Journal Of Proteomics[JOURNAL]
Sun
200 papers
Structural dynamics of the TPS/TPP complex in Saccharomyces cerevisiae: Insights from cross-linking mass spectrometry and computational modeling.
Santos RM, Bastos VA, Bagatelli FFM … +8 more , Valente RH, Kurt LU, Lima DB, Oliveira CC, Nogueira FCS, Eleutherio ECA, Carvalho PC, Gomes-Neto F
J Proteomics · 2026 Jan · PMID 40987384 · Publisher ↗
Trehalose, a ubiquitous disaccharide, plays a vital role in cell viability, including pathogenic fungi, making its synthetic pathway a key target for antifungal drug design. The structural details of the trehalose-phosph...
Trehalose, a ubiquitous disaccharide, plays a vital role in cell viability, including pathogenic fungi, making its synthetic pathway a key target for antifungal drug design. The structural details of the trehalose-phosphate synthase (TPS)/trehalose-phosphate phosphatase (TPP) complex, which is responsible for synthesizing trehalose, have remained elusive. Information on the structure and topology of the TPS/TPP complex remains scarce, significantly limiting the mechanistic understanding of trehalose synthesis. This study presents the first overview of the interactions within the Saccharomyces cerevisiae TPS/TPP complex following a 40 °C heat shock, analyzed by cross-linking mass spectrometry (XL-MS) and computational modeling. Our cross-linking data corroborate the UniProt-available AlphaFold models for isolated subunits. Intrinsically disordered regions are suggested for the regulatory subunits, while cross-linking analysis highlights the disordered N-terminus of Tsl1 as an important region in assembling the TPS/TPP complex. Finally, the phosphorylation prediction indicates that Tps3-disordered regions at the N-terminus and the phosphatase-like domains are preferentially phosphorylated, triggering the inhibition of Tps2 activity and halting T6P accumulation. These insights enhance our understanding of the structural dynamics and flexibility of the TPS/TPP complex, opening new avenues for potential therapeutic applications with diminished or no toxicity to humans. SIGNIFICANCE: This research contributes to a comprehensive structural understanding of the TPS/TPP complex in Saccharomyces cerevisiae, a key enzymatic system involved in trehalose synthesis. Trehalose is essential in cellular viability and stress adaptation, particularly in pathogenic fungi, making its biosynthetic pathway a promising target for antifungal drug development. By integrating cross-linking mass spectrometry (XL-MS) and computational modeling, this study uncovers critical interactions and dynamic features of the TPS/TPP complex, including the involvement of intrinsically disordered regions in its regulatory subunits possibly contributing to complex assembly and regulation under heat shock conditions. Furthermore, the phosphorylation predictions shed light on how disordered regions on Tps3 would participate in modulating Tps2 activity to regulate trehalose-phosphate levels, offering insights into the complex's functional dynamics. This work fills a crucial knowledge gap in understanding trehalose biosynthesis and paves the way for novel antifungal strategies with reduced toxicity to human cells.
Anisomelic acid promotes proteasomal degradation of HPV16 E6 via E3 ligase recruitment: A mass spectrometry-based interactome study.
Santos Silva M, Coelho-Rato LS, Delshad N … +5 more , Tarkhova T, Edman J, Paul P, Meinander A, Eriksson JE
J Proteomics · 2026 Jan · PMID 40967472 · Publisher ↗
Human papillomavirus (HPV) is a major driver of cervical and other epithelial cancers, with the viral oncoprotein E6 playing a central role in tumorigenesis by promoting degradation of the tumor suppressor p53. While pro...
Human papillomavirus (HPV) is a major driver of cervical and other epithelial cancers, with the viral oncoprotein E6 playing a central role in tumorigenesis by promoting degradation of the tumor suppressor p53. While prophylactic vaccines prevent infection, there remains a critical need for therapeutic strategies that eliminate established HPV-positive cells. Here, we identify anisomelic acid (AA), a natural diterpenoid, as a novel pharmacological principle that selectively induces the degradation of HPV16 E6. Using cellular thermal shift assay, we demonstrate that AA directly interacts with E6, likely triggering a conformational change that promotes its ubiquitination. Proteomic analysis of the E6 interactome in AA-treated cells revealed consistent enrichment of E3 ubiquitin ligases, including E6AP, UBR4, CDC20, and TRIP12, as well as proteasomal subunits. To our knowledge, this represents the first comprehensive proteomics framework of the HPV16 E6 interactome under small-molecule treatment conditions. These findings support a model in which AA facilitates proteasome-mediated elimination of E6, and the dataset itself provides a timely and valuable resource for HPV biology and therapeutic development.
Potential protein biomarkers for heat tolerance in wheat at seedling and ear peep stages.
Pratap A, Siddique KHM, Taylor NL
J Proteomics · 2026 Jan · PMID 40945775 · Publisher ↗
Heat stress is a major threat to global wheat (Triticum aestivum L.) production, adversely affecting crop yields and grain quality. Understanding wheat's heat tolerance mechanisms is crucial for developing resilient cult...
