bims-prolim 2025-04-06 papers

bims-prolim

Biomed News

on Protein lipidation, metabolism and cancer

Issue of 2025–04–06
23 papers selected by
Bruna Martins Garcia, CABIMER

The emerging role of protein L-lactylation in metabolic regulation and cell signalling.
Protocol for label-free quantitative lysine lactylproteome profiling.
The role of metabolite sensors in metabolism-immune interaction: New targets for immune modulation.
Functions and mechanisms of non-histone post-translational modifications in cancer progression.
O -GlcNAc modifications regulate lamin A tail processing.
Mechanisms and Functional Implications of ZDHHC5 in Cellular Physiology and Disease.
Flavin transferase ApbE: from discovery to applications.
Potential role of lactylation in intrinsic immune pathways in lung cancer.
Gut flora-derived succinate exacerbates Allergic Airway Inflammation by promoting protein succinylation.
Spatial alteration of metabolites in diabetic cortical cataracts: New insight into lactate.
Targeting lactylation and the STAT3/CCL2 axis to overcome immunotherapy resistance in pancreatic ductal adenocarcinoma.
Defective anterograde protein-trafficking contributes to endoplasmic reticulum-stress in a CLN1 disease model.
Sirt5 regulates chondrocyte metabolism and osteoarthritis development through protein lysine malonylation.
Consistency of gene expression and protein acetylation modification with physiological and biochemical changes responding to the deficiency of histone deacetylase 1 homolog in Monascus ruber.
Protein succinylome analysis identifies citrate synthase as a central regulator of osteoclast metabolic activity.
O-GlcNAc transferase-mediated O-GlcNAcylation of CD36 against myocardial ischemia-reperfusion injury.
Glycolysis-Derived Lactate Induces ACSL4 Expression and Lactylation to Activate Ferroptosis during Intervertebral Disc Degeneration.
In-depth characterization of S-glutathionylation in ventricular myosin light chain 1 across species by top-down proteomics.
In vivo validation of the palmitoylation cycle as a therapeutic target in NRAS -mutant cancer.
Pan-PTM profiling identifies post-translational modifications associated with exceptional longevity and preservation of skeletal muscle function in Drosophila.
Lysyl hydroxylase 2 glucosylates collagen VI to drive lung cancer progression.
Posttranslational Modification in Bone Homeostasis and Osteoporosis.
Factors Affecting Pathological Amyloid Protein Transformation: From Post-Translational Modifications to Chaperones.

Nat Metab. 2025 Apr 02.

The emerging role of protein L-lactylation in metabolic regulation and cell signalling.

Haowen Ren, Yuwei Tang, Di Zhang.

L-Lactate has emerged as a crucial metabolic intermediate, moving beyond its traditional view as a mere waste product. The recent discovery of L-lactate-driven protein lactylation as a post-translational modification has unveiled a pathway that highlights the role of lactate in cellular signalling. In this Perspective, we explore the enzymatic and metabolic mechanisms underlying protein lactylation and its impacts on both histone and non-histone proteins in the contexts of physiology and diseases. We discuss growing evidence suggesting that this modification regulates a wide range of cellular functions and is involved in various physiological and pathological processes, such as cell-fate determination, development, cardiovascular diseases, cancer and autoimmune disorders. We propose that protein lactylation acts as a pivotal mechanism, integrating metabolic and signalling pathways to enable cellular adaptation, and highlight its potential as a therapeutic target in various diseases.

DOI: https://doi.org/10.1038/s42255-025-01259-0
STAR Protoc. 2025 Apr 03. pii: S2666-1667(25)00132-7. [Epub ahead of print]6(2): 103726

Protocol for label-free quantitative lysine lactylproteome profiling.

Yu Ran, Qiqing Yang, Lianqi Zhou, Heyu Li, Bing Yang, Long Zhang.

  L-lactate has been recognized as an essential molecule for signaling and metabolic balance. Lactylation, a post-translational modification (PTM) derived from L-lactate, is commonly observed on various proteins and plays essential roles in cellular processes. Here, we present a protocol to globally profile the lactylation proteome and perform label-free quantification. We provide steps for cell preparation, protein extraction, digestion, peptide desalting, and lactylpeptide enrichment. Additionally, we outline the parameters for liquid chromatography-tandem mass spectrometry (LC-MS/MS) analysis. For complete details on the use and execution of this protocol, please refer to Li et al.1.

Keywords:  Mass spectrometry; Protein expression and purification; Proteomics

DOI:  https://doi.org/10.1016/j.xpro.2025.103726
Clin Transl Med. 2025 Apr;15(4): e70294

The role of metabolite sensors in metabolism-immune interaction: New targets for immune modulation.

Qiqing Yang, Ce Guo, Long Zhang.

