DNA double-strand breaks (DSBs) pose a severe threat to genomic integrity, and cells rely on two major pathways for repair: homologous recombination and non-homologous end joining (NHEJ). While eukaryotic NHEJ requires a multi-component assembly including the Ku70/80 heterodimer, bacterial NHEJ operates with a simpler toolkit comprising a Ku homodimer and the multifunctional LigD. Despite this simplicity, the mechanism by which broken DNA ends are bridged together has remained unclear in bacterial NHEJ. Here, we present a cryo-electron microscopy structure of the Bacillus subtilis Ku (bsKu)-DNA complex at 2.74 Å resolution, capturing two blunt DNA ends bridged by a Ku protein alone. Supported by further biochemical assays, we propose an integrated model in which oligomeric arrays of Ku homodimers bridge and stabilize two DNA ends, facilitating efficient DSB repair in Bacillus subtilis. This work reveals a bsKu-mediated DNA bridging mechanism distinct from the eukaryotic system and provides critical structural insight into prokaryotic DNA repair.

Alzheimer disease (AD) is a progressive neurodegenerative disorder marked by transcriptomic alterations affecting multiple genes. Many researchers have tried to predict major hallmarks of AD pathogenesis for diagnosis but the association between receptor tyrosine kinase (RTK) pathways and AD diagnosis is still unclear. This study aims to identify RTK-associated gene signatures crucial to AD pathogenesis and assess their potential as diagnostic biomarkers for AD. The study investigated changes in RTK pathway gene expression related to AD by analyzing brain transcriptome data from two independent public data sets (GSE84422 and GSE109887). Differentially expressed genes (DEGs) were analyzed from the GSE84422 and GSE109887 data sets and overlapping genes (oDEGs) were identified. RTK-related genes (ooDEGs) were subsequently selected through functional enrichment analysis. These were further refined into AD-related genes (disease-associated genes (DAGs)) through protein-protein interaction network analysis. Logistic regression and receiver operating characteristic analyses were conducted on the selected DAGs to evaluate their diagnostic potential, with additional gene expression validation performed in brain organoids and primary neurons. A total of 145 genes were identified as oDEGs in the above two data sets, and 18 genes were selected as ooDEGs. Six DAGs (ITGB1, AXL, GFAP, NRG1, CAV1, and RHOA) were selected. The diagnostic powers of the six DAGs for AD were 0.825 (GSE84422) and 0.884 (GSE109887). Human brain organoids and primary neuronal models were used to validate the biological relevance of these findings. AXL and ITGB1 were finally selected as key genes for RTK pathway in AD and were significantly increased in AD.

Pathogens deploy effector proteins to manipulate host physiology and promote infection. YopJ family effectors are highly conserved across bacterial genera that cause crop diseases. Nucleotide-binding leucine-rich repeat receptors (NLRs) play a central role in direct or indirect recognition of effectors and trigger immune responses, including hypersensitive cell death (HR). Two NLRs, Nicotiana benthamiana homologues of Pseudomonas tomato race 1 (NbPtr1) and HOPZ-ACTIVATED RESISTANCE 1 (NbZAR1), were recently identified as independently recognising two YopJ family effectors, HopZ5 and AvrBsT/XopJ2. NbZAR1 also detects XopJ4 via the receptor-like cytoplasmic kinase XOPJ4 IMMUNITY 2 (JIM2). Here, we conducted Agrobacterium-mediated transient expression assays with 20 YopJ family effectors from five phytopathogenic bacterial genera and identified 12 YopJ family effectors that are recognised either by NbZAR1 or independently by NbZAR1 and NbPtr1. Furthermore, we show that YopJ family effector-induced HR is differentially suppressed by the deacetylase SUPPRESSOR OF AVRBST-ELICITED RESISTANCE 1, suggesting more than one mechanism for YopJ family effector recognition. This work provides the genetic basis of the recognition of YopJ family effectors in N. benthamiana and lays a foundation for the mechanistic study of NbZAR1/JIM2 and NbPtr1 mode of activation.