Heat stress is a major threat to global wheat (Triticum aestivum L.) production, adversely affecting crop yields and grain quality. Understanding wheat's heat tolerance mechanisms is crucial for developing resilient cultivars. This study used targeted proteomics to validate heat-induced changes to protein abundances in seedling and flag leaves of heat-tolerant (Vixen-T) and heat-sensitive (HD2329-S) wheat genotypes. Proteomics samples were collected on days 1, 3 and 5 of heat exposure (32/16 °C day/night for 3 hours per day over 5 days) and day 12 post-recovery. Flag leaf gas exchange was studied under heat treatment during ear peep and significant genotype × heat treatment interactions were observed for all traits. Significant protein abundance changes occurred under heat stress for 15 and 14 proteins at the seedling and ear peep stages, respectively. Two key proteins-DM2 domain-containing protein (r = 0.99) and Rubisco activase (r = 0.96)-showed consistent responses across both developmental stages. Redox homeostasis and protein chaperone pathways emerged as major contributors to wheat heat tolerance. These findings highlight critical protein biomarkers that can support breeding efforts to develop heat-tolerant wheat varieties, offering valuable strategies for sustaining wheat productivity under climate change. SIGNIFICANCE: This study identifies and validates novel protein biomarkers associated with heat tolerance in wheat. These proteins were discovered in our previous study in the flag leaves of four genotypes with contrasting heat responses (tolerant: RAJ3765, HD2932; susceptible: HD2329, HD2733) under short-term heat stress at the ear peep stage. These biomarkers were further validated in two genotypes (tolerant: Vixen; susceptible: HD2329) under short-term heat stress at both seedling and ear peep stages. The validated protein isoforms span key biological processes, including photosynthesis, redox regulation, chromatin remodelling, protein folding, and carbohydrate and secondary metabolism. This panel of protein biomarkers offers a novel molecular framework for breeding heat-tolerant wheat, providing a strategic avenue, utilising targeted proteomics, to sustain yield under rising temperatures.
Peroxiredoxin 6 (Prdx6) and oxidative stress in post-mortem beef tenderization: A proteomic perspective.
Zuo H, Huang L, Huang M … +6 more , Ma X, Zheng C, Holman BWB, Zhang Y, Luo X, Mao Y
J Proteomics · 2026 Jan · PMID 40935016 · Publisher ↗
This research explored the role of peroxiredoxin 6 (Prdx6)-mediated non‑selenium glutathione peroxidase (NSGPx) activity in modulating the tenderization process of beef during post-mortem aging, extending up to 168 h. Sh...
This research explored the role of peroxiredoxin 6 (Prdx6)-mediated non‑selenium glutathione peroxidase (NSGPx) activity in modulating the tenderization process of beef during post-mortem aging, extending up to 168 h. Shear force, NSGPx activity, differential protein abundance, heat shock proteins (HSP70, HSP27), and troponin-T levels were analyzed in beef longissimus lumborum muscles treated with hydrogen peroxide (HO), N-acetylcysteine (NAC), mercaptosuccinic acid (MA), or saline (Control). MA treatment inhibited NSGPx activity and accelerated tenderization compared to NAC. Proteomics revealed that proteins differentially abundant between 0 and 24 h post-mortem were linked to cytoskeleton, extracellular matrix, amino acid metabolism, and apoptosis pathways.MA upregulated HSP70 abundance, oxidative stress, and troponin-T breakdown. HO upregulated HSP70 and HSP27 abundance only within 6-12 h post-mortem. These results demonstrate that oxidative stress treatments modulate protein dynamics during aging, offering insights into strategies to enhance beef tenderness. SIGNIFICANCE: This study highlights peroxiredoxin 6 (Prdx6) as a crucial regulatory element that affects oxidative stress-associated pathways involved in the meat tenderization process during post-mortem beef aging. We demonstrate that inhibiting Prdx6's on‑selenium glutathione peroxidase (NSGPx) enzymatic activity with mercaptosuccinic acid (MA) increases HSP70 abundance and accelerates troponin-T proteolysis through enhanced oxidative stress and calcium signaling pathways. Conversely, antioxidant N-acetylcysteine (NAC) delays tenderization by preserving cytoskeletal integrity. Our TMT-based proteomics further identifies 35 core proteins linking extracellular matrix remodeling, amino acid metabolism, and apoptosis to tenderness modulation. These findings provide the first mechanistic evidence that targeted manipulation of Prdx6 activity can optimize beef aging efficiency. For the meat industry, MA treatment offers a science-driven strategy to reduce tenderization time by >20 % within 24-72 h post-mortem, lowering processing costs while maintaining quality. This work also establishes HSP70 and troponin-T degradation as novel biomarkers for real-time monitoring of oxidative stress in meat processing systems.
Sequence-driven species identification of ZooMS collagen peptide mass fingerprints.
Lawrence T, Buckley M
J Proteomics · 2025 Oct · PMID 40850354 · Publisher ↗
Developments in biomolecular species identification of animal tissues have been ongoing for decades, with collagen peptide mass fingerprinting becoming increasingly used in recent years. However, establishing confidence...
Developments in biomolecular species identification of animal tissues have been ongoing for decades, with collagen peptide mass fingerprinting becoming increasingly used in recent years. However, establishing confidence in the species biomarkers within these fingerprints requires sequence assignment, usually done via LC-ESI-MS/MS-based approaches and correlation with sequence databases. This study develops an approach that allows collagen fingerprints to be matched to sequence databases directly. To do so we create theoretical spectra from in silico digests of publicly available sequences that are then filtered by previously collected proteomic sequence data. These inform on the likely number of collagen post translational modifications, vastly reducing the number of peaks in the theoretical spectra and so making overlapping peptide signals as well as false positives less likely. We retrieved a database containing 211 mammals and tested this approach with spectra of 29 modern reference species and 98 archaeological examples of 10 different families, some for which the taxa were represented in the sequence database, and others that were not. This approach was found to be at least 93 % accurate for predicting the correct family in both modern and archaeological spectra, and capable of species-level identification in some cases. This sequence-driven analysis allows rapid comparison across whole spectra, rather than small sets of markers for a particular taxon, which removes human error from manual identification and ensures that the selected markers derive from the protein of interest, unlike machine-learning methods. SIGNIFICANCE: Species identification using collagen peptide mass fingerprinting is a MALDI-based mass spectrometric method becoming increasingly popular, largely because of its reliance on the dominant protein in the most enduring of biological tissues, bone and tooth dentine. This endurance has great significance to fields such as bioarchaeology and palaeontology, but also applies to processed foodstuffs, for which proteomics-based species identification has been ongoing for decades. However, with this increase in demand, there are greater explorations into a wider range of vertebrate species under investigation, making biomarker selection more challenging. Although the use of DNA-based gene sequence information has been a cornerstone for probability-based proteomic inferences, their use in fingerprint analysis for species identification has remained indirect, where tools such as Mascot's PMF search application may be suitable for protein identification but often struggle with such species-level inferences. Here we introduce a means to create post-translational modification rules based on observation in LC-MS/MS data to generate improved in silico PMFs from DNA-based sequences that greatly reduces search space and confidence matches to taxonomic interpretations of PMFs. This is applicable beyond collagen to any protein for which species identification is needed.
