Perfluorooctanesulfonic acid (PFOS) elicits adverse effects on numerous organs and developmental processes but the mechanisms underlying these effects are not well understood. Here, we use single-cell RNA-sequencing to assess tissue-specific transcriptomic changes in zebrafish (Danio rerio) larvae exposed to 16 µM PFOS or dimethylsulfoxide (0.01 %) from 3-72 h post fertilization (hpf). Data analysis was multi-pronged and included pseudo-bulk, untargeted clustering, informed pathway queries, and a cluster curated for hepatocyte biomarkers (fabp10a, and apoa2). Overall, 8.63 % (2390/27698) genes were significantly differentially expressed. Results from untargeted analysis revealed 22 distinct clusters that were manually annotated to specific tissues using a weight-of-evidence approach. The clusters with the highest number of significant differentially expressed genes (DEGs) were digestive organs, muscle, and otolith. Additionally, we assessed the distribution of pathway-specific genes known to be involved in PFOS toxicity: the PPAR pathway, β-oxidation of fatty acids, the Nfe2l2 pathway, and epigenetic modifications by DNA methylation, across clusters and identified the blood-related tissue to be the most sensitive. The curated hepatocyte cluster showed 220 significant DEGs and was enriched for the Notch signaling pathway. These findings provide insights into both established and novel sensitive target tissues and molecular mechanisms of developmental toxicity of PFOS.

The characterization of gene sets is a recurring task in computational biology. Identifying specific properties of a hit set compared to a reference set can reveal biological roles and mechanisms, and can lead to the prediction of new hits. However, collecting the features to evaluate can be time consuming, and implementing an informative but compact graphical representation of the multiple comparisons can be challenging, particularly for bench scientists. Here, I present Qarles (quick characterization of large sets of genes), a web server that annotates Saccharomyces cerevisiae gene sets by querying a database of 31 features widely used by the yeast community and that identifies their specific properties, providing publication-ready figures and reliable statistics. Qarles has a deliberately simple user interface with all the functionality in a single web page and a fast response time to facilitate adoption by the scientific community. Qarles provides a rich and compact graphical output, including up to five gene set comparisons across 31 features in a single dotplot, and interactive boxplots to enable the identification of outliers. Qarles can also predict new hit genes by using a random forest trained on the selected features. The web server is freely available at https://qarles.org.

Type 2 diabetes mellitus (T2DM) and Alzheimer's disease (AD) have similar clinical characteristics in the brain and islet, as well as an increased incidence with ageing and familial susceptibility. Therefore, in recent years there has been a great desire for research that elucidates how anti-diabetic drugs affect AD. This work attempts to first elucidate the possible mechanism of action of DPP-IV inhibitors in the treatment of AD by employing techniques from network pharmacology, molecular docking, molecular dynamic simulation, principal component analysis, and MM/PBSA. A total of 463 targets were identified from the SwissTargetPrediction and 784 targets were identified from the SuperPred databases. 79 common targets were screened using the PPI network. The GO and KEGG analyses indicated that the activity of DPP-IV against AD potentially involves the hsa04080 neuroactive ligand-receptor interaction signalling pathway, which contains 17 proteins, including CHRM2, CHRM3, CHRNB1, CHRNB4, CHRM1, PTGER2, CHRM4, CHRM5, TACR2, HTR2C, TACR1, F2, GABRG2, MC4R, HTR7, CHRNG, and DRD3. Molecular docking demonstrated that sitagliptin had the greatest binding affinity of -10.7 kcal/mol and established hydrogen bonds with the Asp103, Ser107, and Asn404 residues in the active site of the CHRM2 protein. Molecular dynamic simulation, PCA, and MM/PBSA were performed for the complex of sitagliptin with the above-mentioned proteins, which revealed a stable complex throughout the simulation. The work identifies the active component and possible molecular mechanism of sitagliptin in the treatment of AD and provides a theoretical foundation for future fundamental research and practical implementation.

N6-Methyladenosine (m6A) is a neuronal-enriched, reversible post-transcriptional modification that regulates RNA metabolism. The m6A-modified RNAs recruit various m6A-binding proteins that act as readers. Differential m6A methylation patterns are implicated in ischemic brain damage, yet the precise role of m6A readers in propagating post-stroke m6A signaling remains unclear. We presently evaluated the functional significance of the brain-enriched m6A reader YTHDF1, in post-stroke pathophysiology. Focal cerebral ischemia significantly increased YTHDF1 mRNA and protein expression in adult mice of both sexes. YTHDF1-/- male, but not female, mice subjected to transient middle cerebral artery occlusion (MCAO) showed worsened motor function recovery and increased infarction compared to sex-matched YTHDF1+/+ mice. YTHDF1-/- male, but not female, mice subjected to transient MCAO also showed significantly perturbed expression of genes related to inflammation, and increased infiltration of peripheral immune cells into the peri-infarct cortex, compared with sex-matched YTHDF1+/+ mice. Thus, this study demonstrates a sexual dimorphism of YTHDF1 in regulating post-ischemic inflammation and pathophysiology. Hence, post-stroke epitranscriptomic regulation might be sex-dependent.

The Gene Ontology (GO) project has been pivotal in providing a structured framework for characterizing genes and annotating them to specific biological concepts. While traditional gene annotation primarily focuses on mapping genes to GO terms, descriptors of biological concepts, there is a growing need for tools facilitating reverse querying. This paper introduces GOReverseLookup, a novel tool designed to identify over- or underrepresented genes in researcher-defined states of interest (phenotypes), described by sets of GO terms. GOReverseLookup supplements the existing power of Gene Ontology by the possibility of orthologous gene querying across several databases, such as Ensembl and UniProtKB. This combination allows for a more nuanced identification of significant genes across a range of cross-species research contexts.

GOReverseLookup queries genes associated with input GO terms. Bundles of GO terms encapsulate user-defined states of interest, e.g., angiogenesis. In the second stage of the analysis, all GO terms associated with each gene are fetched, and finally, the statistical relevance of the genes being involved in one (or all) of the defined states of interests is computed.

The two presented use cases illustrate its utility in discovering genes related to rheumatoid arthritis and genes linked with chronic inflammation and tumorigenesis. In both cases, GOReverseLookup discovered a substantial number of genes significantly associated with the aforementioned states of interest.

