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PubMed2026

Identification of the BrSK gene family in flowering Chinese cabbage and functional characterization of BrSK2 subfamily involvement in heat stress.

Glycogen synthase kinase 3 (GSK3) kinases are evolutionarily conserved regulators of plant development and stress signaling, yet their contributions to thermotolerance in cool-adapted Brassica crops remain poorly understood. Here, we identified 16 BrSK genes in the Caixin (Brassica rapa ssp. chinensis var. parachinensis) genome, all harboring intact catalytic motifs indicative of functional kinase activity. Spatiotemporal expression profiling revealed preferential accumulation of BrSK transcripts in stem apices and floral organs during reproductive transition, while promoter analysis identified abundant heat- and abiotic stress-responsive cis-elements. Under heat stress, BrSK21, BrSK22, and BrSK23 displayed striking genotype-specific expression dynamics. BrSK21/22/23 transcripts were stably suppressed in the heat-tolerant cultivar '49-19' but transiently declined before rapidly rebounding in the heat-sensitive 'Liuye 50', mirroring RNA-seq profiles. Protein-protein interaction assays (Y2H, BiFC, and LCI) demonstrated specific associations between BrSK kinases and BrHSFA1. Functional validation via VIGS revealed that silencing of BrSK21 significantly enhanced thermotolerance, with triple silencing of BrSK21/22/23 conferring additive protection, indicating functional redundancy within the BrSK2 subfamily. Collectively, these findings establish the BrSK2 subfamily as negative regulators of heat tolerance in Caixin, likely via modulation of BrHSFA1 expression. This work identifies high-priority targets for molecular breeding of climate-resilient Brassica vegetables.

Plant Proteins↗

PubMed2026

A 7-mm Covered TIPS Reduces Hepatic Encephalopathy Without Increasing Rebleeding in Cirrhotic Patients With Small Liver: A Randomized Study.

BACKGROUND/AIMS: International guidelines recommend initiating transjugular intrahepatic portosystemic shunt (TIPS) placement with an 8-mm stent. However, there is an evident lack of randomized controlled trials evaluating TIPS diameters < 8 mm in cirrhotic patients with a relatively small liver. The aim of this study was to determine whether 7 mm-covered TIPS, compared with 8-mm stents, could achieve comparable shunt function with a lower incidence of hepatic encephalopathy (HE). METHODS: In this multicenter randomized controlled trial, patients with cirrhosis and relatively small liver were randomized 1:1 to receive TIPS with a 7-mm (n = 92) or 8-mm (n = 92) covered stent to prevent variceal rebleeding. The primary endpoint was the incidence of overt HE after randomization. All-cause rebleeding, orthotopic liver transplantation (OLT)-free survival and a composite of these outcomes, were designated as secondary endpoints. RESULTS: Among the 184 enrolled patients, the predominant etiologies of liver cirrhosis were hepatitis B virus infection (56.0%) and alcohol-related liver disease (20.7%). Over a median follow-up of 26.5 months, overt HE occurred in 19 patients (20.7%) in the 7-mm group and 33 patients (35.9%) in the 8-mm group. The 2-year cumulative incidence of overt HE was significantly lower in the 7-mm group than in the 8-mm group (21.4% vs. 37.2%, p = 0.02). Stent diameter, post-TIPS portosystemic pressure gradient, pre-covert HE and MELD-Na score were identified as independent risk factors for overt HE. The rates of shunt dysfunction were statistically similar between groups (8.7% vs. 8.7%, p = 1.0), as were 2-year rebleeding rates (10.9% vs. 9.8%, p = 0.81) and OLT-free survival rates (91.3% vs. 88.0%, p = 0.82). CONCLUSIONS: A 7-mm covered TIPS demonstrated comparable shunt function to an 8-mm covered stents, with a significantly lower risk of overt HE. These findings support consideration of 7-mm TIPS stents for preventing variceal rebleeding in cirrhotic patients with a small liver who are undergoing TIPS. TRAIL REGISTRATION: ClinicalTrials.gov, NCT02541825.

Humans↗

PubMed2026

TNF-NF-κB signaling mediates immune-biomineralization crosstalk during shell repair under ocean acidification in Mytilus edulis.