Potential kidney biomarkers in urinary extracellular vesicles from end stage renal disease and post-transplant patients.
Verdugo-Molinares MG, Ku-Centurion M, Cortes-Sanabria L … +6 more , Lugo-Melchor YO, Pinault E, Evangelista L, Ramos-Garcia CO, Marquet P, Melo Z
J Proteomics · 2025 Oct · PMID 40834982 · Publisher ↗
Chronic kidney disease is a multifactorial entity characterized by decreased glomerular filtration rate (GFR). The last stage of the disease requires renal replacement therapy or kidney transplantation. As a disease with...
Chronic kidney disease is a multifactorial entity characterized by decreased glomerular filtration rate (GFR). The last stage of the disease requires renal replacement therapy or kidney transplantation. As a disease with no treatment at earlier stages, and few biomarkers available, proteomics represent an excellent tool searching for new more efficient biomarkers. Urinary extracellular vesicles are an important source of information for kidney alterations, and their collection is not invasive. In this exploratory study, we worked on urine samples collected from patients at Centro Medico Nacional de Occidente in Guadalajara, Jalisco, and isolated urinary extracellular vesicles (uEVs) by ultracentrifugation. Our objective was to compare the Proteomic Profile of uEVs between Mexican patients with normal kidney function, end-stage renal disease, or kidney transplantation. High resolution mass spectrometry analysis reveals alterations in end-stage renal disease regarding the energy metabolism, cytoskeleton organization and cell motility. Proteomic alterations in transplant patients point towards the conservation of fibrotic process. Important proteins such as cystatins can be proposed as candidates for kidney transplant monitoring, while Gelsolin, a protein with an important role in assessing podocyte damage, stands out as a probable marker of chronic kidney disease. Data are available via ProteomeXchange with identifier PXD065380. SIGNIFICANCE: Chronic Kidney disease is a growing public health burden, increasing each year, and favored by major chronic diseases such as diabetes and hypertension. Although Mexico is one of the countries with the highest incidence of chronic kidney disease, proteomics studies involving Mexican patients had not yet been conducted. uEVs are features of particular interest to study the disease and discover biomarkers. We characterized the uEVs proteomic profile in Mexican patients, providing new insights into the pathogenesis of chronic kidney disease and kidney transplantation disorders. We identified promising biomarker candidates for transplant monitoring, and one as an early indicator of ESRD progression. uEVs may serve as a non-invasive platform for renal disease investigation, potentially offering non-invasive biomarkers for patient monitoring as well as mechanistic insights for future research into kidney pathophysiology.
Proteoformics: Current status and future perspectives.
Fang Z, Zhang Y, Feng X … +3 more , Li N, Chen L, Zhan X
J Proteomics · 2025 Oct · PMID 40834981 · Publisher ↗
Proteoforms represent the ultimate structural/functional forms of a gene product, defined by multiple factors, including amino acid sequences, post-translational modifications, spatial conformations, and interactions wit...
Proteoforms represent the ultimate structural/functional forms of a gene product, defined by multiple factors, including amino acid sequences, post-translational modifications, spatial conformations, and interactions with other molecules. The human proteoform diversity significantly exceeds the number of human genes/transcripts, emphasizing the need for advanced analytical methods to characterize this complexity. Two-dimensional gel electrophoresis-liquid chromatography/mass spectrometry (2DE-LC/MS) and top-down MS (TD-MS) are complementary to detect, identify, and quantify the large-scale proteoforms. The emerging AI tools for structural biology such as AlphaFold 3 and D-I-TASSER will enable proteoformics to be high-throughput and precisely predict spatial conformations and molecular interactions. Integrating the large-scale experimental data derived from 2DE-LC/MS and TD-MS with AI-driven high-throughput structural analysis paves the way to deeply understand proteoform diversity and functionality. The combination of advanced 2DE-LC/MS, TD-MS, and AI-driven structural analysis represents a pivotal advancement in proteoformics. This integrated approach enables the comprehensive profiling of proteoforms, providing critical insights into their roles in health care. Such advancements hold promise for predictive, preventive, and personalized medicine, particularly through biomarker discovery and therapeutic target identification. Future developments in high-throughput capabilities and dynamic modeling are expected to address current challenges and further expand the applicability of proteoformics in biological and clinical research. SIGNIFICANCE: Proteoformics is the future of proteomics, whose two main complementary analytical approaches are 2DE-LC/MS and TD-MS. The AI-driven large-cale structural analysis enables to high-throughput and precisely analyze spatial conformations and molecular interactions of proteoforms, which helps to deeply understand proteoform diversity and functionality. Proteoformics holds transformative potential to uncover biomarkers, guide targeted therapies, and advance predictive diagnosis in the context of personalized medicine.
Mining the plasma proteome: Evaluation of enrichment methods for depth and reproducibility.
Roger K, Metatla I, Ceccacci S … +4 more , Wahbi K, Motté L, Chhuon C, Guerrera IC
J Proteomics · 2025 Oct · PMID 40829696 · Publisher ↗
Plasma is a complex biological fluid containing extracellular vesicles (EVs), residual platelets, and soluble proteins. While conventional plasma proteomics typically identifies hundreds of proteins, recent enrichment st...