GOReverseLookup proves to be a valuable resource for unraveling the genetic basis of phenotypes, with diverse practical potentials in functional genomics, systems biology, and drug discovery. We anticipate that GOReverseLookup will significantly aid in identifying potential gene targets during the initial research phases.

Eukaryotes achieve a wide range of stable phenotypes by virtue of epigenetic modifications. However, what drives epigenetic diversification in the first place remains an open question. Here, we investigated the dynamic interplay between the production fluxes of epigenetic cosubstrates and histone post-translation modifications (PTMs) in Saccharomyces cerevisiae's Yeast Metabolic Cycle (YMC). We developed a novel approach integrating flux analysis with transcriptomic data to investigate the production fluxes of acetyl-CoA and SAM and their influence on histone marks H3K9Ac and H3K4me3. Our results show that acetyl-CoA and SAM flux dynamics are asynchronous during the YMC, suggesting distinct regulatory roles. Gene ontology analysis revealed that genes whose enrichment of H3K9Ac correlates with acetyl-CoA dynamics are associated with metabolic functions, while genes whose enrichment of H3K4me3 correlates with SAM dynamics are associated with translation processes. Finally, we found evidence that chromatin accessibility on genes promoter regions was a precondition for the metabolic fluxes influencing the enrichment of H3K4me3 and H3K9Ac. These findings support the concept that metabolism provides timely cosubstrates essential for histone PTMs.

Evidence has shown that ibuprofen and celecoxib are effective in improving depressive symptoms, but their mechanisms of action are unclear. In this study, we aimed to determine the relationship between these two drugs and depressive disorder (DD) and elucidate potential mechanisms of action.

Relevant targets for ibuprofen, celecoxib, and DD were obtained and screened from multiple online drug and disease public databases. A protein-protein interaction network was obtained. The Centiscape and CytoHubba plug-ins were applied to screen for core targets. Gene Ontology and Kyoto Encyclopedia of Genes and Genomes analyses were performed. Molecular docking was performed to predict the binding of ibuprofen and celecoxib to core targets. Examined the differences in core target protein expression between DD patients (DDs, n = 18) and healthy controls (HCs, n = 16) as a further experimental validation of the network pharmacology results.

In total, 220 potential targets for ibuprofen and 316 potential targets for celecoxib were identified and associated with DD. The antidepressant effects of both drugs involve many key targets in pathways such as "pathways in cancer" and "neuroactive ligand-receptor interaction," including ALB, BCL2, MAPK3, SRC, STAT3, EGFR, and PPARG. The binding affinity of ALB with ibuprofen is the strongest, and it is connected only by hydrophobic interactions. Celecoxib exhibits higher affinity at multiple targets such as SRC, EGFR, and PPARG, with stronger and more specific intermolecular interactions, including salt bridges and halogen bonds. Clinical trials have found that serum ALB expression in DDs is significantly lower than that in HCs (t = 6.653, p < 0.001), further confirming the potential role of ibuprofen in DD.

Ibuprofen and celecoxib primarily exert their antidepressant effects through targets and pathways related to inflammation, neural signaling, and cancer, with celecoxib showing a stronger potential antidepressant effect. The expression difference of the core target ALB between depression and healthy individuals further supports the potential effect of the drug on DD. Our findings propose new treatment strategies, support the link between inflammation and depression, and encourage reassessing existing medications for depression.

Thrombotic diseases remain the major cause of death and disability worldwide, and the contribution of inflammation is increasingly recognized. Thromboinflammation has been identified as a key pathomechanism, but an unsupervised map of immune-cell states, trajectories, and intercommunication at a single-cell level has been lacking. Here, we reveal innate leukocyte substates with prominent thrombolytic properties by employing single-cell omics measures on human stroke thrombi. Using in vivo and in vitro thrombosis models, we propose a pro-resolving monocyte-neutrophil axis, combining two properties: (1) NR4A1hi non-classical monocytes acquire a thrombolytic and neutrophil-chemoattractive phenotype, and (2) blood neutrophils are thereby continuously recruited to established thrombi through CXCL8-CXCR1 and CXCR2 and adopt a hypoxia-induced thrombus-resolving urokinase receptor (PLAUR)+ phenotype. This immunothrombolytic axis results in thrombus resolution. Together, with this immune landscape of thrombosis, we provide a valuable resource and introduce the concept of "immunothrombolysis" with broad mechanistic and translational implications at the crossroad of inflammation and thrombosis.

Environmental and endogenous stimuli determine plant developmental transitions including flowering through multiple signaling cascades. Although the key activators and repressors of flowering initiation are defined, the components and mechanisms integrating light signaling and flowering pathways are not fully established. This study investigates the role of TANDEM ZINC-FINGER/PLUS3 (TZP), a light-integrating transcriptional regulator, to elucidate how light cues influence the epigenetic regulation of flowering in Arabidopsis thaliana. To dissect the molecular function of TZP, this study employed a combination of genetics, RNA sequencing, chromatin immunoprecipitation sequencing and phenotypic assays. These approaches were used to determine TZP's genomic binding sites, its downstream gene targets and its influence on flowering time and chromatin modifications. TANDEM ZINC-FINGER/PLUS3 was found to directly associate with the promoter regions of chromatin-modifying genes, including FLOWERING LOCUS D (a histone H3K4 demethylase) and histone deacetylase 6 (a histone deacetylase). This regulation promotes a chromatin environment that represses FLOWERING LOCUS C (FLC) transcription, thereby accelerating flowering. TANDEM ZINC-FINGER/PLUS3 thus functions upstream of multiple pathways integrating photoperiodic and autonomous floral cues. TANDEM ZINC-FINGER/PLUS3 mediates crosstalk between light signaling and flowering pathways by modulating chromatin structure at the FLC locus. This provides a mechanistic framework for understanding how environmental signals dynamically influence epigenetic regulation of developmental transitions.