Ocean acidification (OA) impairs biomineralization in bivalves, but its effects on immune-biomineralization crosstalk during shell repair remain unknown. Here, we exposed adult Mytilus edulis bearing standardized shell perforations to three pH levels (8.1, 7.9, and 7.7) for up to 40 days. OA slowed early repair and caused microstructural disorganization and an approximately 87% reduction of compressive strength at pH 7.7, yet the damaged area appeared largely closed by day 15, suggesting a decoupling between morphological closure and functional recovery. In addition, transcriptomic profiling of hemocytes and mantle tissue, based on an average of 6.5 Gb of clean reads per sample mapped to the M. edulis reference genome (NCBI Assembly GCF_000511035.1), revealed that these shell-level defects were accompanied by coordinated immune and metabolic reprogramming. Hemocytes, the primary immune effector cells of bivalves, exhibited pH- and time-dependent shifts with moderate acidification (pH 7.9) promoting inflammatory transcripts, whereas severe acidification (pH 7.7) suppressed these signals while upregulating stress-associated pathways; both treatments consistently downregulated lysosomal proteases and NF-κB negative regulators. The mantle, a primarily mineralizing organ, paradoxically upregulated immune-related genes while suppressing oxidative phosphorylation and extracellular matrix pathways. This tissue-level imbalance, with hemocytes recruited but functionally constrained and mantle metabolically suppressed yet immunologically activated, points to TNF-NF-κB pathway modulation as a key mediator of shell repair under acidification. Our findings demonstrate that visible shell closure masks underlying structural and mechanical failure, and that immune regulation, rather than simple suppression or activation, critically shapes the repair outcome. These results advocate for multifunctional indicators beyond closure area to assess shell integrity in acidified marine environments.

Animals↗

PubMed2026

Genome-wide analysis of the plant-specific PLATZ gene family in Taraxacum kok-saghyz and its roles in response to drought and salt tolerance.

Abiotic stress severely limits plant growth and productivity. Taraxacum kok-saghyz Rodin (TKS), known for its environmental resilience, represents a valuable resource for identifying stress-tolerant genes to improve stress-adaptive crops. Plant AT-rich protein and zinc-binding protein (PLATZ) transcription factors serve as core regulators of plant growth, developmental processes, and adaptive responses to various stress conditions; however, they remain uncharacterized in TKS. Here, we identified 10 TksPLATZ genes through a whole-genome analysis. Phylogenetically, these genes were grouped into five distinct evolutionary branches. Promoter sequence analysis revealed multiple types of cis-acting regulatory elements that are connected with hormonal signal responses and environmental stress adaptation. Integrated analysis of transcriptome datasets and RT-qPCR validation demonstrated that TksPLATZ genes display tissue-specific expression profiles and show distinct responsive patterns to drought and salt stress treatments. Among them, TksPLATZ1, TksPLATZ2 and TksPLATZ7 were markedly induced under both stressors and were selected for further functional study. We demonstrated that TksPLATZ1, TksPLATZ2 and TksPLATZ7 localize to the cell nucleus and act as transcriptional activators and repressors, respectively. Phenotypic data from overexpression experiments in plants confirm that heterologous expression of TksPLATZ1, TksPLATZ2, and TksPLATZ7 enhances the tolerance of Arabidopsis to salt and osmotic stress. These findings provide valuable genetic resources for improving plant tolerance to environmental stresses.

Salt Tolerance↗

PubMed2026

Smoking Cue Reactivity in Relation to Uncertain-Threat and Reward-Anticipation Networks: A Coordinate-Based fMRI Meta-Analysis.

BACKGROUND: Smoking is a concerning medical and social problem, yet how the brain links stress to continued smoking is still not well understood. This coordinate-based meta-analysis identified convergent activations for smoking cues, uncertain threat, and reward anticipation, and examined co-activation patterns to clarify whether smoking-cue activity in smokers relates to threat and reward activity in non-addicted controls. METHODS: We conducted a coordinate-based activation likelihood estimation (ALE) meta-analysis of 102 fMRI studies (N = 3,068), including 30 studies on smoking cue reactivity (n = 945), 48 on reward anticipation (n = 1,413), and 24 on uncertain threat processing (n = 1,621). We performed single, conjunction, and contrast analyses, followed by meta-analytic connectivity modeling (MACM) of key regions. RESULTS: Single analysis revealed smoking engaged bilateral ACC (-1.1, 46.2, -1.1), uncertain threat engaged bilateral insula (left = -33.6, 22, 4.3, right = 41.3, 22, 1), reward anticipation activated thalamus (0.9, 1.3, -3.1) and medial frontal gyrus (2.7, 6.6, 52.1). Conjunction and contrast analyses showed shared or unique activation for each task in its respective regions. MACM showed ACC co-activation with thalamus and medial frontal gyrus, while insula co-activated with ACC and inferior frontal gyrus. CONCLUSIONS: Each process converged in a separate region, with no overlap between the smoking-cue map and either the threat or reward map. The ACC nonetheless co-activated with reward-related regions and shared network membership with the threat-related insula. On this basis we hypothesize a shift in motivation from stress-driven reward toward cue-driven craving, to be tested within subjects, and identify candidate neuromodulation targets for preventing stress-precipitated relapse.