Plasma is a complex biological fluid containing extracellular vesicles (EVs), residual platelets, and soluble proteins. While conventional plasma proteomics typically identifies hundreds of proteins, recent enrichment strategies have expanded coverage to thousands. It is still unclear whether these methods enrich preferentially different classes of protein and whether they allow for reliable quantification. Here, we compared three common advanced proteomic workflows-Proteograph (Seer), Mag-Net (ReSynBio), and ENRICHplus (PreOmics) as well as EV enrichment obtained by centrifugation. We explore the content in soluble proteins, EV cargo, and platelet-derived proteins after the enrichments. Quantification was evaluated comparing each method to neat plasma using protein coefficient of variation and point-biserial correlation. We quantified an average of ∼4500 proteins with EV centrifugation, ∼4000 with Seer, ∼2800 with ENRICHplus, ∼2300 with Mag-Net, and ∼ 900 with neat plasma. Each method enriched distinct sets of protein signatures: EV preparations were enriched with EV markers such as CD81; ENRICHplus predominantly captured lipoproteins; and Proteograph was enriched for cytokines and hormones. Platelet protein intensity was directly correlated with total protein identifications but did not compromise quantification of low-abundance proteins. Across 50 healthy individuals, Proteograph consistently demonstrated reproducible enrichment and depletion patterns, with some reported exceptions. Our results highlight the strengths and biases of different plasma enrichment strategies. SIGNIFICANCE: This study benchmarks corona formation strategies for enriching low-abundance plasma proteins, including those from platelets and EVs. While enabling deeper proteome coverage compared to depletion methods, these approaches also reshape the intensity landscape and reveal method-specific patterns in protein class enrichment and in quantification repeatability.
Expanding the landscape of lysine acetylation stoichiometry and clinical impact.
Encarnación-Guevara S, Gil J
J Proteomics · 2025 Oct · PMID 40812546 · Publisher ↗
Lysine acetylation, once viewed primarily as a histone mark, is now recognized as a widespread regulator of protein function. Recent breakthroughs in chemical labeling, isotopic tagging workflows, and data-independent ac...
Lysine acetylation, once viewed primarily as a histone mark, is now recognized as a widespread regulator of protein function. Recent breakthroughs in chemical labeling, isotopic tagging workflows, and data-independent acquisition mass spectrometry enable precise, site-specific quantification of acetylation stoichiometry. This quantitative "acetylomics" approach reveals a "rheostat" model, where most acetylation sites exhibit low occupancy, acting as subtle modulators, while a subset of highly acetylated lysines (e.g., p53 C-terminus, AKT1, histones) serve as pivotal regulatory switches in gene expression, metabolism, and cell fate. Site-specific occupancy changes (e.g., p53, PKM2) increasingly serve as robust biomarkers for cancer diagnosis, prognosis, and therapeutic monitoring, often surpassing mRNA or total protein levels. Quantitative acetylation data now guide the development of targeted epigenetic therapies, including HDAC and p300/CBP inhibitors. Beyond oncology, acetylomics can pinpoint metabolic bottlenecks in heart failure, epigenetic deficits in neurodegenerative conditions, and inflammatory signaling nodes. With advances in high-throughput workflows, FFPE and liquid biopsy compatibility, and microfluidic platforms, acetylation stoichiometry is poised for clinical translation. We highlight both the promise and challenges of this emerging dimension of precision medicine, emphasizing the need for integrated multi-omics approaches and robust clinical validation to fully realize the potential of quantitative acetylomics in disease diagnosis and therapy. SIGNIFICANCE: Understanding the extent of acetylation occupancy in proteins, beyond simply determining presence or absence of acetylation, has profound implications for biology and medicine. This review emphasizes the importance of acetylation stoichiometry, connecting advanced proteomic technologies with translational science. We emphasize that quantifying site occupancy reveals which acetylation events truly modulate enzyme function. For instance, it can identify which acetylation events truly modulate enzyme activity or gene expression. Additionally, it can highlight molecular changes in diseases like cancer that are not apparent through qualitative analyses. These quantitative insights pave the way for clinical innovations, including novel biomarkers that stratify patients based on their acetylation profiles and targeted therapies that modulate acetylation levels. In summary, this work highlights the evolving landscape of protein acetylation research over the past two decades and its increasing influence on translational proteomics, celebrating milestones achieved by the global research community.
Global profiling of protein lysine Malonylation during goat intramuscular Preadipocyte differentiation.
Yang C, Li H, Xu Z … +5 more , Lin Y, Wang Y, Huang L, Xiang H, Zhu J
J Proteomics · 2025 Oct · PMID 40812545 · Publisher ↗
Enhancing intramuscular fat (IMF) to improve the quality of livestock product has long been a goal in animal breeding. Recent studies have revealed a strong connection between malonylation and lipid metabolism, yet the f...
Enhancing intramuscular fat (IMF) to improve the quality of livestock product has long been a goal in animal breeding. Recent studies have revealed a strong connection between malonylation and lipid metabolism, yet the function of malonylated proteins in ruminants largely unclear. In the present study, we identified the third day of goat intramuscular preadipocyte differentiation as a critical time point for lipid accumulation, with no significant alterations in malonylation levels. We identified 212 and 216 malonylated proteins on day 0 (d0) and day 3 (d3) of differentiation, respectively, enrichment in pathways such as glycolysis/gluconeogenesis, tight junctions, and actin cytoskeleton regulation. Our findings demonstrate the consistent presence of malonylation during preadipocyte differentiation, with minor quantitative variations, and highlight key malonylated proteins closely associated with lipid metabolism, including acetyl-CoA carboxylase (ACACA), translation control tumor protein 1 (TPT1), phosphoglycerate kinase 1 (PGK1), annexin A6 (ANXA6), and annexin A2 (ANXA2). Collectively, our study uncovers critical malonylated proteins during preadipocyte differentiation, establishing a foundation for exploring their roles in intramuscular fat deposition. SIGNIFICANCE: Currently, efforts are being made to improve meat quality by enhancing intramuscular fat (IMF) deposition, thereby promoting the development of the livestock industry. This study addresses a critical gap in our understanding of malonylation, a key post-translational modification, in livestock. By constructing the first malonylation protein modification map in goats and revealing dynamic changes during intramuscular preadipocyte differentiation, this research offers novel insights into the regulatory roles of malonylation in fat deposition. The findings not only advance the field of livestock proteomics but also provide a theoretical foundation for improving meat quality and exploring metabolic regulation in animals.