Mycobacterium tuberculosis poses a serious challenge for human health, and new antibiotics with novel targets are needed. Here we demonstrate that an acylaminooxadiazole, MBX-4132, specifically inhibits the trans-translation ribosome rescue pathway to kill M. tuberculosis. Our data demonstrate that MBX-4132 is bactericidal against multiple pathogenic mycobacterial species and kills M. tuberculosis in macrophages. We also show that acylaminooxadiazole activity is antagonized by iron but is potentiated by zinc. Our transcriptomic data reveal dysregulation of multiple metal homeostasis pathways after exposure to MBX-4132. Furthermore, we see differential expression of genes related to zinc sensing and efflux when trans-translation is inhibited. Taken together, these data suggest that there is a link between disturbing intracellular metal levels and acylaminooxadiazole-mediated inhibition of trans-translation. These findings provide an important proof-of-concept that trans-translation is a promising antitubercular drug target.

The genomic landscape of esophageal squamous cell carcinoma (ESCC) has been characterized extensively, but there remains a significant need for actionable targets and effective therapies.

Here, we perform integrative analysis of genome-wide loss of heterozygosity and expression to identify potential tumor suppressor genes. The functions and mechanisms of one of the candidates, TACC2, are then explored both in vitro and in vivo, leading to the proposal of a therapeutic strategy based on the concept of synthetic lethality.

We reveal that the inactivation of TACC2, due to copy number loss and promoter hypermethylation, is associated with poor prognosis in ESCC patients. TACC2 depletion enhances ESCC tumorigenesis and progression, as demonstrated in Tacc2 knockout mouse models and by increased growth abilities of ESCC cells. Mechanistically, TACC2 interacts with components of the NuRD and CoREST co-repressor complexes, including MTA1, MBD3, and HMG20B, in the cytoplasm. TACC2 loss leads to the translocation of these proteins into the nucleus, facilitating the formation of functional NuRD and CoREST complexes and the epigenetic repression of CDKN1A. This repression results in elevated CDK1/2 activation. Furthermore, TACC2-deficient cells and ESCC patient-derived organoids with reduced TACC2 expression show increased sensitivity to CDK inhibitors, particularly dinaciclib, which is currently in a phase III trial. Notably, the combination of TACC2-specific RNAi and dinaciclib in subcutaneous ESCC models significantly impairs tumor growth.

The findings suggest a strategy for cancer treatment based on synthetic lethality.

Funded by NKRDP, NSFC, GDIIET, GDBABRF, GDECISTP, and SYSUTP.

Colorectal cancer (CRC) is the third-most common cancer and the second-leading cause of mortality due to cancer worldwide. The underlying regulatory mechanism of CCNA2 in CRC was explored through multiomics and experimental analyses, thus facilitating diagnosis, therapy and prognosis. Two gene expression datasets (i.e., GSE9348 and GSE110223) were extracted from GEO. Differentially expressed genes (DEGs) were identified via GEO2R, which were used for enrichment analyses through DAVID. PPI network of DEGs was constructed by STRING, and the core genes were identified. CCNA2, a prognostic core gene for CRC, was validated in TCGA and HPA via transcriptomics and proteomics. ROC analysis was performed to evaluate the diagnostic value of CCNA2 in CRC. The therapeutic value of CCNA2 was evaluated in DGIdb through pharmacogenomics. The correlation between CCNA2 and immune infiltration was determined in TIMER by immunomics. TF-mRNA and miRNA-mRNA networks for CCNA2 were constructed in miRnet and miRDB via transcriptomics. The role and mechanism of CCNA2 in CRC were investigated both in vitro and in vivo. The miR-548x-3p/CCNA2 regulatory axis in CRC was investigated in vitro. CCNA2 showed excellent diagnostic, therapeutic, and prognostic value in CRC. CCNA2 was closely associated with tumor-infiltrating immunocytes, TFs, and miRNAs. The upregulation of CCNA2 was observed in CRC, and the knockdown of CCNA2 inhibited the proliferation, migration, and invasion while inducing apoptosis of CRC cells. The knockdown of CCNA2 could inhibit epithelial-mesenchymal transition (EMT) pathway. CCNA2 acted as a target of miR-548x-3p in regulating the biological behavior of CRC cells via the EMT-signaling pathway. CCNA2 is a potential biomarker for the diagnosis, treatment, and prognosis of CRC and is associated with immune infiltration, TF, and miRNA. The miR-548x-3p/CCNA2 axis plays a pivotal role in regulating the tumorigenesis of CRC through the EMT-signaling pathway.

PANoptosis is a concept of total cell death characterized by pyroptosis, apoptosis, and necroptosis. We aimed to explore the clinical significance of PANoptosis-related genes (PARGs) in glioblastoma multiforme (GBM).

Expression profiles of GBM were downloaded from the XENA database as a training dataset to construct a differentially expressed PARGs (DE-PARGs)-based risk score (RS) model, and the prognostic prediction role was validated in the CGGA database and GSE108474 using Kaplan-Meier (KM) curve and receiver operating characteristic (ROC) curve. Meanwhile, independent prognostic clinical factors were screened, and their prognosis predictive activity was evaluated by a nomogram model. Furthermore, the relationships between key DE-PARGs and immune cell infiltration, as well as chemotherapy drug sensitivity were analyzed.

The RS model consisting of five DE-PARGs was constructed, including NOD2, NLRP2, NLRP7, GATA3, and TERT. ROC and KM curves confirmed the good potency of the RS prognostic model both in XENA database and GSE108474. Three clinical prognostic factors, including chemotherapy, pharmaceutical therapy, and RS model, were selected as individual prognostic factors. The nomogram model showed RS contributed most to survival probability, followed by chemotherapy and pharmaceutical therapy. In high- and low-risk groups, B cell memory, NK cell resting, and macrophage M1 had significant differences. As compared with the immune checkpoint therapy non-responder group, the responder involved a higher ratio of patients sub-grouped into the low-risk group. Three drugs between high- and low-risk groups had significant differences, including Cisplatin, Gefitinib, and Vorinostat.

Our data exhibit the prognostic value of PARGs in GBM and offer new insights for GBM pathogenesis and immune treatment.

A better understanding of the mechanisms regulating CD8+ T cell differentiation is essential to develop new strategies to fight infections and cancer. Using genetic mouse models and blocking antibodies, we uncovered cellular and molecular mechanisms by which Notch signaling favors the efficient generation of effector CD8+ T cells. Fibroblastic reticular cells from secondary lymphoid organs, but not dendritic cells, were the dominant source of Notch signals in T cells via Delta-like1/4 ligands within the first 3 days of immune responses to vaccination or infection. Using transcriptional and epigenetic studies, we identified a unique Notch-driven T cell-specific signature. Early Notch signals were associated with chromatin opening in regions occupied by bZIP transcription factors, specifically BATF, known to be important for CD8+ T cell differentiation. Overall, we show that fibroblastic reticular cell niches control the ultimate molecular and functional fate of CD8+ T cells after vaccination or infection through the delivery of early Notch signals.