Malignancies of the nervous system remain a critical challenge in oncology, contributing substantially to cancer-related mortality in children and adults. Their aggressive behavior is influenced by adhesion heterogeneity of the tumor cells, as distinct patterns of cell-to-cell and cell-to-matrix interactions shape tumor organization, invasion, and resistance to conventional treatments. Here, we investigate tumor adhesion dynamics in a 3D spherical cavity culture platform that enables the rapid and high-throughput formation of uniform spheroids for parallel morphogenesis, adhesion profiling, and drug screening. The morphogenetic patterns of human neuroblastoma and patient-derived glioblastoma observed in 3D cavity culture closely correlated with tumor aggressiveness, ranging from adhesive layers in benign tumor subtype to compact spheroids in malignant cells and irregular aggregates in highly invasive, cancer stem-like counterparts. These morphologies corresponded to adhesion profiles: aggressive cells displayed elevated N-cadherin with variable integrin, while compact spheroids maintained a balance of both molecules. Blocking N-cadherin with ADH-1 disrupted spheroid integrity, and combining ADH-1 with DOX yielded synergistic cytotoxic effects in malignant phenotypes, highlighting N-cadherin upregulation in tumor cells with higher aggressiveness in 3D culture. Overall, our dynamic 3D spherical cavity culture enables reproducible 3D adhesion phenotyping, providing a simple method for malignancy assessment and supporting adhesion-targeted therapeutic strategies for personalized medicine.

Wild plant species are threatened by diverse pathogens, but disease symptoms are rarely observed in nature. This suggests that wild plants harbor valuable sources of resistance. In this study, we show that the model bacterial pathogen Pseudomonas syringae pv. tomato (Pto) DC3000 triggered defense responses in all tested accessions of a wild Solanaceae species, Solanum americanum. Pto DC3000-triggered immunity in S. americanum required a type III secretion system. We show that seven Pto DC3000 effectors (AvrPto, HopAD1, HopAM1, HopC1, HopAA1-1, HopM1, and AvrE1) triggered hypersensitive responses (HR) in S. americanum accession SP2273. Significantly, sequential deletion of the HR-triggering effectors from Pto DC3000 resulted in enhanced virulence in S. americanum. However, the well-conserved effectors, HopM1 and AvrE1, were indispensable for virulence. We conclude that the immunity triggered by multiple effectors contributes to nonhost resistance in S. americanum against P. syringae. We propose that the identification of the corresponding disease resistance genes for HopM1 and AvrE1 in S. americanum would accelerate the development of durable immunity to P. syringae pathogens in Solanaceae crops.

Background/Objectives: Fibroblast growth factor receptors (FGFRs) are frequently dysregulated in diverse cancers and represent important therapeutic targets. Here, we report the design and synthesis of a novel nucleoside-based scaffold which enables irreversible pan-FGFR inhibition as a potential anticancer strategy. Methods: A series of nucleoside analogues was synthesized and assessed through structure–activity relationship studies. Structural analyses, including X-ray co-crystallography and molecular dynamics simulations, were performed to define key determinants of potency and selectivity. Biochemical assays against FGFR1–4 proteins, cellular antiproliferative assays in HCT116 (FGFR1 amplification) and RT4 (FGFR3-TACC3) models, metabolic stability evaluations and covalent bonding confirmation were conducted to characterize representative compounds. Results: SAR studies revealed that fused aromatic substituents and 4′-thio ribose enhanced FGFR potency, whereas enantiomeric inversion of ribose reduced activity. X-ray co-crystallography further demonstrated that two hydroxyl groups form a key water-mediated hydrogen bond network, uniquely stabilizing the ligand and enhancing potency of inhibitors compared to reference compounds. The 7-methoxy-5-methylbenzo[b]thiophene scaffold and ribose moiety emerged as critical features. Compounds 13f, 19e, and 22f demonstrated potent inhibition of FGFR1-4 and dose-dependent suppression of FGFR1-mediated signaling, with strong antiproliferative activity in both FGFR-driven and wild-type cancer models. Compound 22f showed efficient irreversible covalent engagement of FGFRs, confirmed at the protein and cellular levels, and exhibited improved metabolic stability. Conclusions: Nucleoside analogues represent a privileged scaffold for covalent pan-FGFR inhibition. The findings highlight their potential as promising therapeutic candidates for targeting FGFR-driven malignancies. Future efforts will focus on further improving stability and optimizing physicochemical properties to advance these compounds toward translational development.