Proteomic and transcriptomic analyses reveal new insights into allergens in Ligustrum lucidum pollen.
Vizuet-de-Rueda JC, Montero-Vargas JM, López-Calleja AC … +2 more , Manríquez-Ventura MZT, Teran LM
J Proteomics · 2025 Oct · PMID 40803605 · Publisher ↗
Pollen aeroallergens cause up to 40 % of respiratory allergies and are challenging to control due to their widespread distribution in the environment. The pollen of Ligustrum lucidum (privet) is a significant source of i...
Pollen aeroallergens cause up to 40 % of respiratory allergies and are challenging to control due to their widespread distribution in the environment. The pollen of Ligustrum lucidum (privet) is a significant source of inhalant allergens. However, despite its clinical relevance, the protein composition of L. lucidum pollen remains poorly characterized. Therefore, we employed an integrated proteomic and transcriptomic approach to explore its potential allergen composition, focusing on possible cross-reactivity with Olea europea (olive), a well-studied allergenic relative. Using LC-MS/MS-based proteomics and RNA-seq transcriptomics, we detected 13 of the 15 known olive-like allergens, demonstrating high cross-species conservation. Proteomic analysis identified nine homologous allergens, including Ole e 1, Ole e 2, Ole e 3, Ole e 5, Ole e 6, Ole e 9, Ole e 12, Ole e 13, and Ole e 14. Transcriptomic analysis revealed four additional putative allergens: Ole e 8, Ole e 10, Ole e 11, and Ole e 15. These proteins shared 74-95 % sequence identity with their olive counterparts and exhibited multiple isoforms. Our findings provide a set of L. lucidum pollen potential allergens and highlight the utility of multi-omics in allergen discovery. However, further clinical validation of these putative novel allergens is needed to assess their role in sensitization and cross-reactivity. SIGNIFICANCE: Privet (Ligustrum), a genus within the Oleaceae family, is biologically significant due to its role in triggering allergic respiratory diseases worldwide. As a close relative of olive (Olea europaea) and ash (Fraxinus), privet shares allergenic proteins that contribute to cross-reactivity among sensitized individuals. Climate change has been shown to extend their flowering period, increasing pollen exposure and exacerbating allergic symptoms. Ligustrum is widely used in urban landscaping due to its rapid growth, resistance to pollution, and adaptability to diverse soil conditions, which facilitates its global spread across North America, Europe, Asia, and South America. Notably, L. lucidum is a major sensitizing agent in Mexico City, where 37 % of allergic patients react to its pollen. The first identified allergen, Lig v 1, shares homology with Ole e 1 and Fra e 1. At the same time, Lig v 2 (profilin) mirrors Ole e 2, highlighting the molecular basis for cross-reactivity within the Oleaceae family. Recent proteomic studies have uncovered additional allergens, including enolase, β-1,3-glucanase, and ATP synthase subunits, further elucidating privet's allergenic potential. The absence of genomic data for L. lucidum has hindered research; however, advances in transcriptomic and proteomic approaches have enabled the identification of 13 of 15 known olive-like allergens in privet pollen, paving the way for improved diagnostics and targeted therapies. This underscores the need for further investigation into Ligustrum's allergenic components, particularly as climate change and urbanization amplify its public health impact, as well as, the potential for improved diagnosis and targeted therapies.
Off-site processing of data-dependent and data-independent acquisition shotgun proteomics data with MASSyPupX.
García-Chávez JN, Winkler R
J Proteomics · 2025 Oct · PMID 40754233 · Publisher ↗
The rapid pace of shotgun proteomics data generation presents challenges for timely data analysis. In parallel, the scientific community is creating novel data interpretation tools, such as artificial intelligence, that...
The rapid pace of shotgun proteomics data generation presents challenges for timely data analysis. In parallel, the scientific community is creating novel data interpretation tools, such as artificial intelligence, that have not yet been integrated into commercial software. Off-site data processing with free and open-source software (FOSS) enables the decentralization and scaling of informatics workflows. FOSS platforms also lower the costs of education and research. MASSyPupX is a FOSS mass spectrometry (MS) software collection that runs directly from a USB drive. Alternatively, setting up a MASSyPupX workstation or server provides a ready-to-use and reproducible MS analysis platform. Installed programming languages and libraries support the development of custom MS software and workflows. This paper demonstrates using MASSyPupX to convert and process raw shotgun proteomics data. Raw Thermo files were downloaded from ProteomeXchange and converted to the HUPO community format mzML. Data-dependent acquisition (DDA) data were evaluated with Comet, PeptideProphet, ProteinProphet, ProtyQuant, and the Trans-Proteomic Pipeline. Data-independent acquisition (DIA) shotgun proteomics data were analyzed with DIA-NN. Custom Bash, Python, and R scripts were used to post-process and visualize the results. The MASSyPupX project is hosted at https://codeberg.org/LabABI/MASSyPupX, and the current ISO can be downloaded from https://doi.org/10.5281/zenodo.14618430. The MASSyPupX platform significantly advances shotgun proteomics data processing by offering a free and open-source software (FOSS) solution that is portable, scalable, and accessible. Operating directly from a USB drive or server, this Debian-based Linux distribution enables researchers to analyze data-dependent (DDA) and data- independent (DIA) acquisition proteomics data without installation, decentralizing workflows, reducing costs, and fostering collaboration and mass spectrometry data processing training. With pre-installed programming languages, libraries, and support for tools like Comet, PeptideProphet, DIA-NN, and ProtyQuant, MASSyPupX facilitates reproducible analyses, integrates cutting-edge computational techniques, and provides a user-friendly environment for education, research, and custom workflow development. MASSyPupX democratizes access to advanced proteomics analysis, serving as a versatile tool for advancing biological and medical research through decentralized and cost-effective workflows.