fMRI↗

PubMed2026

ADAM10's combined influence on the diagnostic usefulness of IL 22, IL 10, IL-17 A, and IL-17D in autism spectrum disorders: Predicted role on gut leakiness as co-morbidity.

Autism spectrum disorder (ASD) is a complex neurodevelopmental disorder with increasing global prevalence but a lack of reliable diagnostic biomarkers. Emerging evidence suggests that immune dysregulation, gut-brain axis dysfunction, and increased intestinal permeability play key roles in ASD pathophysiology. This study investigated the combined diagnostic value of ADAM10 and cytokines (IL-10, IL-22, IL-17 A, and IL-17D). Multivariable logistic regression produces an improved ROC curve that improves diagnostic accuracy over individual markers by combining numerous predictors into a single risk score (linear predictor). The technique, which frequently raises individual marker AUCs, entails modelling a binary result, calculating the probability, and visualizing ROC based on the projected probabilities. In this case-control study, plasma levels of ADAM10, IL-10, IL-22, IL-17 A, and IL-17D were measured in 37 male children with ASD and 37 age-matched controls. Group comparisons, correlation analyses, and receiver operating characteristic (ROC) curve analyses, including combined ROC models, were performed. ADAM10, IL-22, and IL-17 A levels were significantly reduced in children with ASD compared to controls, whereas IL-10 and IL-17D showed no significant differences. ADAM10, IL-17 A, and IL-22 demonstrated good diagnostic performance, with AUC values of 0.886, 0.855, and 0.812, respectively. In contrast, IL-10 and IL-17D showed poor discriminatory ability, with AUC values of 0.524 and 0.599, respectively. Combined ROC analysis markedly improved diagnostic accuracy, with all panels including ADAM10 achieving AUC values above 0.90, and some reaching as high as 0.988, with high sensitivity and specificity. The combination of ADAM10 with selected cytokines significantly enhances diagnostic performance compared to individual markers, supporting a link between immune dysregulation, barrier dysfunction, and gut permeability in ASD.

Humans↗

PubMed2026

Exploring precision risk in pediatric vesicoureteral reflux: Innate immune gene variations and reflux outcomes in the RIVUR cohort.

INTRODUCTION: Children with vesicoureteral reflux (VUR) are at increased risk for morbidity from recurrent urinary tract infections (UTIs), yet the factors influencing spontaneous VUR resolution remain poorly defined. This study evaluates whether genetic variations in key urinary innate immune effectors (DEFA1A3, DMBT1, and RNASE7) influences VUR resolution and interacts with prophylaxis to alter clinical response. METHODS: We conducted a secondary analysis of 303 RIVUR participants with available DEFA1A3 and DMBT1 copy number variation (CNV) data and RNASE7 rs1263872 genotype. Primary outcomes were (1) VUR improvement (decrease in grade) and (2) VUR resolution at study exit. Multivariable logistic regression models included genotype, treatment, and their interactions, adjusting for age, sex, baseline grade (high vs low), laterality, bowel/bladder dysfunction, and any UTI. Internal validation used 2000-sample bootstrap with bias-corrected and accelerated confidence intervals and influence diagnostics. RESULTS: Clinical covariates did not significantly predict VUR improvement. Children with DEFA1A3 CNV >5 had higher odds of improvement (OR 2.36, 95% CI 1.12-4.96, p = 0.023), an effect that remained significant in bootstrap analyses. High-grade VUR was associated with lower odds of resolution (OR 0.34, 95% CI 0.12-0.94, p = 0.038). A significant interaction was observed between prophylaxis and high DMBT1 copy number for VUR resolution (interaction OR 2.99, 95% CI 1.11-8.04, p = 0.031); no interaction was seen for improvement. RNASE7 rs1263872 was not associated with either outcome. CONCLUSION: Innate immune gene variation may contribute to heterogeneity in VUR outcomes. High DEFA1A3 copy number was associated with reflux improvement and a DMBT1-prophylaxis interaction was associated with reflux resolution. The results of this study is hypothesis-generating and prompt further evaluation to assess whether a subset of children may experience structural benefit from prophylaxis or have a more favorable natural history based on their innate immune genotype.