  Recent advancements in immunometabolism have highlighted the critical role of metabolite sensors in regulating immune responses. Metabolites such as lactate, succinate, itaconate, and β-hydroxybutyrate influence immune cell function by interacting with specific sensors. These metabolites act as signaling molecules, linking cellular metabolic changes to immune responses. Lactate, a metabolite commonly produced under hypoxic conditions, has emerged as a major regulator of innate immunity. Key enzymes, including AARS1 and AARS2, function as intracellular lactate sensors, catalyzing lactylation on proteins like cGAS, which plays a central role in DNA sensing and immune activation. The lactylation of cGAS inhibits its activity, modulating immune responses by balancing inflammation and immune tolerance. Metabolite sensors, like MCT1, also contribute to immune modulation, particularly in cancer and chronic inflammatory diseases. Therapeutically, targeting these sensors offers potential for restoring immune function, especially in cancer immunotherapy. However, challenges in specificity, off-target effects, and long-term safety require further investigation. This article explores the emerging role of metabolite sensors in immune regulation, with a focus on lactate sensors, and outlines potential therapeutic strategies to enhance immune responses in metabolic diseases.

Keywords:  immune modulation; inhibitor research and development; lactylation; metabolite sensors

DOI:  https://doi.org/10.1002/ctm2.70294
Cell Death Discov. 2025 Mar 31. 11(1): 125

Functions and mechanisms of non-histone post-translational modifications in cancer progression.

Zongyang Li, Tao Zhu, Yushu Wu, Yongbo Yu, Yunjiang Zang, Lebo Yu, Zhilei Zhang.

Protein post-translational modifications (PTMs) refer to covalent and enzymatic alterations to folded or nascent proteins during or after protein biosynthesis to alter the properties and functions of proteins. PTMs are modified in a variety of types and affect almost all aspects of cell biology. PTMs have been reported to be involved in cancer progression by influencing multiple signaling pathways. The mechanism of action of histone PTMs in cancer has been extensively studied. Notably, evidence is mounting that PTMs of non-histone proteins also play a vital role in cancer progression. In this review, we provide a systematic description of main non-histone PTMs associated with cancer progression, including acetylation, lactylation, methylation, ubiquitination, phosphorylation, and SUMOylation, based on recent studies.

DOI: https://doi.org/10.1038/s41420-025-02410-2
bioRxiv. 2025 Mar 13. pii: 2025.03.11.642699. [Epub ahead of print]

O -GlcNAc modifications regulate lamin A tail processing.

Katherine Augspurger, Elizabeth Martin, Jason Maynard, Kevin Welle, Sina Ghaemmaghami, Al Burlingame, Barbara Panning, Abby Buchwalter.

Lamin A processing is highly regulated, and necessary for proper assembly of the nuclear lamina facilitating its role in nuclear structure and chromatin organization. Pre-lamin A is first farnesylated, and then a short C-terminal peptide is cleaved to produce mature lamin A. O -GlcNAc Transferase (OGT), a glucose sensitive post-translational modification enzyme, is a potential regulator for lamin A processing. To explore the role of OGT in lamin A biogenesis, we examined the effects of OGT levels and OGT inhibition. Variation in OGT dose or inhibition of its activity did not alter endogenous lamin A abundance or distribution. To more directly test the regulatory effects of O -GlcNAcylation on lamin A, we adapted a tail cleavage assay. Mutation of an OGT binding motif and O -GlcNAc modification sites reduced tail cleavage efficiency, suggesting that O -GlcNAcylation promotes lamin A processing. Our findings add to the understanding of the regulation of lamin A cleavage and identify a potential link between glucose metabolism and lamina biogenesis.

DOI: https://doi.org/10.1101/2025.03.11.642699
J Lipid Res. 2025 Apr 01. pii: S0022-2275(25)00053-7. [Epub ahead of print] 100793

Mechanisms and Functional Implications of ZDHHC5 in Cellular Physiology and Disease.

Huicong Liu, Shuo Wen, Chang Xu, Xiaohong Kang, Eryan Kong.

  Post-translational lipid modification by palmitoylation is a reversible process crucial for maintaining cellular functionality. The palmitoyl acyltransferase zinc finger Asp-His-His-Cys motif-containing 5 (ZDHHC5) has garnered significant attention due to its roles in neurodegenerative diseases, oncogenesis, and cardiac function. ZDHHC5 recognizes substrates through diverse mechanisms and its activity is regulated by multiple factors. Highly expressed in the brain, liver and heart, ZDHHC5 exerts regulatory functions in various cellular processes through self-regulation and substrate palmitoylation. This review focuses on the regulatory roles of ZDHHC5 in the nervous system including circadian rhythm, tumor, lipid metabolism. Dysfunctions in ZDHHC5 are associated with several diseases, thereby highlighting its potential as a target for novel therapeutic strategies against neurological, lipid metabolic, and oncogenesis.

Keywords:  cancer; cardiac function; neurodegenerative diseases; substrate recognition

DOI:  https://doi.org/10.1016/j.jlr.2025.100793
J Biol Chem. 2025 Mar 26. pii: S0021-9258(25)00302-3. [Epub ahead of print] 108453

Flavin transferase ApbE: from discovery to applications.

Xiaoman Fan, Marco W Fraaije.

