Dual cascade signal amplification lights up G4 dimers for fluorescent detection of DNA repair enzyme FEN1.

Single-stranded DNA-binding protein gp32 is essential for DNA replication and widely used in isothermal amplification techniques such as recombinase polymerase amplification (RPA). However, its inherent thermal instability and reaction condition sensitivity restrict its broader applications. Here we develop the Fermentation Integrated Rational Engineering (FIRE-3S) strategy to enhance the functional properties of gp32 through molecular dynamics (MD) simulations, structure-guided fermentation tuning and stability-focused rational design. Starting from a disulfide-bonded mutant (V62C/T80C) that showed enhanced RPA performance but reduced thermostability, MD simulations revealed local structural disruption near the Zn2+-binding motif. Then, structure-guided fermentation tuning was applied, which restored structural integrity and enhanced DNA-binding affinity. Building on 2S-step, a computationally guided clique-based design pipeline rapidly identified a multisite mutant, gp32-M1, with elevated thermostability (ΔTm = +6.03 °C), robust ssDNA-binding, and strong inhibitor tolerance─maintaining >65% activity under 4% isopropanol or ethanol. This work establishes a systematic framework for engineering ssDNA-binding proteins with enhanced biostability and environmental resilience for molecular diagnostics.

DNA polymerase-β (DNA pol-β) plays a critical role in β-amyloid-induced neurodegeneration by mediating aberrant DNA replication in postmitotic neurons. In previous work, we demonstrated that 5-methoxyflavone inhibits DNA pol-β, though computational analyses suggested potential binding to the primase p58 subunit. Through molecular modeling, here, we designed (S)-2-cyclohexyl-5-methoxy-2H-chromene (S-chromene), a novel flavone-derived inhibitor exhibiting strong electrostatic complementarity with DNA pol-β but weak interaction with primase p58, suggesting enhanced selectivity. (R)-2-cyclohexyl-5-methoxy-2H-chromene (R-chromene) exhibited indistinguishable binding properties from S-chromene. The compound was obtained as a racemic mixture (chromene). Since the separated enantiomers were unstable, all biological assays used the racemate. DNA polymerase activity assay confirmed that chromene inhibited selectively DNA pol-β without affecting the primase/DNA pol-α complex activity. Also, the compound amplified methylmethanesulfonate toxicity in wild-type but not DNA pol-β-null fibroblasts, validating target-engagement. In cultured neurons, chromene effectively prevented β-amyloid-induced DNA replication and apoptosis. Ours is the first demonstration of a chromene acting as a selective DNA pol-β inhibitor endowed with a unique mechanism of neuroprotection.

Stepwise DNA damage and repair mechanisms at replication forks in response to topoisomerase I inhibition.

Lipid metabolism disruption as a conserved early response to Benzophenone-1: Pathway sensitivity profiling via dose-dependent yeast functional genomics platform.

The impact of alterations in lamin A on genome integrity.

