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PubMed2026

Human periodontal ligament stem cells promote oral ulcer healing in rats through modulation of TGF-β1/smad signaling.

BACKGROUND: Oral ulcers (OU) often present with prolonged healing, recurrent episodes, and scar formation, posing challenges for clinical management. Human periodontal ligament stem cells (hPDLSCs) have shown potential in oral tissue repair, but further research is needed to clarify their mechanism of action in OU healing. This study aims to elucidate the molecular mechanisms by which hPDLSCs promote oral ulcer healing. METHOD: To identify key regulatory genes, the OU-associated microarray dataset GSE37265 was integrated with hPDLSC genomic data for differential expression analysis. Subsequently, Weighted Gene Co-expression Network Analysis (WGCNA) was used to identify functional modules associated with OU healing. In vivo, hPDLSCs were locally administered into a rat ulcer model, and therapeutic efficacy was assessed by ulcer closure rates and histological evaluation (HE and Masson's trichrome staining). Furthermore, RNA-sequencing (RNA-seq) was performed on oral mucosal tissues to delineate the underlying molecular landscape and critical signaling pathways. The involvement of the TGF-β signaling pathway was confirmed by real-time quantitative PCR (RT-qPCR) and Western blotting (WB) analyses. RESULTS: Bioinformatics analysis identified 92 key genes in hPDLSCs-mediated treatment of OU, highlighting the central role of the TGF-β1/Smad pathway. As shown by the animal studies, hPDLSCs therapy increased the healing rate to 97% by day 8 (vs. 70% in the model). Furthermore, the therapy significantly reduced inflammatory cell infiltration and abnormal collagen deposition while promoting regular collagen arrangement. Transcriptomic and molecular experiments further showed that hPDLSCs simultaneously inhibit TGF-β1/Smad and extracellular signal-regulated kinase (ERK) signaling pathways, thereby alleviating inflammatory responses and suppressing mucosal fibrosis. CONCLUSION: In this study, we reveal a novel role for hPDLSCs in promoting oral ulcer healing. The findings indicate that hPDLSCs suppress inflammation and fibrosis via the TGF-β1/Smad pathway, offering a promising therapeutic strategy for OU and other fibrotic conditions.

TGF-β1↗

PubMed2026

Common Molecular Mechanisms and Candidate Drug Targets in Type 2 Diabetes Mellitus and Atherosclerotic Cardiovascular Disease.

This study examined the mechanisms underlying the comorbidity between type 2 diabetes mellitus (T2DM) and atherosclerotic cardiovascular disease (ASCVD), while identifying potential therapeutic targets. Common differentially expressed genes (C-DEGs) between T2DM and ASCVD were extracted from the GSE78721 and GSE12288 datasets. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analyses, protein-protein interaction (PPI) network construction, hub gene identification, and Drug-Gene Interaction Database (DGIdb) analysis were conducted. The association between hub C-DEGs and immune-infiltrating cells was analyzed using the CIBERSORT method. Expression levels of hub C-DEGs were quantified through qRT-PCR and Western blot analyses. A total of 32 C-DEGs were identified, comprising 20 upregulated and 12 downregulated genes. C-DEGs were predominantly enriched in key pathways, including viral myocarditis, arrhythmogenic right ventricular cardiomyopathy, hypertrophic cardiomyopathy, and dilated cardiomyopathy. PPI analysis revealed 29 nodes and 39 edges, leading to the identification of eight hub C-DEGs (HSP90B1, PLAU, SLPI, TOP3A, NCF4, PRF1, TUBA1C, and CS) across both datasets. Furthermore, hub C-DEGs (TOP3A, SLPI, NCF4, PRF1, and PLAU) demonstrated significant correlations with immune-infiltrating cell levels. Drugs specifically targeting these hub C-DEGs present promising candidates for the treatment of T2DM and ASCVD. Additionally, the expression of hub C-DEGs at both mRNA and protein levels was validated in patients with T2DM and ASCVD. An integrated bioinformatics analysis facilitated the screening of candidate therapeutic targets, mechanisms, and drugs for T2DM and ASCVD, offering new insights into molecular therapies for these conditions.