Humans↗

PubMed2026

GPR3 in neuro-metabolic-immune-reproductive nexus - a potential therapeutic target for Multi-System diseases.

BACKGROUND: GPR3(G-protein-coupled receptor 3), an orphan G-protein-coupled receptor (GPCR) with constitutive Gs activity, is expressed in the brain, liver, ovary, and other tissues, regulating cell proliferation, differentiation, and apoptosis across the nervous, reproductive, immune, and metabolic systems. This review synthesizes evidence on its integrated signaling and physiological functions to address the lack of a comprehensive multisystem pathophysiology overview. METHODS: A systematic literature search was conducted on PubMed and Web of Science, using keywords such as "GPR3", "GPCR", "neurodegeneration", "metabolism", "immune", "reproduction", "agonist", "inhibitor", and "therapeutic target". This search identified GPR3's roles in neurodegenerative diseases, immune inflammation, reproduction, and energy metabolism. The analysis focused on signaling pathways, ligand regulation, and therapeutic potential. RESULTS: The research indicates that GPR3 is involved in neuronal survival, synaptic plasticity, and microglial activity via the cAMP/PKA, PI3K/Akt, and β - arrestin pathways. It promotes amyloid - β formation in Alzheimer's disease (AD), yet provides neuroprotection in Parkinson's disease (PD) models. It may contribute to anxiety/depression - like states, maintain oocyte meiotic arrest in the ovary, and activate thermogenic genes in adipose tissue. GPR3 modulates immune responses. Using oleic acid (OA) and diphenyleneiodonium (DPI) as activators, and AF64394 and cannabidiol (CBD) as antagonists, it shows potential in disease models. CONCLUSION: GPR3 acts as a central molecular hub integrating neural, metabolic, immune, and reproductive signaling, highlighting its potential as a therapeutic target for chronic multisystem disorders. However, its dual roles in certain pathologies and translation challenges necessitate further research.

Humans↗

PubMed2026

Bi-compartmental CSF-serum analysis of NfL and GFAP differentiates central and peripheral pathology in neuroinfectious diseases: A monocentric real-world cohort study.

Neurofilament light chain (NfL) and glial fibrillary acidic protein (GFAP), established biomarkers of neuroaxonal injury and astroglial pathology, are frequently only assessed in blood, which limits conclusions regarding their origin. Bi-compartmental analyses of CSF and serum may help differentiate central or peripheral origin of biomarker elevation. Moreover, studies on NfL and GFAP in distinct neuroinfectious disease (NID) phenotypes, particularly those based on real-world cohorts, are limited. This retrospective monocentric study analyzed CSF and serum from patients with (meningo-)encephalitis/myelitis (TI+; n = 48), meningitis (TI-; n = 80), (cranial) nerve palsies/polyradiculitis (PND; n = 61), and 113 non-neuroinflammatory/non-neurodegenerative controls. A bi-compartmental model using scatter plots and simple linear regression was applied to assess the origin of blood biomarker levels and discriminate between central and peripheral pathology. CSF and serum NfL and GFAP z-scores were significantly higher in TI+ compared with TI- (CSF-GFAP p < 0.001/sGFAP p = 0.0083; CSF-NfL p = 0.003/sNfL p = 0.0004). TI+ and PND differed only in GFAP levels, which were higher in TI+ (CSF-GFAP p = 0.0049/sGFAP p = 0.003). The overall group effect (p ≤ 0.003) and principal findings remained significant after adjustment for age, sex, QAlb, and time since (symptom) onset to LP. Bi-compartmental analysis revealed simultaneous elevation of CSF and serum NfL in TI+, indicating predominantly central origin, whereas PND demonstrated a shift toward higher sNfL levels suggesting peripheral origin. Higher clinical severity (modified Rankin Scale 3-5) was associated with elevated serum and CSF GFAP and NfL (sGFAP p = 0.012/sNfL p = 0.002; CSF-GFAP p < 0.0001/CSF-NfL p = 0.0001), which also predicted unfavorable outcome at discharge (sGFAP p = 0.006/sNfL p = 0.004; CSF-GFAP p = 0.003/CSF-NfL p = 0.012). NfL and GFAP were associated with brain/myelon involvement in NID, predominantly reflecting central pathology. Despite strong CSF-serum correlations, bi-compartmental approaches provide additional insight into biomarker origin and disease compartment.