  ApbE is a unique, membrane-bound enzyme which covalently attaches a flavin cofactor to specific target proteins. This irreversible post-translational modification is crucial for proper functioning of various bacterial proteins. ApbEs have also been identified in archaea and eukaryotes. This review summarizes current knowledge on the structural and mechanistic properties of this unique protein-modifying enzyme and its recent applications. The relatively small flavin transferase is typically anchored to the outer membrane of bacteria and possesses a conserved flavin-binding domain and a catalytic domain. It recognizes a specific sequence motif of its target proteins, resulting in flavinylation of a threonine or serine. For flavinylation, it depends on magnesium and utilizes FAD as substrate to attach the FMN moiety to the target protein, analogous to phosphorylation. ApbE-mediated flavinylation supports critical bacterial respiratory and metabolic pathways. Recently, ApbE was also shown to be a versatile tool for selectively modifying proteins. Using the flavin-tagging approach, proteins can be decorated with FMN or other flavins. Furthermore, it was demonstrated that ApbE can be employed to turn natural noncovalent flavoproteins into covalent flavoproteins. In summary, ApbE is crucial for the maturation of various flavoproteins by catalyzing covalent flavinylation. While great progress has been made in understanding the role and mode of action of ApbE, there are still many bacterial proteins predicted to be flavinylated by ApbE for which their role is enigmatic. Also, exploration of the potential of ApbE as protein modification tool has just begun. Clearly, future research will generate new ApbE-related insights and applications.

Keywords:  FMN; covalent flavinylation; flavin transferase; post-translational modification

DOI:  https://doi.org/10.1016/j.jbc.2025.108453
Front Pharmacol. 2025 ;16 1533493

Potential role of lactylation in intrinsic immune pathways in lung cancer.

Mengdie Huang, Ye Jin, Dandan Zhao, Xingren Liu.

  Lung cancer, one of the most lethal malignancies, has seen its therapeutic strategies become a focal point of significant scientific attention. Intrinsic immune signaling pathways play crucial roles in anti-tumor immunity but face clinical application challenges despite promising preclinical outcomes. Lactylation, an emerging research focus, may influences lung cancer progression by modulating the functions of histones and non-histone proteins. Recent findings have suggested that lactylation regulates key intrinsic immune molecules, including cGAS-STING, TLR, and RIG-I, thereby impacting interferon expression. However, the precise mechanisms by which lactylation governs intrinsic immune signaling in lung cancer remain unclear. This review presents a comprehensive and systematic analysis of the relationship between lactylation and intrinsic immune signaling pathways in lung cancer and emphasizes the innovative perspective of linking lactylation-mediated epigenetic modifications with immune regulation. By thoroughly examining current research findings, this review uncovers potential regulatory mechanisms and highlights the therapeutic implications of targeting lactylation in lung cancer. Future investigations into the intricate interactions between lactylation and intrinsic immunity are anticipated to unveil novel therapeutic targets and strategies, potentially improving patient survival outcomes.

Keywords:  RIG-I; TLR; cGAS-STING; lactylation; lung cance

DOI:  https://doi.org/10.3389/fphar.2025.1533493
Redox Biol. 2025 Mar 28. pii: S2213-2317(25)00136-3. [Epub ahead of print]82 103623

Gut flora-derived succinate exacerbates Allergic Airway Inflammation by promoting protein succinylation.

Chao Wang, Xin Yu, Xiao Yu, Hui Xiao, Yuemeng Song, Xinlei Wang, Haoyu Zheng, Kai Chen, Yiming An, Zhengjie Zhou, Xiaoping Guo, Fang Wang.

  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.

Keywords:  Allergic airway inflammation; Gut flora; Succinate; Succinylation

DOI:  https://doi.org/10.1016/j.redox.2025.103623
Exp Eye Res. 2025 Mar 27. pii: S0014-4835(25)00132-0. [Epub ahead of print]255 110361

Spatial alteration of metabolites in diabetic cortical cataracts: New insight into lactate.

Pengfei Li, Miaomiao Wu, Rong Wang, Guowei Zhang, Lihua Kang, Huaijin Guan, Min Ji.

  This study aimed to use metabolomics to accurately reveal alterations in metabolites and potential regulatory mechanisms in patients with diabetic cortical cataracts (DCC). We first collected cortical samples from different pathological areas of the same lens in DCC patients for metabolomics. Then, we used transcriptomic analysis to study lactate's effect on gene expression in human lens epithelial cells (HLECs). An in vitro rat lens culture assay evaluated lactate's impact on lens transparency, and WB and immunofluorescence assessed lactate-induced apoptosis and oxidative damage in rat LECs. Furthermore, CHIP sequencing and LC-MS identified H3K18la separately modified genes and potential lactylation proteins in HLECs. Immunoprecipitation validated lactylation levels of proteins. Our findings identified 11 upregulated and 18 downregulated metabolites in the opacity zone of LFCs (OZ-LFCs) compared to the clear zone (CZ-LFCs) in DCC patients. We confirmed the differential lactate content between OZ-LFCs and CZ-LFCs and, through transcriptomic analysis, discovered that lactate affects gene expression, protein metabolism, and DNA repair in primary Human Lens epithelial cells (HLECs). Lactate-induced apoptosis and DNA repair hastened lens opacity in a high-sugar rat lens culture model. Lactylation-MS and H3K18la-ChIP sequencing revealed 591 H3K18la-modified genes and 953 lactylation proteins in HLECs. PKM2 and NPM1 lactylation was confirmed through immunoprecipitation. These findings improve our grasp of spatial dynamics in DCC patient metabolomics and suggest a new research path into lactylation modification to understand lactate's role in cataract formation.