Bovine alphaherpesvirus one (BoHV-1) is a primary cause of bovine respiratory disease (BRD), and a leading cause of morbidity and mortality in cattle. The transcriptomic responses of key respiratory and immune associated tissues of dairy calves following experimental challenge with BoHV-1 are unknown. Thus, the study objective was to examine the gene expression profiles of multiple tissue types from dairy calves following an infectious challenge with BoHV-1. Holstein-Friesian bull calves (mean age ±  SD 149.2 days ±  23.8; mean weight ±  SD 174.6 kg ±  21.3 kg were challenged with either BoHV-1 inoculate (6.3 ×  107/mL ×  1.35mL) (n =  12) or sterile phosphate buffered saline (n =  6). Animals were euthanised on day 6 post-challenge and tissue samples collected, including bronchial (BLN) and mediastinal lymph nodes (MLN), pharyngeal tonsil (PGT) and healthy (HL) and lesioned right cranial lung (LL). Total RNA was extracted and libraries sequenced on an Illumina NovaSeq 6000. Differential expression analysis was conducted using edgeR and pathways analysed using DAVID. A weighted gene co-expression network analysis (WGCNA) was conducted separately for each tissue type to identify networks significantly associated with BoHV-1 infection. Differentially expressed genes (DEGs) were identified in all tissues (P <  0.05, FDR <  0.1, FC >  2). Thirty-three DEGs were common to all tissues and enriched pathways included Influenza A and Herpes simplex 1 infection (P < 0.05, FDR < 0.05). Modules enriched for antiviral and innate immune processes were identified for each tissue type. Of the 33 DEGs common to all tissues, 26 were also identified as hub genes in the blood (blue) module. Our use of a controlled experimental challenge allowed for improved understanding of the immune response of dairy calves to a BoHV-1 infection. Furthermore, discovering DEGs that are common to all tissues, including whole blood, indicates future focus areas in research surrounding BRD diagnostic biomarkers.

Toxicological information on moniliformin (MON), an emerging mycotoxin, is limited. This study examined the acute and 28-day toxicity of orally administered MON in male ICR mice. Regarding the acute toxicity, among single oral doses of 0, 20, 40, and 80 mg/kg body weight (BW), MON caused proximal tubular necrosis in the kidneys at ≥ 40 mg/kg BW, and the lethal dose 50 value was estimated as 68.1 mg/kg BW. Regarding the 28-day toxicity, among oral doses of 0, 10, 20, and 40 mg/kg BW/day, MON increased absolute heart weight at 40 mg/kg BW, but histopathological changes were not evident in the heart. In contrast, 40 mg/kg BW MON induced centrilobular liver cell hypertrophy accompanied by increased absolute liver weight. Moreover, MON dose-dependently increased the absolute kidney weight at ≥ 20 mg/kg BW and increased the incidence of renal tubular regeneration at 40 mg/kg BW. RNA sequencing analysis in the renal cortex after a single dose of 40 mg/kg BW MON revealed upregulation of metabolic response-related genes, such as Cyp3a13, Cyp26b1, and Cyp4f15, and oxidative stress-related Gpx7. These results suggest that MON targets the kidneys in mice. Orally ingested MON may be metabolized in the kidneys as well as in the liver, and active intermediates or reactive oxygen species may induce renal tubular toxicity, causing proximal tubular necrosis. Based on kidney changes, the no-observed-adverse-effect-level of MON in the 28-day oral toxicity study of male mice was determined to be 10 mg/kg BW/day.

Elucidating the molecular regulatory mechanisms underlying muscle growth and development is of profound significance in aquaculture. Yesso scallop is a cold-water bivalve of considerable economic importance, having its primary edible component of adductor muscle. In this study, comparative transcriptomics and histological analysis at different sampling times after Myostatin (MSTN) interference were performed to identify the potential candidate genes potentially involved in muscle growth and development. The comparative transcriptomics revealed that growth factors and cytokines, extracellular matrix proteins and ubiquitin-proteasome system are potentially involved in muscle hypertrophy and hyperplasia. After MSTN interference, striated adductor muscle displays significant muscle hypertrophy (51.77 % increase on day 7 and 59.83 % increase on day 21) and muscle hyperplasia (59.36 % increase on day 7 and 61.83 % increase on day 21). WGCNA identifies the key darkolivegreen module, which may play crucial roles in muscle hyperplasia and hypertrophy within the striated muscle of the scallop. Five key transcription factors (zf-CCCH, zf-C2H2, PPP1R10, LRRFIP2, and Gon4) are identified by analyzing the co-expression patterns of core genes within the module. These findings will aid in understanding the regulatory mechanisms of muscle growth in scallops and provide a basis for genetic improvement in shellfish aquaculture.

Nitrogen (N) availability is a key factor in plant growth, but the molecular mechanisms underlying the early responses of poplar (Populus × xiaohei T. S. Hwang & Liang) roots to nitrogen are not well understood. The primary objective of this study was to elucidate these early molecular responses by integrating transcriptome, proteome, and phosphoproteome under low-nitrogen (LN, 0.2 mM NH4NO3) and high-nitrogen (HN, 2 mM NH4NO3) conditions. Specifically, the objectives of this study were: (i) to identify key metabolic pathways involved in nitrogen responses in poplar roots; (ii) to explore the relationship between differentially expressed genes (DEGs) and transcription factors (TFs) within these pathways; and (iii) to construct co-expression networks to uncover the regulatory mechanisms of nitrogen signaling. KEGG pathway enrichment analysis indicated that nitrogen metabolism and phenylpropanoid metabolism were key pathways in RNA-seq and proteome, while starch and sucrose metabolism were crucial in transcriptome and phosphoproteome. Plant hormone signal transduction was a key pathway in transcriptome, and gluconeogenesis/glycolysis was essential in proteome. WGCNA revealed three key modules (MEgreenyellow, MEblack, and MEblue) significantly associated with physiological indices, including NO3-, soluble sugar, and sucrose contents. Co-expression networks highlighted TFs as central regulators of nitrogen-responsive pathways, with distinct expression patterns between LN and HN treatments. These findings elucidate the complexity of nitrogen-regulated metabolic networks in poplar roots and reveal potential links between nitrogen signaling, carbohydrate metabolism, and secondary metabolism. This study provides a foundation for improving nitrogen-use efficiency in forest trees, with implications for sustainable forestry and ecosystem management.