Humans↗

PubMed2026

Emergence of Babesia naoakii infection in Indonesian domestic cattle, a new host record in water buffaloes, and characterization of complete mitochondrial protein-coding genes.

Babesia (B.) naoakii, previously referred to as Babesia sp. Mymensingh, is a recently characterized tick-borne haemoprotozoan parasite of cattle. In Indonesia, we first reported its presence in 2022 from clinically affected cattle in Central Java. To investigate the wider epidemiology of this neglected ruminant-associated Babesia species, we surveyed apparently healthy cattle (Bos indicus) and water buffaloes (Bubalus bubalis) across three districts of Java, Indonesia. A PCR assay targeting the B. naoakii-specific apical membrane antigen 1 (ama1) gene detected the parasite occurrence in 34.39% of assessed cattle (87/253; 95% CI: 28.80-40.44%) and 30.77% of water buffaloes (12/39; 95% CI: 18.47-46.52%). These results represent the first record of B. naoakii infection in water buffaloes in the country and confirm that the parasite circulates in subclinically infected bovine hosts. To characterise this apicomplexan parasite further at the molecular level, we assembled in full length the three mitochondrial protein-coding genes (PCGs): cytochrome c oxidase subunits 1 (cox1) and 3 (cox3), as well as cytochrome b (cytb). These genes were reconstructed by next-generation sequencing of blood DNA collected during the acute haemolytic-phase of B. naoakii infection, from calves that subsequently succumbed to the disease in the endemic area. Phylogenetic analyses of the concatenated amino-acid sequences of cox1, cox3, and cytb placed the Indonesian isolates within a well-supported monophyletic clade, distinct from all previously characterised ruminant-associated Babesia species and sister to the Babesia bigemina/Babesia ovata lineage. This placement confirmed species identity and reinforced the genetic distinctiveness of B. naoakii in Indonesia. Notably, although B. naoakii circulates in peripheral blood and mirrors the diagnostic behaviour of the mild pathogen B. bigemina, its clinical impact more closely resembles that of the severe pathogenic B. bovis, particularly in young animals. This diagnostic-clinical discordance highlights the need for B. naoakii-specific molecular surveillance and species-level differentiation in regions of co-endemicity. Given the high prevalence in subclinically B. naoakii-infected adults, the documented severity of babesiosis in calves, and the potential for substantial economic losses, broader epidemiological investigations and species-specific control measures for B. naoakii are urgently performed. The same holds true for future epizootiological investigations of underdiagnosed B. naoakii-infections possibly circulating in Indonesian endemic ruminant bovids such as the banteng (Bos javanicus), the lowland anoa (Bubalus depressicornis) and the tamaraw (Bubalus mindorensis).

Animals↗

PubMed2026

Transcriptomic and RNAi analyses reveal chloride channel 3-associated osmoregulation in Litopenaeus vannamei under low-salinity stress.

