung cancer by regulating FOXM1 succinylation.

BACKGROUND: Long noncoding RNAs (lncRNAs) play vital regulatory functions in non-small cell lung cancer (NSCLC). Cisplatin (DDP) resistance has significantly decreased the effectiveness of DDP-based chemotherapy in NSCLC patients. This study aimed to investigate the effects of SH3PXD2A antisense RNA 1 (SH3PXD2A-AS1) on DDP resistance in NSCLC. METHODS: Proliferation and apoptosis of DDP-resistant NSCLC cells were detected using cell counting kit-8 and flow cytometry assays. The interaction between SH3PXD2A-AS1 and sirtuin 7 (SIRT7) was assessed using co-immunoprecipitation (Co-IP), RNA pull-down, RNA immunoprecipitation (RIP), RNA fluorescence in situ hybridization, and immunofluorescence assays, while succinylation (SUCC) of Forkhead Box M1 (FOXM1) was analyzed by IP and Western blot assays. The role of SH3PXD2A-AS1 in vivo was explored using a xenografted tumor model. RESULTS: Expression of SH3PXD2A-AS1 was found elevated in DDP-resistant NSCLC cells, while it's knocking down translated into suppression of cell viability and promotion of apoptosis. Moreover, silencing of SH3PXD2A-AS1 resulted in decreased FOXM1 protein level and enhanced FOXM1-SUCC protein level. The SIRT7 was found to interact with FOXM1, translating into inhibition of FOXM1 SUCC at the K259 site in human embryonic kidney (HEK)-293T cells. Overexpressing of SIRT7 reversed the increase of FOXM1-SUCC protein level and apoptosis, and the decrease of cell viability induced by silencing of SH3PXD2A-AS1. In tumor-bearing mice, SH3PXD2A-AS1 inhibition suppressed tumor growth and the protein levels of Ki67, SIRT7, and FOXM1. CONCLUSION: SH3PXD2A-AS1 promoted DDP resistance in NSCLC cells by regulating FOXM1 SUCC via SIRT7, offering a promising therapeutic approach for NSCLC.

SIRT5-mediated PRKAA2 succinylation ameliorates apoptosis of human placental trophoblasts in hypertensive disorder complicating pregnancy.

PURPOSE: Hypertensive disorder complicating pregnancy (HDCP) is a serious clinical disorder syndrome during pregnancy. This study aims at finding novel targets for HDCP therapy. METHODS: HDCP-related mRNAs were firstly screened out and subjected to gene enrichment analysis. We chose protein kinase AMP-activated catalytic subunit alpha 2 (PRKAA2) as the research object. Thirty-nine HDCP patients at 32 to 40 weeks of gestation were selected as the HDCP group, and 39 normal controls who received cesarean section delivery at 37-42 weeks of pregnancy were enrolled in this study. Chorionic villi samples were collected within 30 min of delivery. The apoptosis of isolated placental trophoblasts was monitored to investigate the regulatory role of PRKAA2. RESULTS: PRKAA2 expression was further proven to be enhanced in the placental tissues of HDCP patients compared with that of normal puerpera. Subsequently, the results of flow cytometry analysis and western blot indicated that PRKAA2 overexpression accelerated primary placental cell apoptosis, while its knockdown attenuated cell apoptosis. Mechanistically, we determined that the level of PRKAA2 succinylation was elevated in the placental tissue of HDCP patients. Through in vitro succinylation assay and mutagenesis, we confirmed that sirtuin 5 (SIRT5) interacts with PRKAA2 at K69 and K260 to induce PRKAA2 desuccinylation. SIRT5 regulated primary HDCP cell apoptosis through PRKAA2. Finally, the animal study revealed that PRKAA2 elevates the systolic blood pressure of HDCP rat model. CONCLUSION: Our findings indicated that SIRT5-mediated PRKAA2 succinylation modulates placental cell apoptosis in HDCP, suggesting that PRKAA2 is a potential therapeutic target for HDCP treatment.