Glucocorticoids are a mainstay in the treatment of B-cell acute lymphoblastic leukemia (B-ALL). The glucocorticoid receptor (GR), a ligand-activated transcription factor (TF), mediates their actions. Chromatin occupancy, chromatin-protein networks (chromatomes) and gene programmes of GR are regulated by SUMOylation, a post-translational modification with therapeutic implications in other hematomalignancies. To unravel the GR-SUMOylation crosstalk in B-ALL, we induced hypoSUMOylation in NALM6 B-ALL cells with a SUMOylation inhibitor (SUMOi, ML-792). Genome-wide profiling of GR and SUMO chromatin-binding and chromatin accessibility revealed that hypoSUMOylation augmented GR chromatin occupancy and altered chromatin openness. Association with transcriptome data indicated that the hypoSUMOylation-induced GR-binding sites predominantly repressed genes associated with cell cycle and DNA replication. Consistently, hypoSUMOylation potentiated glucocorticoid-induced cell cycle arrest and growth suppression. Moreover, our proteomic analyses revealed that the protein network of chromatin-bound GR is tightly intertwined with SUMO2/3 and that SUMOylation modulates the stability of the network. The chromatome contained several B-cell TFs with cognate binding motifs found on GR-adjacent chromatin sites, indicating their simultaneous occupancy on chromatin. In sum, our data imply potential for targeting SUMOylation to increase sensitivity to glucocorticoids in B-ALL, supported by ex vivo data of glucocorticoid and SUMOi TAK-981 combination-treated B-ALL patient samples.

Wheat (Triticum aestivum) is a vital cereal crop and a staple food source worldwide. However, wheat grain productivity has significantly declined as a consequence of infestations by Phalaris minor. Traditional weed control methods have proven inadequate owing to the physiological similarities between P. minor and wheat during early growth stages. Consequently, farmers have turned to herbicides, targeting acetyl-CoA carboxylase (ACCase), acetolactate synthase (ALS) and photosystem II (PSII). Isoproturon targeting PSII was introduced in mid-1970s, to manage P. minor infestations. Despite their effectiveness, the repetitive use of these herbicides has led to the development of herbicide-resistant P. minor biotypes, posing a significant challenge to wheat productivity. To address this issue, there is a pressing need for innovative weed management strategies and the discovery of novel herbicide molecules. The integration of computer-aided drug discovery (CADD) techniques has emerged as a promising approach in herbicide research, that facilitates the identification of herbicide targets and enables the screening of large chemical libraries for potential herbicide-like molecules. By employing techniques such as homology modelling, molecular docking, molecular dynamics simulation and pharmacophore modelling, CADD has become a rapid and cost-effective medium to accelerate the herbicide discovery process significantly. This approach not only reduces the dependency on traditional experimental methods, but also enhances the precision and efficacy of herbicide development. This article underscores the critical role of bioinformatics and CADD in developing next-generation herbicides, offering new hope for sustainable weed management and improved wheat cultivation practices. © 2024 Society of Chemical Industry.

Pancreatic ductal adenocarcinoma (PDAC) is a disease with poor prognosis due to diagnostic and therapeutic limitations. We previously identified cystatin A (CSTA) as a PDAC biomarker and have conducted the present study to investigate the antitumor effects of CSTA. PDAC murine models were established with genetically modified PAN02 tumor cell lines to evaluate the antitumor immune response. PDAC mouse survival was significantly longer with CSTA, and its antitumor effect was mediated mainly by CD4+ cells and partly by CD8+ cells. We also observed an increased infiltration of CD4+ and CD8+ cells in tumors of mice overexpressing CSTA. Phenotypically, we confirmed higher T helper type 1 (Th1) cell activity and increased frequency and activity of M1 macrophages and dendritic cells (DCs) in CSTA-overexpressing mice. Gene expression analysis highlighted pathways related to interferon gamma (IFN-γ) induction and Th1 lymphocyte activation that were induced by CSTA. Macrophages and DCs shifted toward proinflammatory antitumor phenotypes. Furthermore, activated splenocytes of PDAC model mice expressing CSTA had increased proapoptotic activity. CSTA also promoted the selective migration of CD4+ and CD11c+ immune cells in an in vitro migration assay. In conclusion, CSTA exerts antitumor effects by enhancing Th1-mediated antitumor effects through promotion of DC and M1 macrophage activity, thereby increasing immune cell chemotaxis. CSTA could be a novel therapeutic candidate for PDAC.

Type 2 diabetes mellitus (T2DM) is characterized by pancreatic β-cell dysfunction and insulin resistance, with pyroptosis emerging as a key contributor to β-cell loss. Jiedu Tongluo Tiaogan Formula (JTTF), a traditional Chinese medicine (TCM), has shown clinical efficacy in T2DM management, but its mechanism linking pyroptosis remains unexplored. This study integrates UPLC-MS/MS, network pharmacology, and in vivo experiments to elucidate JTTF's anti-diabetic mechanisms. UPLC-MS/MS identified 441 compounds in JTTF, predominantly alkaloids, flavonoids, phenols, and terpenoids. Network pharmacology revealed JTTF's multi-target effects on T2DM-associated pyroptosis, particularly via the NLRP3/Caspase-1/GSDMD pathway. In diabetic mice, JTTF dose-dependently reduced fasting blood glucose, insulin resistance, and dyslipidemia, while restoring pancreatic β-cell morphology. Mechanistically, JTTF suppressed NLRP3 inflammasome activation, downregulated Caspase-1 and GSDMD expression, and attenuated IL-1β/IL-18 release. Notably, this study provides the first evidence of JTTF's anti-pyroptotic effects in T2DM, highlighting its unique ability to modulate glycolipid metabolism and inflammatory cell death concurrently. These findings underscore JTTF's translational promise for preserving β-cell function and suggest future exploration of non-classical pyroptosis pathways. Our work bridges traditional medicine and molecular pharmacology, paving the way for clinical trials and integrative T2DM therapies.