Keywords:  Diabetic cortical cataract; Lactate; Lactylation; Metabolomics

DOI:  https://doi.org/10.1016/j.exer.2025.110361
J Clin Invest. 2025 Apr 01. pii: e191422. [Epub ahead of print]135(7):

Targeting lactylation and the STAT3/CCL2 axis to overcome immunotherapy resistance in pancreatic ductal adenocarcinoma.

Qun Chen, Hao Yuan, Michael S Bronze, Min Li.

Metabolic reprogramming in pancreatic ductal adenocarcinoma (PDAC) fosters an immunosuppressive tumor microenvironment (TME) characterized by elevated lactate levels, which contribute to immune evasion and therapeutic resistance. In this issue of the JCI, Sun, Zhang, and colleagues identified nonhistone ENSA-K63 lactylation as a critical regulator that inactivates PP2A, activates STAT3/CCL2 signaling, recruits tumor-associated macrophages (TAMs), and suppresses cytotoxic T cell activity. Targeting ENSA-K63 lactylation or CCL2/CCR2 signaling reprograms the TME and enhances the efficacy of immune checkpoint blockade (ICB) in PDAC preclinical models. This work provides critical insights into the metabolic-immune crosstalk in PDAC and highlights promising therapeutic strategies for overcoming immune resistance and improving patient outcomes.

DOI: https://doi.org/10.1172/JCI191422
Neurobiol Dis. 2025 Mar 28. pii: S0969-9961(25)00106-8. [Epub ahead of print] 106890

Defective anterograde protein-trafficking contributes to endoplasmic reticulum-stress in a CLN1 disease model.

Nisha Plavelil, Abhilash P Appu, K C Gopal, Avisek Mondal, Neil Perkins, Anil B Mukherjee.

  Lysosomal storage disorders (LSDs) represent 70 inherited metabolic diseases, in most of which neurodegeneration is a devastating manifestation. The CLN1 disease is a fatal neurodegenerative LSD, caused by inactivating mutations in the CLN1 gene encoding palmitoyl-protein thioesterase-1 (PPT1). S-palmitoylation, a reversable posttranslational modification by saturated fatty acids (generally palmitate) facilitates endosomal trafficking of many proteins, especially in the brain. While palmitoyl-acyltransferases (called ZDHHCs) catalyze S-palmitoylation, depalmitoylation is mediated by palmitoyl-protein thioesterases (PPTs). We previously reported that in Cln1-/- mice, which mimic human CLN1-disease, endoplasmic reticulum (ER)-stress leads to unfolded protein response (UPR) contributing to neurodegeneration. However, the mechanism underlying ER-stress has remained elusive. The anterograde (ER to Golgi) protein-trafficking is mediated via COPII (coat protein complex II) vesicles, whereas the retrograde transport (Golgi to ER) is mediated by COPI vesicles. We hypothesized that dysregulated anterograde protein-trafficking causing stagnation of proteins in the ER leads to ER-stress in Cln1-/- mice. We found that the levels of five COPII vesicle-associated proteins (i.e. Sar1, Sec23, Sec24, Sec13 and Sec31) are significantly higher in the ER-fractions of cortical tissues from Cln1-/- mice compared with those from their WT littermates. Remarkably, all COPII proteins, except Sec13, undergo S-palmitoylation. Moreover, CLN8, a Batten disease-protein, requires dynamic S-palmitoylation (palmitoylation-depalmitoylation) for ER-Golgi trafficking. Intriguingly, Ppt1-deficiency in Cln1-/- mice impairs ER-Golgi trafficking of Cln8-protein along with several other COPII-associated proteins. We propose that impaired anterograde trafficking causes excessive accumulation of proteins in the ER causing ER-stress and UPR contributing to neurodegeneration in CLN1 disease.

Keywords:  CLN1 disease; ER-stress; Lysosomal storage disease; Neurodegeneration; Palmitoyl-protein thioesterase-1; S-palmitoylation; Unfolded protein response

DOI:  https://doi.org/10.1016/j.nbd.2025.106890
Arthritis Rheumatol. 2025 Apr 02.

Sirt5 regulates chondrocyte metabolism and osteoarthritis development through protein lysine malonylation.

Huanhuan Liu, Anupama Binoy, Siqi Ren, Thomas C Martino, Anna E Miller, Craig R G Willis, Shivakumar R Veerabhadraiah, Joanna Bons, Jacob P Rose, Birgit Schilling, Michael J Jurynec, Shouan Zhu.