Diabetes Mellitus, Type 2↗

PubMed2026

Longitudinal surveillance of antibiotic resistance and virulence evolution in Clostridioides difficile: a 4-year retrospective study of hospitalized patients in a tertiary hospital in China.

UNLABELLED: Clostridioides difficile (C. difficile) is the primary pathogen responsible for nosocomial infectious diarrhea and pseudomembranous colitis. In China, metronidazole and vancomycin are the preferred treatments for C. difficile infection (CDI). This study aimed to investigate the evolution of vancomycin (VA) and metronidazole (MTZ) resistance, as well as the longitudinal changes in virulence over time, using next-generation sequencing, drug susceptibility tests, and analysis of resistance and virulence genes. Additionally, we monitored the emergence of the highly virulent C. difficile strain RT027 and the spread and potential outbreak of C. difficile in the hospital setting. A random stratified sampling method was used to select 114 fecal samples from inpatients at Affiliated Hangzhou First People's Hospital, School of Medicine, Westlake University, between 2021 and 2024. Clinical data from the enrolled patients were also collected. We conducted antigen and toxin protein detection for C. difficile, strain isolation and identification, drug sensitivity tests, whole genome sequencing, and bioinformatics analysis. This included comparisons of drug resistance genes, detection of toxin genes, and the construction of phylogenetic trees based on pan-genome analysis to investigate the resistance and toxin gene variations in C. difficile. Among the 114 samples collected from Affiliated Hangzhou First People's Hospital, School of Medicine, Westlake University, no vancomycin- or metronidazole-resistant strains were identified. However, the average minimum inhibitory concentration (MIC) of C. difficile to vancomycin increased annually (H = 33.208, P < 0.05). The average MIC of C. difficile to metronidazole was highest in 2022 but decreased in 2023 and 2024 (H = 41.990, P < 0.05). Notably, in 2024, one C. difficile strain exhibited an MIC for metronidazole at the resistance threshold (2.00 μg/mL). Further Spearman correlation analysis of the strain years with drug sensitivity results revealed a positive correlation between strain years and the MIC levels of vancomycin and metronidazole (r = 0.528, P < 0.05; r = 0.377, P < 0.05). The proportion of toxin-producing strains increased annually, with 100% of strains in 2024 producing toxins, representing the highest proportion compared to the previous three years (X² =11.75, P < 0.05). Both vancomycin and metronidazole remain effective for the treatment of CDI in clinical practice. However, the sensitivity of C. difficile to these two drugs is gradually decreasing, and the rate of toxin gene carriage is also rising in clinical cases. No hospital outbreaks of C. difficile infections were identified in this study. IMPORTANCE: Clostridioides difficile has developed resistance to multiple antibiotics, including cephalosporins, clindamycin, and fluoroquinolones. This has exacerbated the global antibiotic resistance crisis. In China, according to current treatment guidelines, vancomycin and metronidazole are the preferred first-line drugs for treating C. difficile infections. However, there are reports indicating the emergence of new resistance to both vancomycin and metronidazole. Although there is extensive research on the long-term antibiotic resistance of C. difficile abroad, research on the continuous monitoring of antibiotic resistance and potential outbreaks of C. difficile in China is relatively limited. To fill this gap, we studied positive C. difficile strains from a tertiary general hospital in China. Through Next-Generation Sequencing (NGS), drug sensitivity testing, and analysis of drug resistance and virulence genes, we revealed the evolution of C. difficile's resistance to vancomycin and metronidazole, as well as changes in virulence, and monitored the spread within the hospital and potential outbreaks of C. difficile.

Humans↗

PubMed2026

Exome sequencing and large-scale analysis of electronic medical record-linked biobank data identify candidate deafness genes.