Hutchinson-Gilford progeria syndrome (HGPS) is a rare genetic disorder characterized by premature aging and primarily caused by the accumulation of progerin, a mutant form of lamin A. Although the effects of progerin on multiple tissues have been previously studied, its impact on brain development is not completely understood. We established cortical organoids derived from HGPS patient-induced pluripotent stem cells (iPSCs) from patients with HGPS to investigate the role of progerin in the brain. HGPS cortical organoids showed hallmarks of HGPS pathology, including elevated progerin expression and irregular nuclear morphology during early developmental stages. Additionally, we observed abnormal morphology and increased cellular senescence specifically in the rosette regions of HGPS organoids. This senescence appeared to interfere with normal neuronal differentiation, resulting in a significant reduction in mature neuron development and synapse formation in HGPS cortical organoids. Transcriptome profiling of HGPS cortical organoids revealed the downregulation of key genes related to neural development and synapse formation, with these changes persisting over time, potentially contributing to impaired neuronal differentiation and maturation. These findings suggest the role of progerin in early neural development and establish cortical organoids as a model for studying HGPS-related brain development.

Glutamine synthetase (GS) in astrocytes regulates glutamatergic neurotransmission by maintaining glutamate clearance in the brain. This study determined that GS in astrocytes of the caudate and putamen (CPu) regulates locomotor sensitization after repeated nicotine exposure. Nicotine increased phosphorylated c-Jun N-terminal kinase (pJNK) by stimulating α7 nicotinic acetylcholine receptors in cultured glioma C6 cells and primary astrocytes in a Ca2+-dependent manner. Active JNK phosphorylated metabotropic glutamate receptor 1a (mGluR1a) at the carboxyl terminus of glutathione S-transferase-tagged mGluR1a in vitro. Interference with the pJNK-mGluR1a interaction using the inhibitory peptide, Tat-mGluR1a-i (10 μmol/L), decreased the nicotine-induced increase in GS activity in glioma C6 cells and primary astrocytes. Similar results were obtained by bilateral intra-CPu infusion of the inhibitory peptide (2 nmol/side). Inhibition of GS activity by bilateral intra-CPu infusion of methionine sulfoximine (50 nmol/side) decreased the repeated nicotine-induced increase in locomotor activity. These findings suggest that astrocytes in the CPu upregulate locomotor sensitization by activating GS via the pJNK-mGluR1a interaction, which is linked to α7 nicotinic acetylcholine receptors in response to nicotine.

Over the past two decades, the "hallmarks of cancer" have revolutionized cancer research and highlighted the crucial roles of inflammation and immunity. Protumorigenic inflammation promotes cancer development along with inhibition of antitumor immunity, shaping the tumor microenvironment (TME) toward a tumor-permissive state and further enhancing the malignant potential of cancer cells. This immunosuppressive TME allows tumors to evade immunosurveillance. Thus, understanding the complex interplay between tumors and the immune system within the TME has become pivotal, especially with the advent of immunotherapy. Although immunotherapy has achieved notable success in many malignancies, primary liver cancer, particularly hepatocellular carcinoma, presents unique challenges. The hepatic immunosuppressive environment poses obstacles to the effectiveness of immunotherapy, along with high mortality rates and limited treatment options for patients with liver cancer. In this review, we discuss current understanding of the complex immune-mediated mechanisms underlying liver neoplasms, focusing on hepatocellular carcinoma and liver metastases. We describe the molecular and cellular heterogeneity within the TME, highlighting how this presents unique challenges and opportunities for immunotherapy in liver cancers. By unraveling the immune landscape of liver neoplasms, this review aims to contribute to the development of more effective therapeutic interventions, ultimately improving clinical outcomes for patients with liver cancer.