Increased expression of the cyclin-dependent kinase inhibitor p16Ink4a (p16) is detected in neurons of human Alzheimer's disease (AD) brains and during normal aging. Importantly, selective eliminating p16-expressing cells in AD mouse models attenuates tau pathologies and improves cognition. But whether and how p16 contributes to AD pathogenesis remains unclear. To address this question, we tested whether induction of p16 expression in neurons exacerbates AD pathologies. We created a doxycycline-inducible system to trigger p16 up-regulation in human-induced pluripotent stem cells (iPSCs) and neurons differentiated from iPSCs. We demonstrated that up-regulated p16 expression in iPSCs reduces cell proliferation, down-regulates cell cycle genes, and up-regulates genes involved in focal adhesion, interferon α response and PI3K-Akt signaling. Our approach enables temporal control of p16 induction upon differentiation from iPSCs to neurons. In differentiated cortical neurons, we found that up-regulation of p16 increases tau phosphorylation at Ser202/Thr205 and Thr231 in a cell-autonomous manner, while amyloid beta secretion is not affected. These data suggest a critical role of p16 in regulating tau phosphorylation in neurons, and thereby contributing to pathological progression of AD. As pathological tau tangles have been shown to induce p16 expression, our studies suggest a positive feedback loop between p16 and tau to exacerbate tau pathologies.

Although necroptosis is an emerging mechanism of multiple organ dysfunction in sepsis, data on the mechanistic link between necroptosis and sepsis are scarce. Bioinformatic analysis was performed to compare the gene profiles between the sepsis (n = 133) and healthy control (n = 12) groups and identify necroptosis-related differentially expressed genes (DEGs). The identified necroptosis-related DEGs were verified by three-step molecular experiments: (1) quantitative real-time PCR and enzyme-linked immunosorbent assay; (2) cell culture, transfection and Western blotting; and (3) cytokine array with apoptosis inhibition. Additionally, receiver-operating characteristic curve analyses were performed to evaluate the performance of the corresponding proteins to the necroptosis-related DEGs in diagnosing sepsis and in predicting in-hospital mortality of patients with sepsis. Eight necroptosis-related DEGs, including five upregulated (PYGL, TNF, CYLD, FADD and TLR3) and three downregulated (TP53, FASLG and NLRP6) DEGs, were identified. Moreover, the levels of the corresponding proteins to necroptosis-related DEGs showed excellent or considerable accuracy in diagnosing sepsis and in predicting the mortality of sepsis patients. In cell culture media transfected with plasma from the sepsis and control groups, Western blotting revealed that the levels of the corresponding proteins were increased in the upregulated DEGs and decreased in the downregulated DEGs. The cytokine array revealed cytokines in cell culture media transfected with plasma from patients with sepsis while preventing apoptosis by inhibiting the caspase-8 activity, wherein the transfected cells potentially underwent necroptosis. Eight necroptosis-related DEGs were identified in patients with sepsis by bioinformatic analysis and verified by molecular experiments, implying that necroptosis may be a key mechanism of sepsis.

Moyamoya disease (MMD) is a rare cerebrovascular disease in humans. Although early revascularization can improve symptoms, it cannot reverse the progression of the disease. The current diagnosis still relies on traditional a Digital Subtraction Angiography (DSA) examination, which is invasive and expensive, leading to delayed diagnosis and affecting treatment timing and patient prognosis. The ability to diagnose MMD early and develop personalized treatment plans can significantly improve the prognosis of patients. Here, we have introduced the research on MMD biomarkers. By integrating proteomics and metabolomics data, we have successfully identified over 1700 features from more than 60 serum samples collected at the onset of symptoms in MMD patients. We use multiple computational strategies to interpret complex information in serum, providing a comprehensive perspective for early diagnosis of MMD. Diagnostic ability of our biomarker is significantly better than previous studies, especially when used in combination. In the study of molecular mechanisms, we found that the ferroptosis pathway was significant disruption in MMD patients, which was also confirmed by transcriptomics data. Finally, we validated the metabolites and proteins associated with ferroptosis pathways, as well as the biomarkers screened by machine learning, using another independent MMD cohort. Our research provides important clues for the diagnosis of MMD, and this assay can identify MMD early, thereby promoting stronger monitoring and intervention.

In biological membranes, lipids interact with membrane proteins (MPs) and play important roles in allosterically regulating their structure and function. Analyzing lipid-MP interactions is necessary for understanding these regulatory mechanisms; however, there have been few comprehensive and systematic studies to date. To address this, we developed a high-sensitivity, high-throughput platform that integrates lipid-immobilized beads with advanced proteomics to analyze lipid-MP interactions in detail. We prepared six types of lipid-immobilized beads, including sphingomyelin (SM), ceramide (Cer), dihydrosphingomyelin (DHSM), dihydroceramide (DHCer), phosphatidylcholine (PC), and cholesterol (Chol). In addition, we introduced a novel type of beads that immobilized SM and Chol (SM/Chol beads) to mimic lipid rafts. We first demonstrated that SM/Chol beads coprecipitated with Nakanori, a protein that specifically recognizes and binds to SM/Chol complexes, whereas beads immobilized with SM or Chol alone did not coprecipitate. This indicates the effectiveness of SM/Chol beads for the identification of raft-associated proteins. Next, the cell lysates were incubated with the seven types of lipid-immobilized beads and the recovered proteins were analyzed using shotgun proteomics. This approach successfully identified over 7000 lipid-binding proteins. Filtering based on fold-change values and subsequent enrichment analysis revealed distinct binding protein profiles for each lipid, highlighting the functional diversity of lipid-MP interactions and their roles in cellular processes. In summary, our methodology enables an unprecedented large-scale exploration of lipid-MP interactions. This platform provides a versatile tool for examining the lipid-mediated regulation of MPs and offers new insights into the physiological significance of the lipidome and its implications for health and disease.