INTRODUCTION: Rapid advances in whole-exome sequencing (WES) have enabled large-scale detection of pathogenic variants. Although hundreds of genes are implicated in hearing loss, up to half of inherited cases remain unsolved, limiting eligibility for gene therapy trials that require genetic diagnosis. Biobanks and electronic medical records (EMRs) offer opportunities to integrate genomic and clinical data at scale and expand the spectrum of hearing loss genes. Despite clinical value, EMRs often lack key information such as inheritance patterns, posing challenges for accurate interpretation. METHODS: WES was performed on DNA samples from 1038 hearing-impaired patients enrolled in the Maccabi Research and Innovation Center Tipa Biobank. Clinical data were extracted from EMRs. Audiograms were available for all cases, although data on age of onset, family history and mode of inheritance were mostly unavailable. We applied a scalable bioinformatics analysis strategy for high-throughput annotation, filtering and prioritisation of WES variants across more than 1000 patients, designed to accommodate incomplete and heterogeneous clinical records. RESULTS: Using this approach, 15% of cases were solved or potentially solved through known or novel variants in established deafness genes. Homozygous variants in novel candidate genes were identified in 3% of cases. Functional characterisation was performed for promising candidate genes to validate their role in the ear. CONCLUSION: These findings demonstrate that WES can determine disease aetiology in large, genetically heterogeneous populations, even in the context of incomplete clinical data. This approach supports large-scale genetic screening and provides a framework for identifying patients who may benefit from emerging gene-based therapies.

Genetic Testing↗

PubMed2027

Global Genomic Surveillance.

Global genomic surveillance has emerged as a foundational pillar of public health in the twenty-first century, enabling real-time tracking of pathogen evolution and informing outbreak response. This chapter examines the strategic architecture of global genomic surveillance, focusing on its application to arboviruses such as chikungunya virus (CHIKV). It explores the integration of genomic data with epidemiological, clinical, and environmental information within a One Health framework, while addressing critical challenges in governance, equity, and interoperability. The discussion covers the entire genomic surveillance workflow, from sample collection and sequencing to bioinformatic analysis and phylogenetic inference, and highlights the transformative role of artificial intelligence (AI) in predictive surveillance. By analyzing global initiatives, operational barriers, and emerging technologies, this chapter underscores the necessity of sustainable, equitable, and interoperable genomic systems to proactively address current and future infectious disease threats.

Humans↗

PubMed2025

Dual RNA isolation from blood: an optimized protocol for host and bacterial RNA purification for dual RNA-sequencing analysis in whole blood sepsis samples.

Dual RNA-sequencing (dual RNA-seq) holds significant promise for deciphering bacterial virulence mechanisms during systemic infections. However, its application in sepsis research is hindered by technical challenges, including a low bacterial burden in blood and limited sample volumes and RNA yield from vulnerable populations, such as neonates. We developed an optimized protocol [dual RNA isolation from blood (DRIB)] for simultaneous stabilization, isolation and purification of high-quality host leukocyte and bacterial RNA from low-volume whole blood samples (0.5 ml). This protocol is compatible with clinical sample collection workflows and high-throughput RNA sequencing. The feasibility of DRIB for dual RNA-seq was validated using a pilot cohort of clinical adult sepsis samples, enabling the investigation of host-bacterial gene expression during sepsis. The DRIB protocol yielded 2.10-6.91 µg of total RNA per clinical sample in our pilot cohort. Dual-species ribosomal RNA (rRNA) depletion and RNA-seq generated 16.6-24.8 million filtered reads per sample, with 63±7% of reads uniquely mapped to host or bacterial sequences. Host genes accounted for 51-68% (8.4-10.9 million) reads, while 0.5-6.7% (79,496-789,808 reads) mapped to bacterial genomes. Bioinformatic analysis revealed that both shared and individual transcriptional patterns were identified in host and bacterial responses, including pathways related to immune metabolism and metal-ion binding. Our optimized DRIB protocol and RNA-seq pipeline effectively captured both host and bacterial RNA transcription in clinical sepsis samples. Expanding this approach to larger cohorts and varying disease timepoints will provide crucial new insights into host-bacterial gene co-expression dynamics in sepsis progression and outcomes.

Humans↗

PubMed2026

PDZ-binding kinase promotes ovarian cancer cell proliferation and invasion via CCNB1 regulation.