This study aims to examine intra-household gender dynamics in response to a nutrition intervention (maternal Behaviour Change Communication (BCC), paternal BCC, and food vouchers) aimed at improving IYCF practices using qualitative methods. Participants were drawn from a subset of households enrolled in a larger cluster - randomized controlled trial (RCT) conducted in rural Ethiopia. A total of 40 participants (20 mother-father pairs) from intervention and control households were interviewed separately to explore intra-dyadic beliefs and household decision-making. Furthermore, this study explores plausible mechanisms behind the main RCT finding that greater father involvement, while increasing knowledge, did not consistently improve infant and young child feeding (IYCF) outcomes. We find that BCC mothers had the tendency to adopt more gender-equal beliefs, particularly regarding men's roles in childcare and household chores, whereas fathers were slower to shift their views. This may have led to increased discordance in gender norms within BCC households. By contrast, control couples often retained traditional views, but showed more intra-couple alignment. Unexpectedly, control fathers were sometimes more progressive than control mothers, potentially due to higher education levels. We also find increased paternal engagement occasionally introduced conflicting priorities, as fathers asserted authority over household spending - sometimes at the expense of child-focused nutrition. These findings underscore the complexity of engaging fathers in nutrition interventions and point to the need for strategies that address underlying gender norms and decision-making dynamics within households.

This study investigates the role of 25-hydroxycholesterol (25HC), a metabolite produced by cholesterol hydroxylase encoded by the Ch25h gene, in modulating microglial function and its potential implications in Alzheimer’s disease (AD) pathology. We demonstrated that 25HC impairs microglial surveillance, reduces phagocytic capacity, and increases the production of pro-inflammatory cytokines. In vivo two-photon microscopy revealed that 25HC administration diminishes microglial response to brain lesions, while flow cytometry confirmed reduced phagocytosis in both in vivo and in vitro models. Additionally, amyloid-beta (Aβ) was shown to upregulate Ch25h expression and elevate 25HC levels in microglia, exacerbating these functional impairments. Mechanistically, 25HC was found to enhance cholesterol esterification, disrupt cell membrane dynamics, and further reduce microglial mobility and phagocytosis. Treatment with Avasimibe, a cholesterol esterification inhibitor, restored membrane dynamics and microglial function, leading to attenuated AD pathology in a 5XFAD mouse model. These findings suggest that 25HC-induced changes in microglial function contribute to AD progression, and targeting cholesterol metabolism could offer therapeutic potential.

The impact of microplastics (MPs) on the metabolic functions of the liver is currently unclear and not completely understood. To investigate the effects of the administration of MPs on the hepatic metabolism of normal and obese mice, alterations in the lipid, glucose (Glu), and amino acid regulation pathways were analyzed in the liver and adipose tissues of C57BL/6Korl (wild type, WT) or C57BL/6-Lepem1hwl/Korl mice (leptin knockout, Lep KO) orally administered polystyrene (PS) MPs for 9 weeks. Significant alterations in the lipid accumulation, adipogenesis, lipogenesis, and lipolysis pathways were detected in the liver tissue of MP-treated WT and Lep KO mice compared to the vehicle-treated group. These alterations in their liver tissues were accompanied by an upregulation of the serum lipid profile, as well as alterations in the adipogenesis, lipogenesis, and lipolysis pathways in the adipose tissues of MP-treated WT and Lep KO mice. Specifically, the level of leptin was increased in the adipose tissues of MP-treated WT mice without any change in their food intake. Also, MP-induced disruptions in the glycogenolysis, Glu transporter type 4 (GLUT4)-5′ AMP-activated protein kinase (AMPK) signaling pathway, levels of lipid intermediates, and the insulin resistance of the liver tissues of WT and Lep KO mice were observed. Furthermore, the levels of seven endogenous metabolites were remarkably changed in the serum of WT and Lep KO mice after MP administrations. Finally, the impact of the MP administration observed in both types of mice was further verified in differentiated 3T3-L1 adipocytes and HepG2 cells. Thus, these results suggest that the oral administration of MPs for 9 weeks may be associated with the disruption of lipid, Glu, and amino acid metabolism in the liver tissue of obese WT and Lep KO mice.