Cardiomyocytes undergo proliferation, differentiation, and hypertrophy during fetal development. Current techniques struggle to distinguish cardiomyocyte proliferation from alternative cell fates. In this study, we combined flow cytometry measures of cardiomyocyte ploidy (DAPI) and in vivo DNA replication (EdU) over a 24-h period to evaluate the trajectories of cardiomyocytes from normally growing control fetal sheep and fetuses affected by placental insufficiency and fetal growth restriction (FGR) at 0.9 gestation. We categorized ∼100,000 cardiomyocytes from the left and right ventricles (LV and RV) of each animal as proliferating (2C EdU+), differentiated (4C EdU-), or endoreplicating and polyploid (6C+ EdU+). Compared with controls, FGR hearts had 25%-50% fewer cardiomyocytes that replicated DNA (EdU+) (LV: P = 0.02, RV: P = 0.002). The fraction of proliferating cardiomyocytes, indicated by the population of newly synthesized 2C EdU+ daughter cells, was ∼20% lower in FGR fetuses (LV: P = 0.006, RV: P = 0.02). Instead, the percentage of endoreplicating cardiomyocytes (6C+ EdU+) in FGR hearts was double that of controls (LV: P = 0.004, RV: P = 0.002). Although total EdU+ was not a strong predictor of cardiac growth, LV and RV mass correlated positively with the percentage of 2C EdU+ cardiomyocytes and negatively with 6C+ EdU+ cardiomyocytes across all fetuses. LV mass also correlated positively with the percentage of differentiated cardiomyocytes (4C EdU-), which was lower in FGR hearts compared with controls (P = 0.008). This study is the first to characterize cardiomyocyte fate following DNA replication in fetal sheep. Our findings suggest that FGR cardiomyocytes differentially prioritize their cycling capacity in favor of polyploidization instead of proliferation.NEW & NOTEWORTHY Knowledge of cardiac development has been limited by available methodologies. We used a novel flow cytometry approach to measure DNA replication in utero and distinguish between cardiomyocyte proliferation, differentiation, and endoreplication in growth-restricted (FGR) and normally growing fetal sheep. FGR cardiomyocytes have lower proliferation rates but increased endoreplication compared with controls. Endoreplication and polyploidy are negatively correlated with ventricular mass. Our findings provide insight into fetal cardiac development and how cardiomyocyte fate is altered by FGR.

The protein architecture of Mycobacterium tuberculosis (Mtb) demonstrates remarkable complexity and adaptability, emblematic of its evolutionary refinement as a highly successful pathogen. The Mtb proteome can be broadly classified into four principal categories: core, accessory, transcriptionally plastic, and uncharacterized proteins. Core proteins are highly conserved across all Mtb strains and essential for fundamental cellular functions and bacterial viability; they form the structural and metabolic foundation required for critical processes such as DNA replication, transcription, and cell wall biosynthesis. Conversely, accessory proteins exhibit considerable variability among strains, endowing Mtb with strain-specific traits including virulence, environmental adaptation, and antibiotic resistance. These proteins are vital for enabling the pathogen to thrive in diverse ecological niches and to overcome selective pressures imposed by environmental factors and antimicrobial agents. Distinguished not by strain distribution but by regulatory dynamics, transcriptionally plastic proteins exhibit differential expression in response to environmental changes and host-derived cues, allowing Mtb to modulate its physiological state during infection rapidly. This regulatory flexibility supports the pathogen's ability to enter dormancy, mount stress responses, and transition between metabolic states. A substantial portion of the Mtb proteome remains uncharacterized or annotated as hypothetical, with functions yet to be elucidated. Nevertheless, recent advances in integrative bioinformatics and experimental proteomics have begun to clarify the roles of many such proteins, revealing novel contributions to bacterial survival, pathogenicity, and immune evasion. The complex interplay among these protein categories illustrates a highly sophisticated regulatory network that governs Mtb's growth, dormancy, stress adaptation, and persistence. This dynamic and adaptable protein architecture is fundamental to the bacterium's capacity to endure hostile host environments, evade immune surveillance, and establish chronic infections. Consequently, a comprehensive understanding of the composition, regulation, and functional plasticity of these protein classes is imperative. Such knowledge will drive the development of innovative diagnostics, next-generation vaccines, and targeted therapeutics, ultimately advancing more effective strategies for tuberculosis control and eradication.

Fertility restoration in cytoplasmic male sterile tomato via knockout of either DNA polymerase I or Topoisomerase I, two nuclear-encoded organellar DNA replication genes.

Endotoxin lipopolysaccharide challenge triggers gut microbiota dysbiosis and host immune remodeling in the tiger grouper Epinephelus fuscoguttatus.

KMT2C mutations in breast cancer: molecular mechanisms of cancer promotion, clinical significance, and potential as biomarkers.

Integrative analysis of RNA modification-related gene PUS7 in diagnosis, prognosis, and tumor microenvironment of hepatocellular carcinoma.

Antiparasitic agent emodepside exerts cytotoxicity in human corneal stromal cells by blocking the interaction between mini-chromosome maintenance 6 protein (MCM6) and chromatin licensing and DNA replication factor 1 (CDT1).