OBJECTIVES: Chondrocyte metabolic dysfunction plays an important role in osteoarthritis (OA) development during aging and obesity. Protein post-translational modifications (PTMs) have recently emerged as an important regulator of cellular metabolism. We aim to study one type of PTM, lysine malonylation (MaK) and its regulator Sirt5 in OA development.
METHODS: Human and mouse cartilage tissues were used to measure SIRT5 and MaK levels. Both systemic and cartilage-specific conditional knockout mouse models were subject to high-fat diet (HFD) treatment to induce obesity and OA. Proteomics analysis was performed in Sirt5-/- and WT chondrocytes. SIRT5 mutation was identified in the Utah Population Database (UPDB).
RESULTS: We found that SIRT5 decreases while MAK increases in the cartilage during aging. A combination of Sirt5 deficiency and obesity exacerbates joint degeneration in a sex dependent manner in mice. We further delineate the malonylome in chondrocytes, pinpointing MaK's predominant impact on various metabolic pathways such as carbon metabolism and glycolysis. Lastly, we identified a rare coding mutation in SIRT5 that dominantly segregates in a family with OA. The mutation results in substitution of an evolutionally invariant phenylalanine (Phe-F) to leucine (Leu-L) (F101L) in the catalytic domain. The mutant protein results in higher MaK level and decreased expression of cartilage ECM genes and upregulation of inflammation associated genes.
CONCLUSIONS: We found that Sirt5 mediated MaK is an important regulator of chondrocyte cellular metabolism and dysregulation of Sirt5-MaK could be an important mechanism underlying aging and obesity associated OA development.

DOI: https://doi.org/10.1002/art.43164
Int J Biol Macromol. 2025 Apr 01. pii: S0141-8130(25)03037-5. [Epub ahead of print]308(Pt 4): 142485

Consistency of gene expression and protein acetylation modification with physiological and biochemical changes responding to the deficiency of histone deacetylase 1 homolog in Monascus ruber.

Fufang Tang, Yuehan Zhao, Zejing Mao, Yueyan Huang, Yifan Hu, Baixue Liu, Yanchun Huang, Yanchun Shao.

  Monascus species can produce several secondary metabolites (SMs), which are regulated by a complex network. Histone deacetylase 1 (Hda1) has been demonstrated as a critical regulator of SMs and developmental processes; however, little is known about Hda1 in Monascus spp. (named MrHda1). In this study, strain ΔMrHda1 was generated for phenotypic assays. The results indicated that MrHda1 inactivation led to reduced conidia and ascospores but significantly increased pigment and citrinin production. Transcriptome data showed that MrHda1 deletion upregulated the expression of most genes in the glycolytic pathway as well as the gene clusters involved in pigment and citrinin biosynthesis. Western blotting (WB) and label-free acetylome data indicated that MrHda1 deletion significantly enhanced lysine acetylation modifications on the H3 subunit and enzymes involved in the citric acid cycle. Site-directed mutation demonstrated that the production of pigments and citrinin directly correlates with the acetylation level of histone H3 lysine 18 (H3K18). These findings provide new insights into the essential functions of MrHda1 and its regulation mechanisms on the SMs.

Keywords:  Histone deacetylase; Monascus ruber; Secondary metabolism

DOI:  https://doi.org/10.1016/j.ijbiomac.2025.142485
FEBS J. 2025 Apr 02.

Protein succinylome analysis identifies citrate synthase as a central regulator of osteoclast metabolic activity.

Dayoung Yu, Yue Gao, Marcin Luzarowski, Elisabeth Seebach, Thomas Heitkamp, Michael Börsch, Thomas Ruppert, Katharina F Kubatzky.

  Tumour necrosis factor ligand superfamily member 11 (TNFSF11; RANKL) and macrophage colony-stimulating factor 1 receptor (M-CSF) differentiate macrophages into osteoclasts. This process is characterised by changes in metabolic activity that support energy-consuming processes. Treatment with RANKL triggers a phenotype of accelerated metabolism with enhanced glycolysis and an initial disruption of the tricarboxylic acid cycle (TCA) through increased expression of the enzyme aconitate decarboxylase (ACOD1), which results in an upregulation of intracellular succinate levels. Succinate then causes post-translational succinylation of lysine residues. ACOD1 as an inducer of protein succinylation and the desuccinylase NAD-dependent protein deacylase sirtuin-5, mitochondrial (SIRT5) are regulated differentially, and the initially high expression of ACOD1 decreases towards the end of differentiation, whereas SIRT5 levels increase. To mimic the effect of protein succinylation, diethyl succinate or a SIRT5 inhibitor was added during differentiation, which reduced the formation of large osteoclasts, showing its relevance for osteoclastogenesis. To identify succinylated proteins, we used an immunoaffinity-based liquid chromatography-tandem mass spectrometry (LC-MS/MS) approach. Most lysine succinylated proteins were mitochondrial metabolic enzymes. Citrate synthase (CS), the enzyme catalysing the first reaction of the TCA cycle, showed a notable difference in succinylation levels before and after RANKL stimulation, with succinylation detected exclusively in stimulated cells. Immunoprecipitation assays confirmed CS succinylation. Using whole cell extracts, we observed that RANKL treatment decreased CS activity in a concentration-dependent manner. This suggests that CS could be critical in the context of energy production during osteoclastogenesis and that protein succinylation modulates the differentiation program of osteoclasts.