BACKGROUND: Ovarian cancer is one of the most lethal gynecological malignancies, characterized by late diagnosis, frequent recurrence, and high mortality. PDZ-binding kinase (PBK), a serine/threonine kinase of the mitogen-activated protein kinase kinase (MAPKK) family, has been implicated in the tumorigenesis of multiple cancers, yet its role in ovarian cancer remains incompletely characterized. This study aimed to investigate the effect of PBK on the proliferation and invasion of ovarian cancer cells. METHODS: The expression of PBK and cyclin B1 (CCNB1) in normal ovarian tissues and ovarian cancer tissues was analyzed using online databases including Gene Expression Profiling Interactive Analysis 2 (GEPIA2), Clinical Proteomic Tumor Analysis Consortium (CPTAC), and Kaplan-Meier Plotter. Clinical tissue specimens were collected to detect the expression of PBK and CCNB1 by immunohistochemistry. Quantitative real-time polymerase chain reaction (PCR) was performed to detect PBK messenger RNA (mRNA) expression levels in clinical specimens and cell lines. Western blot was used to detect PBK protein expression in ovarian cancer cell lines. ES2 and A2780 cells with higher PBK expression were selected to construct PBK knockdown cell lines using lentiviral interference vectors. Cell Counting Kit-8 (CCK-8) assay, colony formation assay, and 5-ethynyl-2'-deoxyuridine (EdU) assay were performed to explore the effect of PBK knockdown on cell proliferation. Transwell assay was used to investigate the effect on cell invasion. The Cancer Genome Atlas (TCGA) and Kyoto Encyclopedia of Genes and Genomes (KEGG) databases were utilized to analyze PBK-related pathways and predict CCNB1 as the gene most closely related to PBK. RESULTS: PBK was significantly overexpressed in ovarian cancer tissues and cell lines compared with normal controls, and high PBK expression was associated with poor overall survival (OS) and progression-free survival (PFS). Knockdown of PBK expression inhibited the proliferation, colony formation, and invasion of ovarian cancer cells. Bioinformatics analysis revealed that CCNB1 was significantly overexpressed in ovarian cancer and high CCNB1 expression was associated with poor OS. CCNB1 was also significantly highly expressed in ovarian cancer tissues as validated by immunohistochemistry and was associated with lymph node metastasis. PBK and CCNB1 expression showed a significant positive correlation in TCGA ovarian cancer datasets. Knockdown of PBK inhibited CCNB1 expression in ovarian cancer cells. CONCLUSIONS: PBK promotes ovarian cancer cell proliferation and invasion. PBK knockdown leads to CCNB1 downregulation. These findings suggest that CCNB1 contributes to PBK-mediated oncogenic effects and identify the PBK-CCNB1 axis as a potential therapeutic target for ovarian cancer treatment.

PDZ-binding kinase (PBK)↗

PubMed2026

The prognostic value and molecular mechanisms of Porphyromonas gingivalis infection-associated differentially expressed genes in oral squamous cell carcinoma.

BACKGROUND: Increasing evidence suggests that Porphyromonas gingivalis (Pg) is associated with oral squamous cell carcinoma (OSCC) development and progression. This study aimed to identify Pg-associated genes with prognostic relevance in OSCC through integrated bioinformatics analysis. METHODS: OSCC-related differentially expressed genes (DEGs) were identified from the The Cancer Genome Atlas (TCGA)-OSCC cohort and intersected with Pg supernatant-associated DEGs from GSE192887. Raw count data were analyzed with DESeq2, whereas transcripts per million (TPM)-transformed expression values were used for downstream visualization and model construction. Weighted gene co-expression network analysis (WGCNA), univariate Cox regression, least absolute shrinkage and selection operator (LASSO) regression, and multivariable Cox modeling were used to develop a seven-gene prognostic signature, which was externally evaluated in GSE41613. Additional analyses examined treatment-associated expression changes in the seven model genes, pairwise correlations among the model genes, and correlations between Pg supernatant-associated differentially expressed gene (PgSDEG)-derived module eigengenes and immune-cell fractions. Quantitative reverse-transcription polymerase chain reaction (qRT-PCR) was performed in eight paired OSCC and adjacent non-tumor tissues and in supplemented-brain heart infusion (BHI) vehicle-control and Pg culture-supernatant-treated HOK, HSC-3, and CAL-27 cells. RESULTS: A prognostic signature comprising CXCL8, GAST, HBQ1, PADI3, STC1, TEX19, and TMEM92 was established. The signature showed limited-to-moderate discrimination in the TCGA training cohort, with 1-, 3-, and 5-year areas under the curve (AUCs) of 0.68, 0.69, and 0.69, respectively, and limited discrimination in the GSE41613 external cohort (AUCs: 0.66, 0.67, and 0.61). Kaplan-Meier analysis showed poorer survival in the high-risk group in both cohorts. The GSE192887 analysis showed significant treatment-associated expression changes in all seven genes after Pg culture-supernatant exposure. In paired tissues, CXCL8 and TMEM92 were significantly higher in OSCC tissues, whereas STC1 was not significant after Holm correction. In CAL-27 cells, CXCL8, STC1, and TMEM92 increased significantly after culture-supernatant treatment, whereas the corresponding comparisons were not significant in HOK or HSC-3 cells after adjustment. CONCLUSIONS: This study developed a seven-gene Pg-associated prognostic signature for OSCC and provided complementary transcriptomic, immune-correlation, tissue, and cell-based evidence that placed the signature in biological context. The model showed limited-to-moderate discrimination and is not ready for clinical use. The enrichment, gene-correlation, and immune-correlation findings are hypothesis-generating rather than mechanistic evidence. Further independent validation and dedicated functional studies are required.