Multi-omics integrative analysis elucidates the molecular characteristics of SMARCA4-deficient lung adenocarcinoma and the derived therapeutic options.

Exostosin glycosyltransferase 1 (EXT1) and exostosin glycosyltransferase 2 (EXT2) catalyze heparan sulfate chain elongation and are increasingly implicated in cancer biology, but their roles in gliomas remain incompletely defined. Here, we performed an integrative multi-omics analysis to dissect the transcriptional, epigenetic, and microenvironmental landscape of EXT1 and EXT2 across gliomas. Bulk transcriptomic data from The Cancer Genome Atlas (TCGA) and the Chinese Glioma Genome Atlas (CGGA) revealed that both EXT1 and EXT2 are upregulated in high-grade gliomas and associate with adverse survival, with EXT1 showing the strongest and most consistent prognostic impact. Gene set enrichment analysis (GSEA) and gene set variation analysis (GSVA) indicated that EXT1-high tumors are enriched for DNA damage and replication stress programs, cell cycle progression, inflammatory response, and stromal activation pathways, whereas EXT2 expression is preferentially linked to extracellular matrix remodeling, cytoskeletal organization and angiogenesis-related signaling. Single-cell RNA sequencing and Immune deconvolution using Cell-type Identification By Estimating Relative Subsets Of RNA Transcripts (CIBERSORT) and Estimation of STromal and Immune cells in MAlignant Tumor tissues using Expression data (ESTIMATE) showed that EXT1 correlates with increased stromal and immune scores, and reduced cytotoxic T cell signatures, consistent with an immunosuppressive tumor microenvironment. EXT2 expression is enriched in gliomas with pronounced vascular and mesenchymal features, supporting a complementary role in invasive growth and tissue remodeling. Immunohistochemistry on a glioma tissue microarray validated the upregulation of EXT1 protein in high-grade tumors. The study findings identified EXT1 as a central glycosylation-linked regulator of replication stress tolerance and immune remodeling in gliomas, and suggest that EXT2 contributes to extracellular matrix and cytoskeletal reprogramming. The exostosin axis represents a promising source of prognostic biomarkers and potential therapeutic targets in glioma.

Recent progress in developing ATR inhibitors as anticancer agents.

Discovery of coumarin-conjugated hydrazonoindoles as new type of potential antibacterial agents.

Effect of Nitrosomonas europaea on Chlorella vulgaris in bio-hydrogels for startup of microalgal-bacterial granular sludge: Performance and microscopic mechanism.

Lung cancer remains the leading cause of cancer mortality. The AP-1 adaptor complex, including AP1AR, AP1S1, AP1S2, AP1S3, AP1M1, AP1M2, AP1B1, and AP1G1, functions as a conserved hub of vesicular trafficking, selecting cargo and coordinating clathrin-mediated transport. By shaping receptor recycling, membrane composition, and signal duration, AP-1 influences core cancer phenotypes such as proliferation, migration, and therapy response. However, the family-level role of AP-1 adaptors in lung cancer is incompletely defined. We systematically profiled all eight AP-1 adaptor genes using multi-omics datasets, survival resources, pharmacogenomic panels, Human Protein Atlas data, pathway enrichment, and single-cell RNA sequencing with cell-cell communication modeling. AP1AR was consistently upregulated in lung adenocarcinoma and independently associated with poorer overall survival. It was linked to cell-cycle progression, DNA replication checkpoints, hypoxia, and epithelial-to-mesenchymal transition (EMT). At single cell resolution, AP1AR also regulate malignant epithelial and fibroblast cell types. Pseudotime analyses revealed progressive activation along proliferative and EMT axes, and CellChat modeling indicated enhanced stromal and epithelial signaling. AP1S3 and AP1S1 showed complementary roles, associated with oncogenic/inflammatory signaling and immune-metabolic programs, respectively. These findings identify AP1AR as a clinically relevant biomarker and highlight AP-1 adaptor biology as an underexplored contributor to lung adenocarcinoma progression and therapeutic stratification.

Roles of FEN1 in tumor biology: Mechanisms, therapeutic implications, and emerging strategies.