Chloride channels and transporters are important for cellular volume regulation and salinity adaptation in euryhaline crustaceans, yet the intestinal transcriptional relationship between plasma-membrane and intracellular chloride pathways remains unclear in Litopenaeus vannamei. In this study, RNA interference of anoctamin 1 (ANO1) was combined with intestinal transcriptome sequencing under the production-relevant low-salinity condition of salinity 3. ANO1 silencing produced a focused transcriptional response, with 16 differentially expressed genes (DEGs) identified (11 upregulated and 5 downregulated). Functional enrichment indicated that these genes were associated with transporter activity, cytoskeletal organization, extracellular matrix-receptor interaction, membrane lipid metabolism, and vesicular processes. Notably, a transcript encoding chloride channel protein 3 (CLC-3) was significantly upregulated following ANO1 knockdown, suggesting a potential transcriptional relationship between ANO1 and CLC-3 in chloride homeostasis. Based on this finding, CLC-3 was selected for full-length cDNA cloning, sequence characterization, salinity-gradient expression analysis, and RNAi-based functional assessment. The cloned CLC-3 cDNA was 2883 bp in length and encoded an 850 amino acid protein containing a conserved voltage-gated chloride channel (Voltage-CLC) domain and two cystathionine β-synthase domains. Phylogenetic analysis placed LvCLC-3 within the intracellular CLC-c clade, and tissue distribution analysis showed the highest CLC-3 expression in the intestine. Intestinal CLC-3 expression responded nonlinearly to salinity variation, peaking at salinity 20. Under salinity 3, CLC-3 knockdown reduced ANO1, Na+/K+-ATPase alpha subunit, and Na+-K+-2Cl- cotransporter transcript levels, whereas glutamate-gated chloride channel expression increased. Mild hepatopancreatic structural alterations were also observed after CLC-3 knockdown. These findings suggest that CLC-3 is a salinity-responsive intracellular chloride-transporter candidate associated with intestinal ion-transport-related transcriptional responses after ANO1 suppression in L. vannamei, although the underlying physiological mechanism requires further validation.

Animals↗

PubMed2026

Proteomic analysis of cisplatin-induced spermatogenesis defects in mice.

BACKGROUND: Cisplatin is a crucial chemotherapeutic agent used for treating various cancers; however, its excessive use can cause irreversible damage to the reproductive system, and the protein expression profile of cisplatin-induced testicular injury remains unclear. METHODS: Male C57BL/6 mice were treated with cisplatin at various doses, and testes were collected for histological, immunofluorescence, and proteomic analyses. Germ cell loss and apoptosis were assessed using H&E staining, TUNEL assays, and immunofluorescence for LIN28A, SYCP3, MVH, and CDK1. Label-free quantitative proteomics identified differentially expressed proteins, which were analyzed for functional enrichment and protein-protein interactions. RESULTS: We observed that cisplatin treatment led to smaller testes, reduced sperm count, and a significant decrease in the number of spermatocytes and spermatids in mice. Label-free quantitative proteomic analysis revealed that cisplatin significantly reduced the expression of cyclin-dependent kinase 1 (CDK1), a key spermatogenesis regulator, in the testes. Reduction in CDK1 expression is correlated with spermatogenic arrest, particularly in spermatocytes. CONCLUSION: These findings highlight the critical role of CDK1 in cisplatin-induced spermatogenic dysfunction and provide new insights into fertility preservation strategies for patients with cancer undergoing chemotherapy.

Animals↗

PubMed2026

Inducible flocculation in Komagataella phaffii enables enhanced biomass separation for biopharmaceutical production.

Biomass separation represents a critical bottleneck in Komagataella phaffii-based biopharmaceutical processes, as typically high cell densities of 40 - 50 % create significant operational, technical and economic challenges for harvest operations. Yeast cell aggregation (flocculation) provides a solution to accelerate cell sedimentation by increasing particle size, thus allowing to improve biomass-supernatant separation efficiency during both natural gravity settling and (continuous) centrifugation operations. This study demonstrates successful engineering of K. phaffii strains with an inducible flocculation phenotype using CRISPR/Cas9-based genome editing to integrate the Saccharomyces cerevisiae FLO1 (ScFLO1) gene under control of various regulatory elements, including methanol-inducible and derepressible promoters. Flocculation strength could be enhanced by implementing transcriptional positive feedback circuits based on the methanol-inducible AOX1 promoter. To address methanol-free production requirements, we developed alternative systems to retrofit PAOX1-based ScFLO1 expression and exploited the derepressible PDF promoter, offering broader compatibility with biopharmaceutical manufacturing facilities. Flocculating cells cultivated in a bioreactor demonstrated significantly improved sedimentation behavior, with considerably lower supernatant turbidity after short low-speed centrifugation or gravity sedimentation compared to non-flocculating controls. Crucially, cell flocculation had no negative impact on product amount and quality when expressing a multivalent NANOBODY® VHH molecule with pharmaceutical relevance. Thus, this work establishes the first genetically engineered flocculation system in K. phaffii compatible with recombinant protein production, providing the basis for an innovative approach to streamline harvest operations in biopharmaceutical processes.

Flocculation↗

PubMed2026

Inhibitory mechanism of phloretin on the AgrA LytTR domain-agr operon complex formation and its application in beef.