Oral squamous cell carcinoma (OSCC)↗

PubMed2026

Genomic Characterization of ETV6::RUNX1-Positive Childhood B-ALL in a Chinese Cohort: Novel Fusion Partners, Co-Occurring Mutations, and Risk-Stratifying Biomarkers.

BACKGROUND: ETV6::RUNX1 is the most common genetic abnormality in pediatric B-cell acute lymphoblastic leukemia (ALL; ∼25%), yet the comprehensive genetic architecture and molecular predictors of intermediate-risk (IR) stratification remain incompletely characterized. METHODS: We performed whole-transcriptome sequencing (Illumina NovaSeq 6000, rRNA depletion, 41.70 Gb/sample) on bone marrow samples from 93 pediatric ETV6::RUNX1-positive B-ALL patients. Bioinformatics analysis included STAR alignment, MuTect2 variant calling, FusionCatcher fusion detection, and VEP annotation. The Jaccard index with permutation testing assessed mutation co-occurrence; logistic regression identified independent predictors of IR classification. RESULTS: Beyond ETV6::RUNX1, we identified 51 distinct fusion genes across the cohort, including the reciprocal RUNX1-ETV6 (73.1%), chr8::KLF1210 (38.7%), and KLF12-chr8 (34.4%). Somatic mutations in 249 genes were detected; the most frequent were KIAA1715 (17.2%), KRAS (11.8%), and NSD2 (10.8%). Network analysis revealed significant chromatin modifier co-occurrence (KIAA1715-KMT2C: J = 0.136, p = 0.015) and KRAS-NRAS mutual exclusivity (J = 0.000, p = 0.042). PTCH1 (OR = 3.50, 95% CI 0.21-58.49, p = 0.41) and GNB1 (OR = 6.5, 95% CI 1.2-34.8, p = 0.029) mutations independently predicted IR classification. chr8::KLF1210 fusion correlated with higher Day-19 MRD levels (p = 0.038). CONCLUSIONS: GNB1 mutation represents a novel independent predictor of IR stratification in ETV6::RUNX1-positive B-ALL. The chromatin modifier co-occurrence module and extensive fusion architecture reveal biological heterogeneity within this favorable-risk subtype, with potential implications for risk-adapted therapeutic strategies.

B‐ALL↗

PubMed2023

De novo missense variants in ZBTB47 are associated with developmental delays, hypotonia, seizures, gait abnormalities, and variable movement abnormalities.

The collection of known genetic etiologies of neurodevelopmental disorders continues to increase, including several syndromes associated with defects in zinc finger protein transcription factors (ZNFs) that vary in clinical severity from mild learning disabilities and developmental delay to refractory seizures and severe autism spectrum disorder. Here we describe a new neurodevelopmental disorder associated with variants in ZBTB47 (also known as ZNF651), which encodes zinc finger and BTB domain-containing protein 47. Exome sequencing (ES) was performed for five unrelated patients with neurodevelopmental disorders. All five patients are heterozygous for a de novo missense variant in ZBTB47, with p.(Glu680Gly) (c.2039A>G) detected in one patient and p.(Glu477Lys) (c.1429G>A) identified in the other four patients. Both variants impact conserved amino acid residues. Bioinformatic analysis of each variant is consistent with pathogenicity. We present five unrelated patients with de novo missense variants in ZBTB47 and a phenotype characterized by developmental delay with intellectual disability, seizures, hypotonia, gait abnormalities, and variable movement abnormalities. We propose that these variants in ZBTB47 are the basis of a new neurodevelopmental disorder.