Astragaloside IV protects renal tubular epithelial cells against oxidative stress-induced injury by upregulating CPT1A-mediated HSD17B10 lysine succinylation in diabetic kidney disease.

Tubular injury and oxidative stress are involved in the pathogenesis of diabetic kidney disease (DKD). Astragaloside IV (ASIV) is a natural antioxidant. The effects and underlying molecular mechanisms of ASIV on DKD have not been elucidated. The db/db mice and high-glucose-stimulated HK2 cells were used to evaluate the beneficial effects of ASIV in vivo and in vitro. Succinylated proteomics was used to identify novel mechanisms of ASIV against DKD and experimentally further validated. ASIV alleviated renal dysfunction and proteinuria, downregulated fasting blood glucose, and upregulated insulin sensitivity in db/db mice. Meanwhile, ASIV alleviated tubular injury, oxidative stress, and mitochondrial dysfunction in vivo and in vitro. Mechanistically, ASIV reversed downregulated 17beta-hydroxysteroid dehydrogenase type 10 (HSD17B10) lysine succinylation by restoring carnitine palmitoyl-transferase1alpha (Cpt1a or CPT1A) activity in vivo and in vitro. Molecular docking and cell thermal shift assay revealed that ASIV may bind to CPT1A. Molecular dynamics simulations demonstrated K99 succinylation of HSD17B10 maintained mitochondrial RNA ribonuclease P (RNase P) stability. The K99R mutation of HSD17B10 induced oxidative stress and disrupted its binding to CPT1A or mitochondrial ribonuclease P protein 1 (MRPP1). Importantly, ASIV restored the interaction between HSD17B10 and MRPP1 in vivo and in vitro. We also demonstrated that ASIV reversed high-glucose-induced impaired RNase P activity in HK2 cells, which was suppressed upon K99R mutation of HSD17B10. These findings suggest that ASIV ameliorates oxidative stress-associated proximal tubular injury by upregulating CPT1A-mediated K99 succinylation of HSD17B10 to maintain RNase P activity.

Proteomic analysis of baicalin intervention on protein expression and modification in the hippocampus of Alzheimer's disease model rat.

OBJECTIVE: We aimed to explore the treatment effect and therapeutic mechanisms of baicalin in Alzheimer's disease (AD). METHODS: The AD rat model was established by intracerebroventricular injection of Aß1-40, with rats in the baicalin group receiving baicalin intraventricular injections. Morris Water Maze and Hematoxylin-eosin (H&E) Staining were employed to detect the successful model construction and baicalin treatment effect. The proteins extracted from the hippocampus were subjected to proteomics analysis. Bioinformatics technology was employed for differential protein screening, functional classification, and enrichment. Western Blot was employed to validate the expressions of differentially expressed proteins (DEPs) and the protein modification alternations. RESULTS: Water maze test confirmed the successful AD model construction and baicalin can improve learning and memory abilities. A total of 26 DEPs associated with 28 Gene Ontology (GO) functions were identified in the model and 32 DEPs were obtained between the baicalin group and the model. Bioinformatics analysis demonstrated that AD occurrence resulted in neuronal dysfunction and was associated with immune responses. The baicalin therapeutic effect on AD may be associated with metabolic processes, vitamin response, angiogenesis regulation, and fatty acid response. Immunoglobulin heavy constant mu (Ighm) and Immunoglobulin G2a (IgG2a) exhibited significant increases in AD and baicalin attenuated their expressions, while Fatty acid desaturase 1 (Fads1) exhibited a significantly diminished expression and baicalin could reverse the trend. Succinylation detection exhibited the differentially expressed at 35 kD between the model and baicalin group. CONCLUSION: Baicalin intervention may ameliorate cognitive impairment in AD rats by modulating the expressions of proteins and the succinylation modifications.