Crataegus azarolus L. (Rosaceae), commonly known as Mediterranean hawthorn, has long been valued in Traditional Medicine for treating cardiovascular and inflammation-related diseases, including diabetes, cancer, and rheumatism. Pharmacological benefits of Crataegus azarolus L. are notably linked to its anti-inflammatory properties. Fupenzic acid, a pentacyclic triterpene isolated from its leaves, holds significant pharmacological potential that remains elusive. This study investigates the unexplored capacity of fupenzic acid as a promising anti-inflammatory agent. Using a multidisciplinary approach that integrates network pharmacology, molecular docking, in vitro assays, and predictive in silico analyses of drug-like properties, ADME, and toxicity, the mechanisms and properties of fupenzic acid have been elucidated. Network pharmacology analysis identified the potential targets for fupenzic acid, with enrichment analyses revealing key processes like inflammatory response, cytokine signaling, innate immune system, and MAPK cascade regulation. Transcription factors such as RELA, SP1, and NFKB1 were predicted to play crucial roles in its therapeutic effects. PPI network analysis underscored NF-κB as a central hub, linking these pathways to its anti-inflammatory effects. In vitro experiments demonstrated that fupenzic acid effectively suppressed inflammatory mediators like NOS-2 and COX-2, through the NF-κB pathway. Molecular docking further confirmed its favorable interaction with NF-κB, reinforcing its mechanism of action. Additionally, in silico ADMET profiling revealed favorable drug-like properties including pharmacokinetics and toxicity profiles, emphasizing its suitability as a drug candidate. This study represents a major step forward in understanding the therapeutic potential of fupenzic acid, establishing it as a distinctive and promising anti-inflammatory agent. The findings identified it as a pharmacological agent for clinical development targeting inflammation-driven diseases and also provide a foundation for future translational research.

Acute myeloid leukemia (AML) is an aggressive hematological malignancy. Patients with wild-type FLT3 relapsed or refractory (R/R) AML face significant therapeutic challenges due to the persistent lack of effective treatments. A comprehensive understanding of the mechanisms underlying chemotherapy resistance is needed to the development of effective treatment strategies. Therefore, we investigated the molecular mechanisms underlying cytarabine (Ara-C) resistance and daunorubicin (DNR) tolerance in Ara-C-resistant RHI-1 cells derived from the wild-type FLT3 AML cell line SHI-1. Quantitative analysis of intracellular drug concentrations, proteomics, and phosphoproteomics showed that DNR resistance in Ara-C-resistant RHI-1 cells is driven by metabolic remodeling toward mitochondrial metabolism, upregulation of DNA repair pathways, and enhanced reactive oxygen species (ROS) detoxification rather than reduced drug uptake. Moreover, targeting these compensatory mechanisms, particularly the OXPHOS complex I proteins, significantly improved the efficacy of both Ara-C and DNR. Conclusively, these findings highlight mitochondrial metabolism and DNA repair as critical factors in chemotherapy resistance and offer valuable insights into potential therapeutic targets for enhancing treatment outcomes in patients with wild-type FLT3 R/R AML.

Under stress conditions, such as ex vivo culture, chemotherapy, irradiation and infection, haematopoietic stem cells (HSCs) actively divide to maintain blood cell production. This process leads to production of reactive oxygen species (ROS) that causes HSC exhaustion and haematopoietic failure. Here we show that ferumoxytol (FMT; Feraheme), a Food and Drug Administration-approved nanodrug, is a powerful ROS scavenger capable of relieving ROS in stressed HSCs, facilitating their post-injury regeneration. Mechanistically, the catalase-like activity of FMT reduces intracellular levels of H2O2 and diminishes H2O2-induced cytotoxicity. Moreover, FMT maintains long-term regenerative capacity of transplanted HSCs in pre-conditioned leukaemic mice and shows potential to effectively eliminate leukaemia in vivo while preserving HSCs. Our study highlights FMT as a powerful clinical tool to promote haematopoietic cell recovery in patients undergoing stress-generating treatments.

Cichlid fishes inhabiting the East African Great Lakes, Victoria, Malawi, and Tanganyika, are textbook examples of parallel evolution, as they have acquired similar traits independently in each of the three lakes during the process of adaptive radiation. In particular, 'hypertrophied lip' has been highlighted as a prominent example of parallel evolution. However, the underlying molecular mechanisms remain poorly understood. In this study, we conducted an integrated comparative analysis between the hypertrophied and normal lips of cichlids across three lakes based on histology, proteomics, and transcriptomics. Histological and proteomic analyses revealed that the hypertrophied lips were characterized by enlargement of the proteoglycan-rich layer, in which versican and periostin proteins were abundant. Transcriptome analysis revealed that the expression of extracellular matrix-related genes, including collagens, glycoproteins, and proteoglycans, was higher in hypertrophied lips, regardless of their phylogenetic relationships. In addition, the genes in Wnt signaling pathway, which is involved in promoting proteoglycan expression, was highly expressed in both the juvenile and adult stages of hypertrophied lips. Our comprehensive analyses showed that hypertrophied lips of the three different phylogenetic origins can be explained by similar proteomic and transcriptomic profiles, which may provide important clues into the molecular mechanisms underlying phenotypic parallelisms in East African cichlids.

Oxygen shortage, known as hypoxia, occurs commonly in both physiological and pathological conditions. Transcriptional regulation by hypoxia-inducible factors is a dominant regulatory mechanism controlling hypoxia-responsive genes during acute hypoxia; however, recent studies suggest that post-transcriptional regulation, including RNA degradation, also involves hypoxia-induced gene expression during the chronic hypoxia. In this study, we developed a method to quantify the contributions of RNA synthesis and degradation to differential gene expression, and identified 102 genes mainly regulated via RNA degradation under chronic hypoxia in HCT116 cells. Bioinformatics analysis showed that the genes mainly regulated by RNA degradation were involved in glycolysis. We examined changes in the RNA-binding ability of RNA-binding proteins by RNA interactome capture and statistical analysis using public databases. We identified fragile X messenger ribonucleoprotein 1 (FMRP) as an RNA-binding protein involved in the chronic hypoxia-induced increase in mRNAs encoding rate-limiting enzymes. This study emphasizes the importance of post-transcriptional gene regulation under chronic hypoxia in HCT116 cells.