Child↗

PubMed2026

Unveiling Aziridine-Containing Natural Products by Genomic and Spectroscopic Approaches.

Aziridine-containing natural products are prized for their potent bioactivities, yet their scarcity and poorly understood biosynthesis have limited systematic exploration. Here, we address this by integrating genome mining with a 1H-13C coupled HSQC metabolomic approach that exploits the distinctive NMR signatures of aziridines, enabling their direct detection from complex extracts. This strategy unveiled the desertolides, the first macrolides incorporating a rare terminal 2-methyl-aziridine-2-carboxylate moiety. Genetic and isotopic studies identified a dedicated biosynthetic subcluster (desA-desN) that assembles and installs this unit from glutamate, and heterologous expression confirmed the self-sufficiency of this subcluster. Direct MS evidence reveals the aziridine moiety covalently bound to the active-site Cys113 of DesN, establishing this KAS III homolog as the first dedicated aziridine-transferase and a promising tool for polyketide engineering. Bioinformatic analysis uncovered over 50 biosynthetic gene clusters, suggesting that this aziridine-associated biosynthetic logic may be more widespread than currently appreciated. This work establishes a tractable platform for the targeted discovery and engineered biosynthesis of aziridine-containing natural products, opening this underexplored pharmacophore to systematic interrogation.

Aziridines↗

PubMed2025

ATAC-seq in Emerging Model Organisms: Challenges and Strategies.

The Assay for Transposase-Accessible Chromatin with sequencing (ATAC-seq) is a versatile and widely utilized method for identifying potential regulatory regions, such as promoters and enhancers, within a genome. ATAC-seq has been successfully applied to a wide range of established and emerging model organisms. However, implementing this method in emerging model systems, such as arthropods, can be challenging due to several factors that influence data quality. These factors include the availability of a sufficient amount and quality of tissue or cells, the need for species- and tissue-specific protocol optimization, the completeness and accuracy of the reference genome, and the quality of the genome annotation. In this article, we emphasize the key steps in the ATAC-seq protocol that, based on our experience, have the greatest impact on data quality when adapting this method for emerging model organisms. Specifically, we discuss the importance of nuclei isolation, the incubation conditions of the Tn5 transposase, and PCR amplification of the library. Furthermore, we outline essential quality checkpoints during the bioinformatic analysis of ATAC-seq data to assist in assessing data integrity and consistency. Given that many emerging model organisms may not be readily available in laboratory cultures, we also emphasize the importance of evaluating how different preservation methods affect ATAC-seq data quality. Based on examples in one spider and one ant species, we demonstrate that replication and thorough quality controls at all steps of the protocol and data analysis are essential to assess the usability of ATAC-seq data. Our data highlights the importance of isolating the right number of intact nuclei, as well as ensuring optimal amplification conditions during library preparation to obtain good-quality sequence data for downstream analyses. We recommend using fresh tissue samples if possible because we show that direct cryopreservation of the tissue may affect chromatin integrity. This effect could be avoided or reduced by preserving the homogenate in cell culture medium. Overall, we explain the ATAC-seq protocol and downstream analyses in detail and give step-by-step advice to researchers who are new to the field and want to implement this method. With careful planning and validation, ATAC-seq can reveal the regulatory landscape of a genome and aid in identifying elements that govern gene expression.

Animals↗

PubMed2026

Novel TCOF1 Frameshift Variant and Phenotypic Heterogeneity in a Chinese Family With Treacher Collins Syndrome.

