Molecular Genetics and Genomics最新文献

Aging is a major biological process underlying increased risk of chronic and neurodegenerative diseases, yet its molecular mechanisms remain incompletely defined. Our study systematically investigates the conserved functions and pathways of W06A7.4 in Caenorhabditis elegans and its human homolog TMEM144 in the regulation of aging, combining genetic manipulation in model organisms, analysis of human clinical samples, and functional assays in cell lines. The results demonstrate that W06A7.4 promotes longevity in C. elegans through synergistic effects with dietary restriction, reduction of oxidative damage, modulation of IIS and mTOR signaling, and maintenance of mitochondrial membrane potential. In human samples and cellular models, TMEM144 expression increases with age and in Alzheimer's disease. Our results suggest that TMEM144 may be involved in the regulation of glucose transport and mitochondrial respiration via the downstream protein TIMMDC1. These findings advance our understanding of evolutionarily conserved aging pathways and identify W06A7.4/TMEM144 as promising molecular targets for anti-aging and neurodegenerative disease interventions.

Antimicrobial resistance (AMR) presents a critical global challenge, causing over 1.27 million deaths annually, with projections reaching 10 million by 2050. Among the most concerning contributors are Enterobacteriaceae, particularly Escherichia coli and Klebsiella pneumoniae, which harbour Extended-Spectrum β-Lactamase (ESBL) genes-enzymes that hydrolyse β-lactam antibiotics and confer resistance-such as bla-CTX-M, bla-SHV, and bla-TEM. These genes confer resistance to β-lactam antibiotics, including penicillins and cephalosporins, limiting treatment options for urinary tract infections, bloodstream infections, and pneumonia. The World Health Organisation has classified these pathogens as critical targets for new drug development. In this study, we comprehensively analysed all known variants of bla-CTX-M, bla-SHV, and bla-TEM genes along with their wild-type sequences. Using a multi-step computational approach, we assessed guanine-cytosine (GC) content, single nucleotide polymorphisms (SNPs; single-base changes in DNA), insertion and deletion (InDel) variants (mutations involving nucleotide addition or removal), codon usage patterns, transcription factor binding sites (TFBS; DNA regions regulating gene expression), amino acid composition, protein stability, mutational hotspots, nucleotide and amino acid mutation frequencies, hydrophobicity, isoelectric point, aromaticity, aliphatic index, and molecular flexibility. The integrated dataset maps conserved regions and identifies residues frequently associated with resistance phenotypes. Our findings provide a framework for predicting resistance-associated mutation patterns and identifying genomic regions suitable for resistance-free drug targeting. These insights support prioritising drug target sites, optimising screening libraries, and generating high-quality datasets for machine learning-based precision drug design.

Diamond-Blackfan anaemia (DBA) is a rare inherited disorder marked by early-onset macrocytic anaemia and erythroid hypoplasia, resulting from mutations in ribosomal protein genes. Despite growing genetic insights, data on functional validation remain limited in India; here we report a novel RPS10 mutation with functional validation and provide genotype-phenotype correlation by integrating our findings with all previously reported RPS10 variants. A clinically suspected Diamond-Blackfan anaemia (DBA) case was evaluated through haematological profiling, bone marrow examination, and erythrocyte adenosine deaminase (eADA) activity measurement. Whole exome sequencing (WES) was followed by Sanger sequencing to identify and validate a novel pathogenic variant. Gene expression of ribosomal and regulatory genes was analysed by quantitative RT-PCR, and rRNA processing analysis was carried out to assess functional impact. The proband presented with severe macrocytic anaemia, reticulocytopenia, and erythroid hypoplasia consistent with Diamond-Blackfan anaemia (DBA). Whole exome sequencing identified a novel heterozygous nonsense variant in RPS10 (c.206G > A; p.Trp69Ter), and Sanger sequencing confirmed the variant as de novo. Gene expression analysis revealed significant upregulation of TP53 and downregulation of RPS10 and GATA1, indicating ribosomal dysfunction and activation of the p53 pathway. Additionally, the rRNA processing defect validated the pathogenicity of the novel RPS10 variant. This study identifies a novel de novo nonsense variant in RPS10 associated with Diamond-Blackfan anaemia, with supporting functional evidence of haploinsufficiency and p53 pathway activation. These findings expand the mutational spectrum of RPS10 and underscore the diagnostic value of integrating genomic and functional analyses in rare haematological disorders, while also contributing to ongoing efforts to delineate genotype-phenotype correlations in DBA.