Q2C: A software for managing mass spectrometry facilities.
Lima DB, Ruwolt M, Santos MDM … +3 more , Pu K, Liu F, Carvalho PC
J Proteomics · 2025 Oct · PMID 40752643 · Publisher ↗
We present Q2C, an open-source software designed to streamline mass spectrometer queue management and assess performance based on quality control metrics. Q2C provides a fast and user-friendly interface to visualize proj...
We present Q2C, an open-source software designed to streamline mass spectrometer queue management and assess performance based on quality control metrics. Q2C provides a fast and user-friendly interface to visualize projects queues, manage analysis schedules and keep track of samples that were already processed. Our software includes analytical tools to ensure equipment calibration and provides comprehensive log documentation for machine maintenance, enhancing operational efficiency and reliability. Additionally, Q2C integrates with Google™ Cloud, allowing users to access and manage the software from different locations while keeping all data synchronized and seamlessly integrated across the system. For multi-user environments, Q2C implements a write-locking mechanism that checks for concurrent operations before saving data. When conflicts are detected, subsequent write requests are automatically queued to prevent data corruption, while the interface continuously refreshes to display the most current information from the cloud storage. Finally, Q2C, a demonstration video, and a user tutorial are freely available for academic use at https://github.com/diogobor/Q2C. Data are available from the ProteomeXchange consortium (identifier PXD055186). SIGNIFICANCE: Q2C addresses a critical gap in mass spectrometry facility management by unifying sample queue management with instrument performance monitoring. It ensures optimal instrument utilization, reduces turnaround times, and enhances data quality by dynamically prioritizing and routing samples based on analysis type and urgency. Unlike existing tools, Q2C integrates queue control and QC in a single platform, maximizing operational efficiency and reliability.
Proteomics reveals the role of the EamB transporter from Aeromonas hydrophila LP-2 in biofilm formation.
Xu Q, Tian F, Wang X … +4 more , Lian J, Zhang X, Lin X, Liu Y
J Proteomics · 2025 Oct · PMID 40744271 · Publisher ↗
Biofilms play a pivotal role in the survival and persistence of microorganisms, endowing them with heightened resistance to environmental stressors and antimicrobial agents. The EamB protein, which encodes an inner membr...
Biofilms play a pivotal role in the survival and persistence of microorganisms, endowing them with heightened resistance to environmental stressors and antimicrobial agents. The EamB protein, which encodes an inner membrane transporter, acted as a negative regulator of biofilm formation, and the gene eamB deletion in the pathogen Aeromonas hydrophila LP-2 resulted in a significant increase in biofilm formation. Proteomic analysis revealed a total of 616 differentially abundant proteins between the ΔeamB and wild-type (WT) strains, with 308 downregulated and 308 upregulated. RT-qPCR was employed to verify the stability and accuracy of the proteomics data. Bioinformatic analysis indicated that EamB is involved in critical bacterial biological processes, including flagellar assembly, amino acid metabolism, and fatty acid degradation. Biofilm formation assays further revealed that supplementation with exogenous lysine significantly inhibited biofilm formation in the ΔeamB strain, conversely, exogenous cysteine and O-acetylserine obviously increased biofilm formation in the ΔeamB strain. These findings demonstrated that EamB may modulate bacterial biofilm formation in A. hydrophila through the regulation of amino acid metabolism. This finding provides novel insights into the regulatory mechanism underlying biofilm formation and highlights potential targets for the development of future antibacterial strategies. SIGNIFICANCE STATEMENT: This study elucidates the critical role of the eamB gene in Aeromonas hydrophila, a significant aquatic pathogen, by demonstrating its impact on biofilm formation and physiological traits. Through comparative proteomic analysis, we identified 616 differentially abundant proteins in the ΔeamB mutant, revealing its involvement in key metabolic pathways such as amino acid metabolism, flagellar assembly, and fatty acid degradation. Notably, eamB deletion enhanced biofilm formation, while exogenous amino acids like cysteine and O-acetylserine obviously increased biofilm formation in the ΔeamB strain. These findings highlight EamB as a regulator of biofilm formation, offering novel molecular insights into bacterial pathogenicity. This research advances our understanding of biofilm-associated antibiotic resistance and provides potential targets for developing strategies to mitigate infections caused by A. hydrophila in aquaculture and public health.
Proteomics analysis reveal early event molecular effectors of anti-CD19 CAR-T cell therapy in hematological cancer.
Teibo JO, Silveira RM, Silvestrini VC … +13 more , Archiolli I, Masson AP, de Morais BP, Schmidt D, Dos Santos MH, Ferreira GA, Thomé CH, Helm D, Nirujogi RS, Alessi DR, Picanço-Castro V, de Souza LEB, Faça VM
J Proteomics · 2025 Oct · PMID 40716489 · Publisher ↗
Chimeric antigen receptor T-cell (CAR-T) therapy is at the forefront of the field of cell immunotherapy. In this study, we generated an anti-CD19 CAR-Jurkat T cell line using a locally produced second-generation anti-CD1...