BACKGROUND: Treacher Collins syndrome (TCS) is a congenital craniofacial disorder characterized by malar and mandibular hypoplasia, downward-slanting palpebral fissures, and conductive hearing loss. Pathogenic variants in TCOF1 account for most cases, with POLR1D, POLR1C, and POLR1B also implicated. METHODS: Whole-exome sequencing was performed in a two-generation Chinese family with TCS, followed by Sanger sequencing validation. Clinical features were systematically evaluated, and bioinformatic analyses combined with structural modeling were employed to assess the potential pathogenicity of the identified variant. RESULTS: In this study, a novel heterozygous frameshift variant in TCOF1 (NM_001371623.1:c.1601_1602delCC, p.Pro534Leufs*15) was identified in the proband and his affected father. The proband presented classic TCS features including craniofacial skeletal hypoplasia, downward-slanting palpebral fissures, and conductive hearing loss. He also carried a right-sided preauricular fistula, a nonclassical feature of TCS. The same variant was detected in his affected father with a substantially milder phenotype, indicating marked intrafamilial phenotypic variability. Bioinformatic analysis and structural modeling predicted that this variant produces a severely truncated Treacle protein lacking key functional domains, which is predicted to disrupt nucleolar localization and ribosome biogenesis. CONCLUSION: Our findings expand the variant spectrum of TCOF1, highlight phenotypic heterogeneity in TCS, and reinforce the critical role of molecular diagnosis in distinguishing TCS from phenotypically overlapping craniofacial syndromes.

Humans↗

PubMed2025

A Practical Approach to High-Throughput and Accurate Mapping-by-Sequencing in Arabidopsis.

Forward-directed genetic screens are extremely powerful in identifying novel genes involved in a specific biological process, including various chromatin regulatory pathways. However, the traditional ways of genetic mapping are time- and cost-demanding. Recently, the whole process was revolutionized by the development of mapping-by-sequencing (MBS) protocols. In MBS, the causal mutations and their positions within genes are identified directly by whole-genome sequencing and bioinformatics analysis of the bulk of mutant plants selected based on the mutant phenotype from a segregating population. MBS increases precision and economizes the mapping. Here, we describe a general protocol and provide practical tips on how to proceed with the mapping-by-sequencing on the example of Arabidopsis forward-directed genetic screen designed to identify mutants sensitive to a specific type of DNA damage. The described protocol is generally applicable to a wide range of genetic screens in various inbreeding species with a reference genome sequence.

Arabidopsis↗

PubMed2026

The pseudokinase domain PK1 of UNC-89/obscurin is required for mitochondrial morphology and function in C. elegans.

UNC-89 is a giant modular protein located at the sarcomeric M-line of C. elegans striated muscle and is required for sarcomere organization and function. UNC-89 contains two protein kinase domains, PK1 and PK2, separated by 850 residues, that includes a 645-residue long intrinsically disordered sequence that acts like an elastic spring. Bioinformatic analysis suggests that PK2 is an active kinase whereas PK1 is a pseudokinase. We recently reported that a genome-edited worm, unc-89(sf22), that expresses UNC-89 carrying a kinase-inactivating point mutation in PK2 has an unusual phenotype with normally organized sarcomeres and SR, normal muscle function and yet fragmented mitochondria, increased ATP levels, increased glycolysis and alterations in electron transport chain complexes and respiration. Here, we show that a genome-edited worm unc-89(sf23), that expresses UNC-89 with an in-frame deletion of the C-lobe of PK1 has approximately the same phenotype as the PK2 catalytically dead mutant. The fact that mutations in two different regions of UNC-89 result in a mitochondrial phenotype is further evidence of communication between the sarcomere and mitochondria. We further demonstrate that in vitro PK2 interacts with full length PK1 and the C-lobe of PK1. The protein kinase domains of giant sarcomeric proteins are autoinhibited by parts of their own sequence, and this is also likely for PK2, but the mechanism by which PK2 would be activated is unknown. Our data is compatible with a model in which PK1 interacts with PK2 and thereby stimulates PK2 kinase activity.

Animals↗

PubMed2026

Expression of PIEZO1 in lung adenocarcinoma correlates with PD-L1 expression, cell migration, and poor prognosis: an exploratory study.