Keywords:  PTM scan; RANKL; citrate synthase; metabolism; mitochondria; osteoclast; post‐translational modification; succinylation

DOI:  https://doi.org/10.1111/febs.70090
Tissue Cell. 2025 Mar 23. pii: S0040-8166(25)00158-2. [Epub ahead of print]95 102878

O-GlcNAc transferase-mediated O-GlcNAcylation of CD36 against myocardial ischemia-reperfusion injury.

Kechao Zhao, Laisha Yan, Xinyi Sun, Xiaoyan Hu.

  CD36 affects lipid metabolism and is involved in the development of myocardial infarction (MI). O-GlcNAcylation is a promising therapeutic target for myocardial ischemia-reperfusion (I/R) injury. This study aimed to investigate the effects of CD36 on myocardial I/R injury and its O-GlcNAcylation. H9C2 cardiomyocytes were induced by hypoxia/reoxygenation (H/R), and phenotypes were evaluated using cell counting kit-8, EdU assay, flow cytometry, and TUNEL assay. The O-GlcNAcylation was evaluated by immunoprecipitation, immunoblotting, and cycloheximide chase assay. The role of CD36 in vivo was analyzed by TTC staining and TUNEL assay. The results showed that CD36 protein levels were downregulated in I/R rats and H/R-induced H9C2 cells. OGT and O-GlcNAcylation levels were decreased by H/R. Overexpression of CD36 or OGT promoted cell proliferation and inhibited apoptosis of H/R-treated cells. Moreover, OGT facilitated the O-GlcNAcylation of CD36 at S195 site and enhanced CD36 protein stability. Knockdown of CD36 abrogated the effects of cellular behaviors caused by OGT, and CD36 mutation at S195 site reversed the promotion of proliferation and lipid uptake and the inhibition of apoptosis induced by wild-type CD36. Additionally, overexpression of CD36 attenuated infarction and apoptosis in the myocardium of rats. In conclusion, OGT-mediated O-GlcNAcylation of CD36 attenuates myocardial I/R injury through promoting the proliferation and inhibiting apoptosis of cardiomyocytes. The findings suggest that targeting CD36 O-GlcNAcylation may be a promising therapy for MI.

Keywords:  CD36; Hypoxia/reoxygenation; Myocardial ischemia-reperfusion injury; O-GlcNAcylation; OGT

DOI:  https://doi.org/10.1016/j.tice.2025.102878
Adv Sci (Weinh). 2025 Apr 02. e2416149

Glycolysis-Derived Lactate Induces ACSL4 Expression and Lactylation to Activate Ferroptosis during Intervertebral Disc Degeneration.

Kaiqiang Sun, Yangyang Shi, Chen Yan, Shunmin Wang, Linhui Han, Fudong Li, Ximing Xu, Yuan Wang, Jingchuan Sun, Zijian Kang, Jiangang Shi.

  The abnormal activation of the inflammatory microenvironment is frequently accompanied by metabolic changes that affect the development of various diseases. However, the relationship between metabolic reprogramming and intervertebral disc degeneration (IVDD) remains unclear. This study aims to reveal the metabolic changes in nucleus pulposus (NPCs) during IVDD and investigate the mechanism of glycolysis-derived lactate on NPCs. Single-cell RNA sequencing reveals that during IVDD, NPCs are characterized by excessively elevated glycolysis, and the resultant lactate causes the dysfunction of NPCs via ferroptosis activation. Mechanistically, lactate results in the transcription of Acyl-CoA Synthetase Long Chain Family Member 4 (ACSL4) via promoting Histon H3K18 lactylation. Interestingly, lactate can also increase the lactylation of ACSL4 at K412 site. In addition, lactate-induced decreased expression of Sirtuin-3 (SIRT3), and further cause the elevation of ACSL4 lactylation. Finally, animal experiments demonstrate that inhibiting glycolysis through gene silencing with adenoviral-associated viruses 9 (AAV9)-si-Ldha or chemical treatment using 2-deoxy-D-glucose can suppress lactate production and lactylation, thereby ameliorating ferroptosis and NPC dysfunction. The findings of this study indicate that lactate plays a crucial role in IVDD by activating ferroptosis and that interventions aimed at lactate production can offer a potential therapeutical option for patients with IVDD.

Keywords:  ACSL4; H3K18la; SIRT3; intervertebral disc degeneration; lactylation; metabolic reprogramming; therapy

DOI:  https://doi.org/10.1002/advs.202416149
J Mol Cell Cardiol. 2025 Mar 30. pii: S0022-2828(25)00058-6. [Epub ahead of print]

In-depth characterization of S-glutathionylation in ventricular myosin light chain 1 across species by top-down proteomics.

Emily A Chapman, Holden T Rogers, Zhan Gao, Hsin-Ju Chan, Francisco J Alvarado, Ying Ge.

  S-glutathionylation (SSG) is increasingly recognized as a critical signaling mechanism in the heart, yet SSG modifications in cardiac sarcomeric proteins remain understudied. Here we identified SSG of the ventricular isoform of myosin light chain 1 (MLC-1v) in human, swine, and mouse cardiac tissues using top-down mass spectrometry (MS)-based proteomics. Our results enabled the accurate identification, quantification, and site-specific localization of SSG in MLC-1v across different species. Notably, the endogenous SSG of MLC-1v was observed in human and swine cardiac tissues but not in mice. Treating non-reduced cardiac tissue lysates with GSSG elevated MLC-1v SSG levels across all three species.