Staphylococcus aureus (S. aureus) represents a major foodborne pathogen whose enterotoxin production poses significant challenges to food safety due to its high environmental resistance and limited efficacy of conventional sterilization. Since the expression of enterotoxins is predominantly governed by the agr quorum sensing system, targeting this regulatory pathway has become a strategic choice for virulence control. This study elucidated the mechanism by which phloretin, a potential quorum sensing inhibitor, interferes with the agr system to attenuate virulence. To achieve this, the recombinant AgrA LytTR domain was expressed and purified, and its interaction with phloretin was characterized using thermal shift assays (TSA), electrophoretic mobility shift assays (EMSA), and molecular dynamics (MD) simulations. The results showed that phloretin specifically binds to the AgrA LytTR domain, enhancing its thermal stability and disrupting AgrA LytTR-agr operon binding by reducing the free energy of interaction between them, without causing significant structural rearrangement. Mechanistic analysis indicated that phloretin sterically hinders key β-sheet turn residues (HIS169, ASN201, ARG233), thereby impairing DNA recognition, downregulating RNAIII transcription, and inhibiting agr signaling. In cooked beef, phloretin significantly inhibited the secretion of enterotoxins and α-hemolysin, while delaying lipid oxidation and protein degradation, and maintaining the meat texture. These findings suggested that phloretin is a multifunctional substance with anti-virulence, antioxidant, and preservative properties, demonstrating its potential as a natural food preservative.

Phloretin↗

PubMed2026

The gonadal matrisome and its correlation with sex change in the ricefield eel Monopterus albus.

The matrisome is a comprehensive list of genes in the genome of an organism, which encodes proteins constituting or interacting with the extracellular matrix (ECM). The gonadal ECM is important for folliculogenesis and spermatogenesis. This study characterized the composition of the matrisome and the expression of matrisome genes in the gonad of ricefield eel, a protogynous sex-changing teleost, during sex change. A total of 838 matrisome genes were identified in the genome of ricefield eel through an in-silico orthology-based approach, of which 482, 443, 429, and 570 matrisome genes were shown to be expressed in the gonads of female (F), early intersexual (EI), mid-intersexual (MI), and late intersexual (LI) fish, respectively. Differentially expressed matrisome genes (DEMGs) were observed across all the sexual stages as well as in each category of ECM components. Analysis of DEMGs in the comparison between EI and F revealed dramatic upregulation of adam8a, mmp9, and s100a11 while downregulation of col4a5, col15a1b, clec3ba, f13a1, and ccl44, which were further confirmed by qPCR analysis. Together, these findings revealed significant changes in the expression of many matrisome genes, particularly three regulator genes, adam8a, mmp9 and f13a1, as female ricefield eels initiate sex change, suggesting that gonadal tissues undergo dramatic remodeling involving the regression of ovarian tissues and the development of testicular tissues to facilitate this process. These data provide valuable resources for further unraveling the roles of matrisome genes in gonadal development of ricefield eel and other vertebrates.

Animals↗

PubMed2026

On-filter fractionation by empFASP improves identification of membrane peptides in proteomic experiments.

Membrane proteins remain among the most analytically challenging targets in bottom-up proteomics due to their limited solubility and low abundance of protease-accessible sites within transmembrane domains. In addition, hydrophobic peptides are frequently lost during detergent removal and the on-filter processing steps. Here, we present empFASP, a straightforward on-filter-fractionation-based modification of the enhanced filter-aided sample preparation (eFASP) workflow that enhances recovery of membrane-embedded peptides otherwise lost during digestion and cleanup. The method combines controlled on-filter inversion with sequential ethyl acetate extraction at defined pH values, enabling recovery of peptide material retained on the filter and redistributed into detergent micelles. Compared with SP3 and SP4 in HEK293T lysates, empFASP increased unique hydrophobic peptide identifications by up to 48% and increased the proportion of detected transmembrane peptides. Application to mouse mitochondrial membranes and phosphatidylethanolamine-deficient and PE-containing Escherichia coli membranes showed that the additional fractions of empFASP contribute complementary recovery of hydrophobic and membrane-associated peptides, with the strongest gains observed at the peptide level. Because empFASP requires no specialized reagents or instrumentation, it can be readily implemented in standard proteomics workflows to improve coverage of membrane-embedded regions. SIGNIFICANCE: The empFASP (enhanced membrane peptide) workflow offers a practical solution to one of the persistent limitations in membrane proteomics-the underrepresentation of hydrophobic and transmembrane peptides in standard digests. By integrating simple pH-controlled extractions into an on-filter format, empFASP recovers peptides otherwise lost through adsorption or detergent micelle retention, substantially improving coverage of the membrane proteome. This method expands the analytical reach of bottom-up proteomics without requiring specialized instrumentation, making it immediately applicable for studies of membrane topology, protein-lipid interactions, and the structural consequences of altered membrane composition.