BACKGROUND AND AIMS: Lung adenocarcinoma (LUAD) treatment is challenging process. and the function of PIEZO1, a mechanically sensitive ion channel has not been systematically determined. In this study we aimed to explore the expression, potential associations, and clinical significance of PIEZO1 in LUAD. METHODS AND AIMS: A comprehensive bioinformatics analysis was performed using data from the Cancer Genome Atlas (TCGA) database, the Gene Expression Omnibus (GEO) database and other databases. Experimental validation was performed to confirm the expression patterns and preliminarily examine the associations of PIEZO1 in LUAD cell lines. RESULTS: PIEZO1 expression was significantly lower in LUAD tissues than in normal lung tissues (P < 0.05). Its expression was correlated with advanced pathological stage, lymph node involvement, and distant metastasis. High PIEZO1 expression was associated with a distinct immune-related tumor microenvironment, characterized by correlations with the expression levels of multiple immune checkpoint molecules, and was identified as an independent factor associated with poor overall survival (HR = 1.49; 95% CI 1.11-2; P < 0.007). In vitro experiments confirmed the downregulated PIEZO1 expression in LUAD cell lines, and functional knockdown experiments revealed its association with cell migration and PD-L1 expression. CONCLUSION: This exploratory study revealed that PIEZO1 expression in LUAD cell lines correlated with the expression of immune-related features and EMT-related genes, as well as poor prognosis. In vitro, PIEZO1 knockdown is associated with reduced cell migration and decreased PD-L1 expression. These findings provide a basis for future investigations into the potential role of PIEZO1 in LUAD.

Gene expression↗

PubMed2025

A general framework to over-express tRNA-derived fragments from their parental tRNAs in mammalian cells.

tRNA-derived fragments (tRFs), generated from the cleavage of mature or precursor tRNAs are a category of regulatory noncoding RNAs with diverse functions in physiological or pathophysiological conditions. Here we describe a framework for the over-expression of tRFs from their parental tRNAs in mammalian cells. The process involves bioinformatics analysis to identify specific tRNAs that produce the tRF, PCR amplification of corresponding tRNA genes, and insertion into expression vectors. Transfection is carried out in HEK293T cells and detection of tRFs is achieved through northern blotting and dual luciferase reporter assays. In the latter, a complementary sequence to the tRF of interest is inserted into the luciferase reporter. By observing the reduction in luciferase activity, we can validate the expression of tRFs. This method enables precise study of tRF functions and their roles in cellular processes.

Humans↗

PubMed2026

MX1 promotes gastric cancer cell migration via inhibiting ANXA2 ubiquitination and degradation.

Gastric cancer (GC) is a globally lethal malignancy, with invasion and metastasis driving treatment failure and poor prognosis. MX dynamin like GTPase 1 (MX1) shows tumor-specific functional heterogeneity, while its expression, biological functions and molecular mechanisms in GC remain unclear. Here, we explored MX1's clinical significance and its regulatory mechanism in GC cell migration. We integrated public databases and institutional paired clinical samples for bioinformatics analysis of MX1's correlation with clinical outcomes, and verified its pro-migratory effect via Transwell and wound healing assays. Co-immunoprecipitation/mass spectrometry (Co-IP/MS), immunofluorescence and ubiquitination assays were used to identify MX1-interacting proteins and dissect the underlying mechanism, and the Genomics of Drug Sensitivity in Cancer database was applied for chemosensitivity analysis. MX1 was aberrantly upregulated in GC tissues and served as an independent prognostic biomarker, with high expression associated with shortened overall, first-progression and post-progression survival. MX1 promoted GC cell migration and epithelial-mesenchymal transition pathway enrichment, and directly bound Annexin A2 (ANXA2) in the cytoplasm; both were co-enriched in endothelial and epithelial cells by single-cell sequencing. MX1 dose-dependently upregulated ANXA2 protein (without affecting its mRNA) by inhibiting NEDD4L/TRIM65-mediated ANXA2 ubiquitination and degradation, enhancing ANXA2 stability. Additionally, high MX1 expression correlated with increased paclitaxel sensitivity in GC patients based on database analysis, and CCK-8 assays confirmed that MX1 overexpression significantly reduced the paclitaxel IC50 in gastric cancer cells, supporting its potential as a predictive biomarker for paclitaxel efficacy. This study demonstrates that MX1 promotes GC cell migration by suppressing ANXA2 ubiquitination and degradation, highlighting the critical role of the MX1-ANXA2 axis in GC progression. These findings provide novel molecular targets and theoretical support for GC prognostic evaluation, individualized chemotherapy and targeted therapy.

ANXA2↗