Keywords:  Mass spectrometry; Myosin light chain; Post-translational modification; S-glutathionylation; Sarcomere; Top-down proteomics

DOI:  https://doi.org/10.1016/j.yjmcc.2025.03.012
bioRxiv. 2025 Mar 21. pii: 2025.03.20.644389. [Epub ahead of print]

In vivo validation of the palmitoylation cycle as a therapeutic target in NRAS -mutant cancer.

Matthew Decker, Benjamin J Huang, Timothy Ware, Christopher Boone, Michelle Tang, Julia Ybarra, Aishwarya C Ballapuram, Katrine A Taran, Pan-Yu Chen, Marcos Amendáriz, Camille J Leung, Max Harris, Karensa Tjoa, Henry Hongo, Sydney Abelson, Jose Rivera, Nhi Ngo, Dylan M Herbst, Radu M Suciu, Carlos Guijas, Kimia Sedighi, Taylor Andalis, Elysia Roche, Boer Xie, Yunlong Liu, Catherine C Smith, Elliot Stieglitz, Micah J Niphakis, Benjamin F Cravatt, Kevin Shannon.

Normal and oncogenic Ras proteins are functionally dependent on one or more lipid modifications 1,2 . Whereas K-Ras4b farnesylation is sufficient for stable association with the plasma membrane, farnesylated H-Ras, K-Ras4a, and N-Ras traffic to the Golgi where they must undergo palmitoylation before regulated translocation to cell membranes. N-Ras palmitoylation by the DHHC family of palmitoyl acyl transferases (PATs) and depalmitoylation by ABHD17 serine hydrolases is a dynamic process that is essential for the growth of acute myeloid leukemias (AMLs) harboring oncogenic NRAS mutations 3-6 . Here, we have tested whether co-targeting ABHD17 enzymes and Ras signal output would cooperatively inhibit the proliferation and survival of NRAS -mutant AMLs while sparing normal tissues that retain K-Ras4b function. We show that ABD778, a potent and selective ABHD17 inhibitor with in vivo activity, selectively reduces the growth of NRAS -mutant AML cells in vitro and is synergistic with the allosteric MEK inhibitor PD0325901 (PD901) 7,8 . Similarly, ABD778 and PD901 significantly extended the survival of recipient mice transplanted with three independent primary mouse AMLs harboring an oncogenic Nras G12D driver mutation. Resistant leukemias that emerged during continuous drug treatment acquired by-pass mutations that confer adaptive drug resistance and increase mitogen activated protein kinase (MAPK) signal output. ABD778 augmented the anti-leukemia activity of the pan-PI3 kinase inhibitor pictilisib 9 , the K/N-Ras G12C inhibitor sotorasib 10 , and the FLT3 inhibitor gilteritinib 11 . Co-treatment with ABD778 and gilteritinib restored drug sensitivity in a patient-derived xenograft model of adaptive resistance to FLT3 inhibition. These data validate the palmitoylation cycle as a promising therapeutic target in AML and support exploring it in other NRAS -mutant cancers.

DOI: https://doi.org/10.1101/2025.03.20.644389
NPJ Aging. 2025 Mar 30. 11(1): 23

Pan-PTM profiling identifies post-translational modifications associated with exceptional longevity and preservation of skeletal muscle function in Drosophila.

Suresh Poudel, Chia-Lung Chuang, Him K Shrestha, Fabio Demontis.

Skeletal muscle weakness is a major component of age-associated frailty, but the underlying mechanisms are not completely understood. Drosophila has emerged as a useful model for studying skeletal muscle aging. In this organism, previous lab-based selection established strains with increased longevity and reduced age-associated muscle functional decline compared to a parental strain. Here, we have applied a computational pipeline (JUMPptm) for retrieving information on 8 post-translational modifications (PTMs) from the skeletal muscle proteomes of 2 long-lived strains and the corresponding parental strain in young and old age. This pan-PTM analysis identified 2470 modified sites (acetylation, carboxylation, deamidation, dihydroxylation, mono-methylation, oxidation, phosphorylation, and ubiquitination) in several classes of proteins, including evolutionarily conserved muscle contractile proteins and metabolic enzymes. PTM consensus sequences further highlight the amino acids that are enriched adjacent to the modified site, thus providing insight into the flanking residues that influence distinct PTMs. Altogether, these analyses identify PTMs associated with muscle functional decline during aging and that may underlie the longevity and negligible functional senescence of lab-evolved Drosophila strains.

DOI: https://doi.org/10.1038/s41514-025-00215-2
J Clin Invest. 2025 Apr 01. pii: e189197. [Epub ahead of print]135(7):

Lysyl hydroxylase 2 glucosylates collagen VI to drive lung cancer progression.