Moraxella catarrhalis is a Gram-negative diplococcus bacterium and a common respiratory pathogen, implicated in 15-20% of otitis media (OM) cases in children and chronic obstructive pulmonary disease (COPD) in adults. The rise of drug-resistant Moraxella catarrhalis has highlighted the urgent need for the potent vaccine strategies to reduce its clinical burden. Despite a mortality rate of 13%, there is no FDA-approved vaccine for this pathogen. The aim of this study was to computationally identify novel antigens and design a multi-epitope peptide-based vaccine candidate against M. catarrhalis using an immunoinformatics-driven subtractive proteomics and reverse vaccinology approaches. The core proteome of 12 M. catarrhalis genomes were analyzed, identifying 360 host non-homologous proteins. Subsequent screening revealed 30 metabolic pathway-dependent and 7 independent drug targets, along with 7 membrane and extracellular proteins as potential vaccine candidates. A prioritized protein target (WP_081569984.1) was selected for vaccine design. The predicted B-cell, MHC-I, and MHC-II epitopes were linked using adjuvants and linkers to construct four vaccine candidates (V1-V4). These constructs were assessed for physicochemical properties, allergenicity, antigenicity, secondary structures, and immune receptor interactions. As a result, V1 emerged as the most promising candidate. Molecular docking and molecular dynamics (MD) simulations evaluated the interactions of V1 with human toll-like receptors (TLR2 and TLR3). MD trajectories including RMSD, RMSF, Radius of gyration (Rg), SASA, binding free energy (MM-PBSA), PCA, free energy landscapes, and DCCM, showed a strong interaction of vaccine with the TLR recptors. Immune simulations predicted significant immune responses against the proposed vaccine. Additionally, the vaccine construct was in-silico tested in an E. coli plasmid vector (pET-28a(+) for its cloning potential. These findings highlight the potential of the proposed multi-epitope vaccine V1 as a safe and effective preventive strategy against M. catarrhalis-associated infections, and additionally laid the groundwork for future in vitro, in vivo, and clinical studies to validate its immunogenicity and protective efficacy.

Accurate variant calling is essential for next-generation sequencing (NGS)-based diagnosis of rare diseases, yet most benchmarking studies have focused on standard cell lines or trio-based samples, with limited relevance to sporadic cases. Here, we systematically compared the performance of DeepVariant and GATK HaplotypeCaller in two Chinese cohorts of patients with sporadic epilepsy (EP) and autism spectrum disorder (ASD). DeepVariant exhibited higher precision and sensitivity in detecting single nucleotide variants (SNVs), while GATK showed a distinct advantage in identifying rare variants, which are often key to understanding the genetic basis of rare diseases. Comparative analyses based on disease-related gene panels further highlighted differences in the identification of potentially deleterious variants. These findings reveal important trade-offs between variant callers and emphasize the need to tailor variant-calling strategies to specific research and clinical contexts. Our study provides practical vision for optimizing germline variant detection pipelines in sporadic neurodevelopmental disorders, offering broader insights for precision medicine applications.

Mosquito reproductive biology is an underexplored area with potential for developing novel vector control strategies. In this study, we investigated the role of the testis-specific serine/threonine-protein kinase (tssk) family, an essential regulator of spermiogenesis in mammals, in mosquitoes. We identified tssk homologues, As_tssk3 and Aea_tssk1, in Anopheles stephensi and Aedes aegypti, respectively and analyzed their expression across different developmental stages. Functional characterization of As_tssk3 was performed by RNAi mediated gene silencing in An. stephensi. The efficiency of As_tssk3 knockdown was analyzed using qRT-PCR, and the testes of knockdown and control males were compared for phenotypic changes. Additionally, the effect of As_tssk3 knockdown on the reproductive potential of male mosquitoes was analyzed via mating analysis. Further, in-silico studies were conducted to identify interactions between As_TSSK3 and the homologues of known Drosophila TSSK targets involved in the process of spermatogenesis, in Anopheles. As_tssk3 and Aea_tssk1 were found to express in a male-specific manner throughout development. Efficient knockdown in the expression of As_tssk3 gene was observed upon feeding An. stephensi larvae with target dsRNA. The knockdown of As_tssk3 led to a reduced sperm reservoir in adult males. As_TSSK3 was found to interact with the Nucleosome Assembly Protein1 like-4 (NAP1like-4), a putative transcription factor in our in-silico analysis. We hypothesize that As_TSSK3 might regulate the transcription of the protamine gene via the NAP protein. The study presented here is the first report of the characterization of tssk homologues in mosquitoes' and demonstrates a possible role of As_tssk3 in maintaining sperm stores.