Although the phosphorylation of serine (S), threonine (T), and tyrosine (Y) is well-established, arginine phosphorylation (pR) has recently garnered significant attention due to its crucial role in bacteria pathogenicity and stress response. Mycolicibacterium smegmatis, a nonpathogenic surrogate of Mycobacterium tuberculosis, serves as a model for studying mycobacterial pathogenesis. A recent proteomics study identified six pR proteins in M. smegmatis. To gain a more comprehensive understanding, we performed pR profiling using mass spectrometry in combination with two distinct phosphopeptide enrichment strategies: titanium-immobilized metal ion affinity chromatography (Ti4+-IMAC) and Fe-NTA cartridge purification. This approach led to the identification of 1192 shared pR peptides with 1553 pR sites in M. smegmatis following both competitive and non-competitive scoring assessments for pR and pS/T/Y. Further stringent filtering through manual verification resulted in 58 high-confident pR sites across 57 proteins. These confirmed pR-proteins are functionally related, particularly in DNA binding and ATP binding. Alterations in the modification of three pR sites during the logarithmic and stationary phases at the phosphorylation level, but not at the total cell protein level, further suggest the role of pR in the bacterium's functional adaptation to its environment. SIGNIFICANCE: Our findings reveal that pR proteins are prevalent and play roles in DNA-binding and ATP-binding activities, providing insights into the broader biological functions of pR peptides in other genetically diverse species. The reliable identification of bacterial pR events in M. smegmatis not only propels the study of pR within the realm of proteomics but also paves the way for exploring its detailed function in bacteria.

The essence of the plant drought tolerance mechanism lies in determining protein expression patterns, identifying key drought-tolerant proteins, and elucidating their association with specific functions within metabolic pathways. So far, there is limited information on the long-term drought tolerance of Haloxylon ammodendron and Haloxylon persicum grown in natural environments, as analyzed through proteomics. Therefore, this study conducted proteomic research on H. ammodendron and H. persicum grown in natural environments to identify their long-term drought-tolerant protein expression patterns. Totals of 71 and 348 differentially expressed proteins (DEPs) were identified in H. ammodendron and H. persicum, respectively. Bioinformatics analysis of DEPs reveals that H. ammodendron primarily generates a large amount of energy by overexpressing proteins related to carbohydrate metabolism pathways (pyruvate kinase, purple acid phosphatases and chitinase), and simultaneously encodes proteins capable of degrading misfolded/damaged proteins (tam3-transposase, enhancer of mRNA-decapping protein 4, and proteinase inhibitor I3), thus adapting to long-term drought environments. For H. persicum, most DEPs (enolase and UDP-xylose/xylose synthase) involved in metabolic pathways are up-regulated, indicating that it mainly adapts to long-term drought environments through mechanisms related to positive regulation of protein expression. These results offer crucial insights into how desert plants adapt to arid environments over the long term to maintain internal balance. In addition, the identified key drought-tolerant proteins can serve as candidate proteins for molecular breeding in the genus Haloxylon, aiming to develop new germplasm for desert ecosystem restoration.

Wenweishu (WWS) is a traditional Chinese medicine compound formulated for chronic atrophic gastritis (CAG) treatment by warming the stomach and alleviating pain. However, its pharmacological mechanisms remain underexplored.

This study investigated the therapeutic effects and potential mechanisms of WWS on CAG with spleen-stomach cold deficiency syndrome (SSCDS).

To achieve this, an SSCDS-CAG rat model and a human gastric mucosal epithelial cells (GES-1) cell model were established using multi-factor modeling and N-Methyl-N'-nitro-N-nitrosoguanidine (MNNG) induction, respectively. WWS's effects on gastric injury were evaluated through pathology, inflammation, serum biomarkers, and apoptosis. Additionally, MNNG's effects on GES-1 cells were analyzed. Network pharmacology, involving protein-protein interaction networks, GO/KEGG enrichment, and molecular docking, was employed to predict WWS's potential targets and mechanisms in SSCDS-CAG. Mechanistic insights were further validated using immunohistochemistry, quantitative reverse transcription polymerase chain reaction, and western blotting.

In vivo results showed that WWS alleviated symptoms in SSCDS-CAG rats, lowering symptom scores and improving gastric histopathology. It modulated serum biomarkers and reduced inflammation and apoptosis in both in vivo and in vitro studies. Network pharmacology results revealed 263 overlapping targets between WWS and SSCDS-CAG, associated with apoptosis, inflammation, and the PI3K/AKT pathway. Molecular docking revealed strong binding affinity between the core target and active WWS components. In SSCDS-CAG rats and GES-1 cells, WWS inhibited PI3K/AKT phosphorylation, increased PTEN expression, and regulated Bcl-2, Bax, and cleaved caspase-3 levels.

WWS reduces inflammation and apoptosis in multi-factor CAG rats and MNNG-induced GES-1 cells by modulating the PTEN/PI3K/AKT signaling pathway and apoptosis-related proteins.

Mutations in the tumor suppressor gene Folliculin (FLCN) are responsible for Birt-Hogg-Dube' (BHD) syndrome, a rare inherited condition that predisposes affected individuals to skin tumors, pulmonary cysts, and kidney tumors. FLCN regulates key cellular pathways, including TFEB, TFE3, and mTORC1, which are critical for maintaining cell homeostasis. Loss of FLCN leads to both hyperactivation of mTORC1 and constitutive activation of TFEB and TFE3, contributing to tumorigenesis. While previous studies showed that Flcn liver-specific conditional knockout (FlcnLiKO) mice are protected from developing liver fibrosis and damage upon high-fat diet exposure, the potential role of FLCN loss in liver carcinogenesis remained unexplored. Here, we demonstrate that hepatic loss of FLCN in mice results in cancer associated with inflammation and fibrosis with features of cholangiocarcinoma (CCA). This phenotype emerges in mice over 90-week-old, with a male predominance. Moreover, FlcnLiKO mice are more prone to develop diethylnitrosamine (DEN)- or 3,5-diethoxycarbonyl-1,4-dihydrocollidine (DDC)- induced liver tumors with heterogenous histological features. Notably, depletion of TFE3, but not TFEB, in the liver of FlcnLiKO mice fully rescues the cancer phenotype and normalized mTORC1 signaling, highlighting TFE3 as the primary driver of liver cancer and mTORC1 hyperactivity in the absence of FLCN.