The impact of microplastics (MPs) on the metabolic functions of the liver is currently unclear and not completely understood. To investigate the effects of the administration of MPs on the hepatic metabolism of normal and obese mice, alterations in the lipid, glucose (Glu), and amino acid regulation pathways were analyzed in the liver and adipose tissues of C57BL/6Korl (wild type, WT) or C57BL/6-Lepem1hwl/Korl mice (leptin knockout, Lep KO) orally administered polystyrene (PS) MPs for 9 weeks. Significant alterations in the lipid accumulation, adipogenesis, lipogenesis, and lipolysis pathways were detected in the liver tissue of MP-treated WT and Lep KO mice compared to the vehicle-treated group. These alterations in their liver tissues were accompanied by an upregulation of the serum lipid profile, as well as alterations in the adipogenesis, lipogenesis, and lipolysis pathways in the adipose tissues of MP-treated WT and Lep KO mice. Specifically, the level of leptin was increased in the adipose tissues of MP-treated WT mice without any change in their food intake. Also, MP-induced disruptions in the glycogenolysis, Glu transporter type 4 (GLUT4)-5' AMP-activated protein kinase (AMPK) signaling pathway, levels of lipid intermediates, and the insulin resistance of the liver tissues of WT and Lep KO mice were observed. Furthermore, the levels of seven endogenous metabolites were remarkably changed in the serum of WT and Lep KO mice after MP administrations. Finally, the impact of the MP administration observed in both types of mice was further verified in differentiated 3T3-L1 adipocytes and HepG2 cells. Thus, these results suggest that the oral administration of MPs for 9 weeks may be associated with the disruption of lipid, Glu, and amino acid metabolism in the liver tissue of obese WT and Lep KO mice.

The impact of microplastics (MPs) on the metabolic functions of the liver is currently unclear and not completely understood. To investigate the effects of the administration of MPs on the hepatic metabolism of normal and obese mice, alterations in the lipid, glucose (Glu), and amino acid regulation pathways were analyzed in the liver and adipose tissues of C57BL/6Korl (wild type, WT) or C57BL/6-Lepem1hwl/Korl mice (leptin knockout, Lep KO) orally administered polystyrene (PS) MPs for 9 weeks. Significant alterations in the lipid accumulation, adipogenesis, lipogenesis, and lipolysis pathways were detected in the liver tissue of MP-treated WT and Lep KO mice compared to the vehicle-treated group. These alterations in their liver tissues were accompanied by an upregulation of the serum lipid profile, as well as alterations in the adipogenesis, lipogenesis, and lipolysis pathways in the adipose tissues of MP-treated WT and Lep KO mice. Specifically, the level of leptin was increased in the adipose tissues of MP-treated WT mice without any change in their food intake. Also, MP-induced disruptions in the glycogenolysis, Glu transporter type 4 (GLUT4)-5′ AMP-activated protein kinase (AMPK) signaling pathway, levels of lipid intermediates, and the insulin resistance of the liver tissues of WT and Lep KO mice were observed. Furthermore, the levels of seven endogenous metabolites were remarkably changed in the serum of WT and Lep KO mice after MP administrations. Finally, the impact of the MP administration observed in both types of mice was further verified in differentiated 3T3-L1 adipocytes and HepG2 cells. Thus, these results suggest that the oral administration of MPs for 9 weeks may be associated with the disruption of lipid, Glu, and amino acid metabolism in the liver tissue of obese WT and Lep KO mice.