Proteomics↗

PubMed2026

Candidate biomarkers for early Giardia duodenalis infection revealed by time-resolved secretome proteomics.

Giardia duodenalis is a zoonotic protozoan parasite that causes giardiasis in humans and other mammals. Early diagnosis remains challenging because current diagnostic methods, including microscopy and enzyme-linked immunosorbent assays (ELISAs), primarily detect established infections. Consequently, a critical diagnostic gap exists during the early stage of infection within the first 2-48 h following exposure. To address this limitation, we characterized the proteins released by in vitro-cultured G. duodenalis trophozoites under serum-free conditions and evaluated their potential as early diagnostic biomarkers. Proteomic analysis of culture supernatants collected during early trophozoite incubation identified 31,773 peptides corresponding to 2504 quantifiable proteins. Temporal profiling showed distinct secretion patterns, including proteins that peaked during the early stage, progressively accumulated over time, or remained persistently abundant throughout the incubation period. Based on their secretion characteristics and predicted immunogenic properties, five candidate biomarkers were selected for further evaluation. Polyclonal antibodies raised against selected candidates successfully detected the corresponding proteins in serum-free culture supernatants, providing preliminary evidence for their potential utility as early-stage diagnostic targets. These findings identify stage-associated candidate proteins that may serve as a resource for future early giardiasis diagnostic development, provide a valuable resource for investigating host-parasite interactions, and establish a foundation for future diagnostic assay development. However, further validation in clinical and biological samples is required to confirm their diagnostic applicability. SIGNIFICANCE: Giardiasis, caused by Giardia duodenalis, is a major diarrheal disease worldwide. Although enzyme-linked immunosorbent assays (ELISAs) provide rapid detection, their diagnostic utility is limited by the lack of biomarkers capable of identifying infection during its earliest stages, creating a critical gap in the detection of active infection within 2-48 h following exposure. Using data-independent acquisition proteomics, this study provides a time-resolved characterization of proteins released by G. duodenalis trophozoites into serum-free culture supernatants. Our findings reveal temporal secretion dynamics of protein secretion and identify candidate biomarkers with potential utility for the development of early-stage diagnostic assays pending rigorous biological and clinical validation. In addition, this proteomic resource provides a foundation for investigating host-parasite interactions and may facilitate the development of future point-of-care diagnostic strategies.

Giardiasis↗

PubMed2026

Engineering bubble structures as Cas12a activators for highly sensitive monitoring of WRN helicase function.

The Werner syndrome helicase (WRN) is a critical synthetic lethal target in microsatellite instability cancers, essential for resolving complex genomic structures like replication bubbles and R-loops. However, strategies to simultaneously discriminate WRN activity on DNA versus DNA-RNA substrates in living cells are lacking. Here, we developed a structure-specific CRISPR/Cas12a biosensing strategy to visualize WRN functional activity by engineering bubble-structure probes. These probes were rationally designed to structurally mimic DNA replication bubbles and R-loop associated DNA-RNA hybrids. Upon specific unwinding by WRN, the probes release a sequestered activator strand that triggers Cas12a trans-cleavage, effectively converting the unwinding event into an amplified fluorescent signal. This assay achieves low picomolar sensitivity (LODs: 5.6-6.0 pM) and exceptional selectivity against homologous RecQ helicases. Uniquely, this strategy enables the parallel quantification of WRN activity on both substrate types, providing insights into distinct WRN-mediated pathways for resolving genomic stress. We further demonstrated the strategy's utility by visualizing endogenous WRN dynamics in living cells and profiling the efficacy of small-molecule inhibitors. This work offers a powerful molecular toolkit for dissecting WRN biology and facilitating high-throughput drug screening in targeted cancer therapy.

Werner Syndrome Helicase↗