Shike Wang, Houfu Guo, Reo Fukushima, Masahiko Terajima, Min Liu, Guan-Yu Xiao, Lenka Koudelková, Chao Wu, Xin Liu, Jiang Yu, Emma Burris, Jun Xu, Alvise Schiavinato, William K Russell, Mitsuo Yamauchi, Xiaochao Tan, Jonathan M Kurie.

  Lysyl hydroxylase 2 (LH2) is highly expressed in multiple tumor types and accelerates disease progression by hydroxylating lysine residues on fibrillar collagen telopeptides to generate stable collagen cross links in tumor stroma. Here, we show that a galactosylhydroxylysyl glucosyltransferase (GGT) domain on LH2-modified type-VI collagen (Col6) to promote lung adenocarcinoma (LUAD) growth and metastasis. In tumors generated by LUAD cells lacking LH2 GGT domain activity, stroma was less stiff, and stable types of collagen cross links were reduced. Mass spectrometric analysis of total and glycosylated peptides in parental and GGT-inactive tumor samples identified Col6 chain α3 (Col6a3), a component of the Col6 heterotrimeric molecule, as a candidate LH2 substrate. In gain- and loss-of-function studies, high Col6a3 levels increased tumor growth and metastatic activity and enhanced the proliferative, migratory, and invasive activities of LUAD cells. LH2 coimmunoprecipitated with Col6a3, and LH2 glucosylated Col6 in an in vitro reaction. Glucosylation increased the integrin-binding and promigratory activities of Col6 in LUAD cells. Col6a3 K2049 was deglucosylated in GGT-inactive tumor samples, and mutagenesis of Col6a3 K2049 phenocopied Col6a3 deficiency or LH2 GGT domain inactivation in LUAD cells. Thus, LH2 glucosylates Col6 to drive LUAD progression. These findings show that the GGT domain of LH2 is protumorigenic, identify Col6 as a candidate effector, and provide a rationale to develop pharmacological strategies that target LH2's GGT domain in cancer cells.

Keywords:  Cell biology; Collagens; Integrins; Lung cancer; Oncology

DOI:  https://doi.org/10.1172/JCI189197
MedComm (2020). 2025 Apr;6(4): e70159

Posttranslational Modification in Bone Homeostasis and Osteoporosis.

Yuzhe Lin, Shide Jiang, Yuming Yao, Hengzhen Li, Hongfu Jin, Guang Yang, Bingzhou Ji, Yusheng Li.

  Bone is responsible for providing mechanical protection, attachment sites for muscles, hematopoiesis micssroenvironment, and maintaining balance between calcium and phosphorate. As a highly active and dynamically regulated organ, the balance between formation and resorption of bone is crucial in bone development, damaged bone repair, and mineral homeostasis, while dysregulation in bone remodeling impairs bone structure and strength, leading to deficiency in bone function and skeletal disorder, such as osteoporosis. Osteoporosis refers to compromised bone mass and higher susceptibility of fracture, resulting from several risk factors deteriorating the balanced system between osteoblast-mediated bone formation and osteoclast-mediated bone resorption. This balanced system is strictly regulated by translational modification, such as phosphorylation, methylation, acetylation, ubiquitination, sumoylation, glycosylation, ADP-ribosylation, S-palmitoylation, citrullination, and so on. This review specifically describes the updating researches concerning bone formation and bone resorption mediated by posttranslational modification. We highlight dysregulated posttranslational modification in osteoblast and osteoclast differentiation. We also emphasize involvement of posttranslational modification in osteoporosis development, so as to elucidate the underlying molecular basis of osteoporosis. Then, we point out translational potential of PTMs as therapeutic targets. This review will deepen our understanding between posttranslational modification and osteoporosis, and identify novel targets for clinical treatment and identify future directions.

Keywords:  bone formation; bone resorption; osteoporosis; posttranslational modification

DOI:  https://doi.org/10.1002/mco2.70159
Biochemistry (Mosc). 2025 Jan;90(Suppl 1): S164-S192

Factors Affecting Pathological Amyloid Protein Transformation: From Post-Translational Modifications to Chaperones.

Vladimir I Muronets, Sofiya S Kudryavtseva, Lidia P Kurochkina, Evgeniia V Leisi, Yulia Yu Stroylova, Elena V Schmalhausen.

  The review discusses the influence of various factors (e.g., post-translational modifications and chaperones) on the pathological transformation of amyloidogenic proteins involved in the onset and development of neurodegenerative diseases (Alzheimer's and Parkinson's diseases) and spongiform encephalopathies of various origin with special focus on the role of α-synuclein, prion protein, and, to a lesser extent, beta-amyloid peptide. The factors investigated by the authors of this review are discussed in more detail, including posttranslational modifications (glycation and S-nitrosylation), cinnamic acid derivatives and dendrimers, and chaperonins (eukaryotic, bacterial, and phage). A special section is devoted to the role of the gastrointestinal microbiota in the pathogenesis of amyloid neurodegenerative diseases, in particular, its involvement in the transformation of infectious prions and possibly other proteins capable of prion-like transmission of amyloidogenic diseases.

Keywords:  chaperons; microbiota; post-translational modifications; prion protein; α-synuclein; β-amyloid

DOI:  https://doi.org/10.1134/S0006297924604003