BACKGROUND:Macrophages engineered with chimeric antigen receptors (CAR) are suitable for immunotherapy based on their immunomodulatory activity and ability to infiltrate solid tumours. However, the production and application of genetically edited, highly effective, and mass-produced CAR-modified macrophages (CAR-Ms) are challenging. METHODS:Here, we used homology-independent targeted insertion (HITI) for site-directed CAR integration into the safe-harbour region of human pluripotent stem cells (hPSCs). This approach, together with a simple differentiation protocol, produced stable and highly effective CAR-Ms without heterogeneity. FINDINGS:These engineered cells phagocytosed cancer cells, leading to significant inhibition of cancer-cell proliferation in vitro and in vivo. Furthermore, the engineered CARs, which incorporated a combination of CD3ζ and Megf10 (referred to as FRP5Mζ), markedly enhanced the antitumour effect of CAR-Ms by promoting M1, but not M2, polarisation. FRP5Mζ promoted M1 polarisation via nuclear factor kappa B (NF-κB), ERK, and STAT1 signalling, and concurrently inhibited STAT3 signalling even under M2 conditions. These features of CAR-Ms modulated the tumour microenvironment by activating inflammatory signalling, inducing M1 polarisation of bystander non-CAR macrophages, and enhancing the infiltration of T cells in cancer spheroids. INTERPRETATION:Our findings suggest that CAR-Ms have promise as immunotherapeutics. In conclusion, the guided insertion of CAR containing CD3ζ and Megf10 domains is an effective strategy for the immunotherapy of solid tumours. FUNDING:This work was supported by KRIBB Research Initiative Program Grant (KGM4562431, KGM5282423) and a Korean Fund for Regenerative Medicine (KFRM) grant funded by the Korean government (Ministry of Science and ICT,Ministry of Health and Welfare) (22A0304L1-01).

Owing to their superior hydrophilicity, zwitterionic molecules are known to form a strong hydration layer serving as an effective antifouling barrier. Despite their widespread use, the immobilization of zwitterionic molecules on various surfaces has posed a considerable challenge. Herein, we designed zwitterionic polyethers functionalized with mussel-inspired catechol moieties, which can be applied as versatile coatings with enhanced antifouling properties for biomedical surfaces. A series of block polyethers were synthesized via sequential anionic ring-opening polymerization of catechol-acetonide glycidyl ether and N,N-diisopropyl ethanolamine glycidyl ether, followed by post-polymerization modification to afford the desired zwitterionic brushes. The versatile surface adsorption and superior antifouling effects of the synthesized polyether brushes were evaluated using atomic force microscopy, quartz crystal microbalance with dissipation, and bioassay on the biomedical device surfaces. This study highlights the superior antifouling properties of zwitterionic polyether brushes, with potential applications in biomedical surfaces. Zwitterionic polyether brushes bearing mussel-inspired catechol moieties are developed for versatile coatings with improved antifouling properties on biomedical surfaces.

Topical treatment offers a viable alternative for melanoma patients incompatible with surgical interventions. Herein, the study develops a skin-penetrating peptide (SPP)-based peptide drone (PD) for the transdermal delivery of antitumor proteins. To achieve cost-effective therapeutics, Concanavalin A (ConA), which can be prepared through an extraction method, is loaded onto PDs. Compared with free proteins, ConA-PD complexes (CPCs) demonstrate significantly greater skin permeation. Moreover, the CPCs exhibit potent anticancer activity both in vitro and in vivo, which substantially surpass the efficacy of the Aldara (imiquimod) cream. The CPC-treated mouse skin remains clear, whereas Aldara cream induces side effects, including psoriasis-like skin inflammation. The antimetastatic activity of the PD is identified as another advantage of CPC. The use of ConA as an anticancer agent has been significantly limited due to its high hepatotoxicity; however, the transdermal delivery strategy minimizes its hepatic absorption and, in turn, results in negligible hepatotoxicity. These results demonstrate the potential of the CPC as a novel therapeutic agent or an adjunct to other modalities for melanoma treatment, overcoming the limitations of existing materials while exhibiting superior efficacy. A skin-penetrating peptide-based peptide drone (PD) delivers the antitumoral but hepatotoxic protein, ConA, beyond the skin barrier, thereby exhibiting significantly superior antitumor (melanoma) efficacy to Aldara (imiquimod) cream. This transdermal strategy minimizes hepatic absorption of the proteins, resulting in negligible hepatotoxicity. Furthermore, the PD-ConA complex avoids inducing the side effects associated with Aldara cream, including psoriasis-like skin inflammation. image