Chimeric antigen receptor T-cell (CAR-T) therapy is at the forefront of the field of cell immunotherapy. In this study, we generated an anti-CD19 CAR-Jurkat T cell line using a locally produced second-generation anti-CD19 CAR construct, which allowed us to analyse early proteomic changes that are crucial for comprehending the signalling pathways and mechanism of action of this CAR-T cell. SILAC-heavy tagged Raji B-cells and anti-CD19 CAR-Jurkat T-cells were co-cultured for ten minutes. The proteomic profiles were acquired via DIA methodology on the Orbitrap Astral LC-MS/MS platform. The proteome was extensively covered, resulting in about 8800 protein identifications at 1 % FDR. The effector CAR-Jurkat cells showed proteomic changes involving antigen presentation by CD74. The target Raji B-cells exhibited more significant alterations. Effector proteins, namely CD247, CD28, DAP, LCK, p38 MAPK, and CASP3, were validated, as they have critical roles in antigen presentation, T-cell activation, and apoptosis. Pharmacological inhibition of LCK using Dasatinib further suggested its pivotal role in early CAR-T signalling. This study led us to identify proteins that function as molecular effectors of anti-CD19 CAR-T cell therapy during the initial phases of CAR-T-target cell engagement, advancing our knowledge of the mechanism and signalling pathways that will support CAR-T cell development. SIGNIFICANCE: Chimeric antigen receptor T-cell (CAR-T cell) therapy is state-of-the-art in cell and immunotherapy. Determining important players in cellular communication and signalling mediated by membranes and intracellular proteins requires understanding the connection between tumours and modified cells. We employed global proteomics in this study to better grasp the functional protein networks using a high-sensitivity mass spectrometric platform for protein identification and quantification. We identified proteins as molecular effectors of anti-CD19 CAR-T cell treatment during the early stages of CAR-T-target cell interaction. Our understanding of the mechanism and signalling pathways will promote the development of new CAR constructs and improve the efficacy and ability to overcome the resistance of this innovative cancer treatment strategy, which will advance the identification of adjuvant molecules for the regulation of CAR-T responses.
Quantitative proteomic analysis of the Toxoplasma gondii cytoskeleton and bioinformatic identification of highly antigenic proteins.
Araujo-Ruiz K, López-Flores DI, Martínez-Muné MK … +7 more , García-Sánchez BY, Ramírez-Flores CJ, Sandoval-Rodríguez FE, Ríos-Castro E, Mondragón-Castelán ME, González-Pozos S, Mondragón-Flores R
J Proteomics · 2025 Oct · PMID 40716488 · Publisher ↗
The Toxoplasma gondii cytoskeleton is a highly organized structure essential for parasite motility, replication, and host cell invasion. To identify its components, a highly enriched fraction of tachyzoite cytoskeletons...
The Toxoplasma gondii cytoskeleton is a highly organized structure essential for parasite motility, replication, and host cell invasion. To identify its components, a highly enriched fraction of tachyzoite cytoskeletons was obtained and quantitatively analyzed by mass spectrometry. We identified 623 proteins classified into 18 functional groups, including 30 IMC proteins, 34 cytoskeleton proteins, and 14 uncharacterized proteins. A comprehensive bioinformatic analysis was conducted to assess protein abundance (fmol), antigenicity, accessibility, interactome, and homology, with the aim of identifying immunogenic targets. Among the top vaccine candidates were -GRA12, IMC1, ROP8, and -IMC4, with ROP8 emerging as the most promising based on epitope prediction. Data are available via ProteomeXchange with identifier PXD063409. SIGNIFICANCE: Toxoplasma gondii represents one of the most virulent and successful parasites in human and veterinary pathogenesis. Since T. gondii is a highly dynamic parasite that depends on its cytoskeleton to invade and disseminate through tissues, knowledge of its cytoskeleton composition is essential for understanding the biological mechanisms involved in parasite-host interactions and for the design of pharmaceutical and vaccination strategies. Quantitative proteomic analysis of the T. gondii cytoskeleton provided new and extensive information on its composition and, through bioinformatics approaches, allowed us to suggest several candidate molecules for future immunoprotective design.
E cadherin appears to be an essential on/off switch for initiating bile canaliculi formation.
Chevallet M, Rabilloud T, Diemer H … +3 more , Bertile F, Fuchs A, Deniaud A
J Proteomics · 2025 Oct · PMID 40714199 · Publisher ↗
The mechanisms underlying cell polarization are fundamental to biology but remain incompletely understood. This is especially true for hepatocytes, which display a particularly complex polarization that enables the forma...
The mechanisms underlying cell polarization are fundamental to biology but remain incompletely understood. This is especially true for hepatocytes, which display a particularly complex polarization that enables the formation of the bile canaliculi (BC) network crucial for liver excretory functions. To identify key proteins involved in hepatocyte polarization, BC formation, structure or function, we employed a proteomic approach comparing the human hepatocyte cell line HepG2 to its sub clone HepG2/C3A known for its markedly greater efficiency in forming mature BCs. Through this analysis, we localized LimA1 and Espin to the BC for the first time, suggesting their important role in this compartment, and confirmed the presence of NHE-RF1. Using a targeted protein repression strategy, we identified E cadherin as essential for the initiation of BC formation, unlike other adherens junction components such as N cadherin or α-catenin. Our findings demonstrate, for the first time, that in the absence of E cadherin, hepatocytes lose the capacity to form BCs. SIGNIFICANCE: This study aims to deepen our understanding of the highly specialized polarization of hepatocytes in relation to bile canaliculus formation. The major finding is the key role of E cadherin in this process, where it appears to be essential for bile canaliculus formation in both 2D and 3D culture models. Additionally, the study led to the identification of several proteins potentially localized to the bile canaliculi, whose functions remain to be elucidated.
Redox proteomics workflow to unveil extracellular targets of oxidation in vascular endothelial cells.
Vileigas DF, da Silva RP, Dempsey B … +3 more , Massafera MP, Pinz MP, Meotti FC
J Proteomics · 2025 Oct · PMID 40712755 · Publisher ↗
Redox regulation has emerged as a key process in cellular signaling. The role of extracellular cell surface redox-sensitive proteins in redox regulation and intracellular communication has been supported by secretion of...