The aim of this study was to investigate three unrelated Simmental calves with atypical white coat color, identify potential genetic causes using a trio-based whole-genome sequencing approach, and assess the prevalence of the identified variants in the breed. Several inherited alleles affecting coat color, ranging from fawn to red spotted and white-headed, have been described in Simmental cattle originating from Switzerland. However, no genetic variant has yet been associated with an almost completely white coat in this breed. Clinical examination revealed different syndromic disorders of white coat color in Simmental in all three cases, and pedigree records indicated recessive inheritance. Filtering for rare protein-changing variants revealed an independent homozygous variant that could be the cause in each case: a likely pathogenic missense variant in TYR (NP_851344.1:p.Pro428Leu) in case 1 with oculocutaneous albinism type 1, a likely pathogenic missense variant in GRID1 (XP_024842694.1:p.Pro489His) in case 2 with short stature-auditory depigmentation syndrome, and a frameshift variant of uncertain significance in RAD54B (NP_001179884.1:p.Ala722_Gly724delinsAsnLeuIlePheCys*) in case 3 with a multisystem depigmentation syndrome. Validation by Sanger sequencing confirmed the variant genotypes, and parental heterozygosity supported recessive inheritance. These variants were almost entirely absent from other breeds, and the allele frequency of the three candidate causal variants was less than 1% in the current Swiss Simmental population. This study identified three novel recessive alleles associated with syndromic forms of albinism or depigmentation, revealing unexpected heterogeneity. The investigation did not reveal any indications of possible dominant de novo mutations impacting protein coding genes including known candidate genes for depigmentation phenotypes. These findings possibly expand the list of pigmentation related genes in mammals, but further investigation is needed. We also highlight the biomedical relevance of investigating rare congenital disorders in livestock.

Male infertility affects a significant number of couples worldwide, yet the precise causes and genetic mechanisms underlying this condition remain largely unknown. To investigate the monogenic causes of primary male infertility, we performed exome sequencing (ES) in a cohort of 100 unrelated Pakistani male patients with non-obstructive, non-syndromic primary infertility. ES identified potential causal variants in established infertility-associated genes in 17/100 patients, resulting in a diagnostic yield of 17%. This highlights the effectiveness of next-generation sequencing technologies, particularly the ES approach, in unraveling the genetic basis of male infertility. Overall, this study provides valuable insights into the genetic underpinnings of male infertility. Our findings explore the diagnostic potential of ES and expand the spectrum of causal variants underlying male infertility.

Variant classification in genetic testing often culminates in "uncertain" calls, known as variants of uncertain significance (VUS), which remain a major clinical challenge. Among the established criteria for variant classification, population allele frequency (AF) is fundamental, yet under-representation of non-European groups hinders accurate interpretation. In this study, we evaluated the impact of population-specific AF on the reclassification of VUS and conflicting variants in hereditary cancer genes. From ClinVar, we curated 487 variants present in both the Brazilian cohort ABraOM and gnomAD v4.1 databases. Comparative population analysis revealed that 43% of shared variants showed significantly different AFs (q ≤ 0.01), with 113 (23%) exhibiting large effect sizes (OR ≥ 4), including 39 VUS. Among these, 20 VUS had higher AF in the Brazilian cohort and exceeded benignity thresholds (BS1), while remaining rare in other populations. Functional prediction tools such as REVEL and CADD failed to distinguish these variants from globally rare VUS. Integrating Brazilian‑specific AF with ClinGen VCEP rules downgraded 15% (3/20) of candidate VUS and resolved five conflicting calls. These findings argue for routine incorporation of regional reference datasets in diagnostic curation to reduce uncertainty and avoid inappropriate clinical management in diverse populations.