Redox regulation has emerged as a key process in cellular signaling. The role of extracellular cell surface redox-sensitive proteins in redox regulation and intracellular communication has been supported by secretion of oxidoreductases that modulate thiol-disulfide switches. Despite these advances, redox-sensitive targets on the cell surface remain little explored. We established a comprehensive redox proteomic workflow using plasma membrane impermeable thiol labeling where we identified 1159 cell surface and extracellular proteins susceptible to oxidation. Treatment with diamide or urate hydroperoxide (HOOU) resulted in 377 and 12 differentially abundant redox-modulated proteins compared to control. Such proteins represent chaperones, adhesion molecules, vesicle-associated proteins, channels, receptors, cytoskeleton, and others, which may play a relevant role in several signaling pathway. Eleven oxidoreductases were redox-modulated by diamide, including members of the protein disulfide isomerase (PDI), peroxiredoxin (PRDX), and quiescin sulfhydryl oxidase (QSOX) families, with a particular focus on PDI TMX3 (TMX3), which provides the first evidence of its secretion in endothelial cells. In conclusion, our findings not only revealed potential redox-sensitive targets on the cell surface but also offer a useful tool for future investigations aiming to analyze redox regulation in the extracellular environment across diverse biological contexts. SIGNIFICANCE: Redox signaling at the cell surface is emerging as a crucial regulator of vascular function, emphasizing its role in cardiovascular disease. However, the extracellular redox proteome remains underexplored because of the complexity of the method. We developed a reproducible workflow combining differential thiol labeling and mass spectrometry to systematically map oxidized extracellular proteins in endothelial cells exposed to oxidants. Hundreds of proteins were identified as redox-sensitive targets. Key functional groups included molecular chaperones, adhesion molecules, vesicle-associated proteins, channels, receptors, and cytoskeleton. This work reveals novel insights into extracellular redox regulation, expands the repertoire of known redox-sensitive proteins, and establishes a versatile platform to investigate redox dynamics at cell surface both in vascular biology and other pathophysiological contexts.
Comparative proteomic analysis of subcutaneous and intraperitoneal cysts of Echinococcus multilocularis.
Zhang J, Cao S, Tian Z … +6 more , Zhang Z, Zhou Y, Wu Y, Luo X, Wang S, Guo X
J Proteomics · 2025 Sep · PMID 40701200 · Publisher ↗
Alveolar echinococcosis is a zoonotic disease that poses serious threats to public health. We observed subcutaneous cysts (SCs) of E. multilocularis had fewer protoscoleces (PSCs) compared to intraperitoneal cysts (ICs)...
Alveolar echinococcosis is a zoonotic disease that poses serious threats to public health. We observed subcutaneous cysts (SCs) of E. multilocularis had fewer protoscoleces (PSCs) compared to intraperitoneal cysts (ICs) at 60 days post-infection. However, the mechanisms underlying the development of E. multilocularis cysts in different tissues remain unclear. In this study, we compared the proteomic profiles of E. multilocularis cysts derived from mice intraperitoneally and subcutaneously infected with PSCs at 30 days post-infection, prior to the development of mature PSCs. Proteomic analysis identified 284 differentially expressed proteins (DEPs) in SCs compared to ICs, with 147 upregulated DEPs and 137 downregulated DEPs. Enzymatic proteins involved in carbohydrate and amino acid metabolism were predominantly upregulated in SCs compared to ICs, whereas proteins associated with protein folding, sorting, a degradation were downregulated. Western blotting analysis confirmed that phosphoenolpyruvate carboxykinase (PEPCK) and fructose-bisphosphate aldolase (FBA) were upregulated, whereas transitional endoplasmic reticulum ATPase (TER ATPase) was downregulated in SCs compared to ICs. The identified DEPs may play crucial roles in shaping the unique characteristics of E. multilocularis cysts. This study offers valuable insights into exploring the mechanisms underlying the occurrence and development of metacestodes.
Palaeoproteomic characterization of archaeological dental calculus reveals precarious periodontal health in pre-Roman Italy (7th-4th century BCE).
Riccomi G, Rosselli L, Marchesi M … +5 more , Guidi F, Taloni M, Ricci G, Pellegrino C, Wilkin S
J Proteomics · 2025 Sep · PMID 40691993 · Publisher ↗
Periodontitis, a chronic inflammatory disease affecting the tooth-supporting structures, is a key indicator of oral health in palaeopathology. While poor oral hygiene, systemic diseases, and genetics are well-established...
Periodontitis, a chronic inflammatory disease affecting the tooth-supporting structures, is a key indicator of oral health in palaeopathology. While poor oral hygiene, systemic diseases, and genetics are well-established contributors, the dietary impact has often been underestimated. Clinical studies, however, link diets high in fermentable carbohydrates and meat to inflammation. We investigated periodontal disease by analyzing interdental septa in 63 individuals from elite and non-elite groups in pre-Roman Italy (7th-4th centuries BCE), a period of social stratification, intensified agriculture, and increased cereal consumption. Macroscopic analysis was combined with proteomics of dental calculus from 33 individuals. Of the 1890 septa considered, 23 % displayed signs of periodontitis, with significantly higher rates in males. Prevalence increased with age in both sexes. Proteomic findings identified Porphyromonas gingivalis, a key periodontal pathogen, in 10 of 19 well-preserved dental calculus samples. While plaque accumulation is the main trigger for periodontitis, our findings highlight the dietary role in disease susceptibility. Carbohydrate-rich foods adhere to teeth and nourish bacteria, worsening periodontal conditions. At the same time, greater access to animal protein, particularly among emerging elites, may have contributed to inflammation. We propose that a proinflammatory diet may have been a major contributor to the proliferation of pathogenic oral microbiota.
About
- Frequency
- Sun
- Papers found
- 200
- RSS feed
- Subscribe