Molecular Genetics and Genomics最新文献

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.

Salt mine workers are occupationally exposed to DNA damaging agents at their workplace. The present study estimates the extent of DNA damage and expression deregulation of related genes in mine workers, staff, community living nearby and unexposed controls. Blood samples were collected from all groups. Expression analysis of AGT, H2AX and Mre11 genes was done using RT-PCR. DNA damage was detected by comet assay. Relative expression of selected genes was upregulated in mine, staff and community group compared to control. Expression of all three genes increased significantly with increasing age, total exposure time and smoking. DNA damage was higher in mine workers compared to control, staff and community groups. Elevated serum levels of sodium, potassium, chloride and total ROS were observed in mine and staff group compared to control and community group. Positive correlation was observed between gene expression versus total exposure time. Moreover, significant dependable regression was observed between gene expression versus comet parameters. The present study anticipated a negative impact of mine environment on the genomic stability of mine workers and staff group. Moreover, age, exposure time and smoking act synergistically to enhance the extent of DNA damage, ROS production, electrolyte imbalance and expression deregulation of selected genes. In addition, current research will provoke thoughtful insights to rethink the risk assessments for genetic integrity of community living nearby mines.

Fennel (Foeniculum vulgare Mill.) is a widely cultivated medicinal and aromatic plant valued for its essential oils used in pharmaceutical, culinary, and industrial applications. Breeding activities for fennel have been historically limited, but recent genomic advances have revealed substantial genetic diversity and variability among its populations, offering new opportunities to improve yield, oil composition, and stress resilience. Studies using molecular markers including RAPD, ISSR, SSR, and SNPs have characterized the genetic structure of fennel germplasm and identified key loci for traits such as seed yield, essential oil profile, and disease tolerance. Quantitative trait locus (QTL) mapping and principal component analysis (PCA) have refined genotype selection. Transcriptomic studies related to t-anethole biosynthesis and expression profiles under stress conditions have enabled functional gene discovery. Biotechnological tools such as callus induction, doubled haploid protocols, and in vitro selection techniques have emerged as adjunct strategies to accelerate breeding outcomes. Integration of classical breeding methods with molecular and biotechnological approaches enables precision breeding of fennel cultivars tailored for modern agricultural needs. Enhancing genetic diversity utilization and targeting key traits will support the development of high-performing, resilient varieties. This direction advances both the sustainability of fennel cultivation and its utility in agro-industrial sectors.

An introgression from Moricandia arvensis is known to restore male fertility to Brassica juncea cytoplasmic male sterile lines carrying M. arvensis, Diplotaxis berthautii, D. catholica or D. erucoides cytoplasm. We have previously mapped the fertility-restorer gene (Rfm) to the distal end of A09 chromosome of B. juncea but the restorer gene remains to be discovered. This study was undertaken to identify and clone the restorer gene(s) using next-generation sequencing approach, leveraging its known chromosomal location and flanking markers. We assembled the draft genome of the B. juncea fertility restorer line (MRS15), carrying the M. arvensis introgression. Alignment of the MRS15 genomic scaffolds to B. juncea reference genome identified six scaffolds aligned to the terminal region of chromosome A09 (between 51 and 58.5 Mb) harbouring the Rfm. The high-density linkage map of Rfm locus confirmed the correct orientation of these scaffolds. Based on segregation of tightly linked flanking markers, namely, the earlier reported BjESSR06 and a newly identified SRB17 marker, the Rfm gene was assigned to Scaffold-547. In silico analysis revealed six pentatricopeptide repeat (PPR)-encoding Restorer-of-Fertility-Like (RFL) genes in the ~ 300 kb region delimited by the above stated markers. Based on the expression profiles of these genes in CMS and fertility restorer lines, and in a segregating population, PPR-640 was identified as the Rfm gene. Further, we designed a gene-based, co-dominant marker perfectly co-segregating with fertility restorer trait through collinearity analysis of the genomic region spanning PPR-640 and the B. juncea genome. The Rfm gene and the marker reported here are critical for utilising this CMS system in hybrid breeding and to clone and study evolution of restorer genes in other Brassicaceae members.

Oncogenic Ras mutations are prevalent in human cancers, yet the mechanisms by which Ras promotes tumorigenesis remain incompletely understood. In Drosophila, oncogenic Ras (Ras^V12) induces tissue overgrowth and metastasis, but the cellular restraints it must overcome are unclear. We have identified Drosophila CRIF, the homolog of mammalian CR6-interacting factor 1 (CRIF1), as a modifier of Ras^V12-induced lethality and Ras^V12-induced overgrowth and cell proliferation. Knockdown of CRIF exacerbated Ras^V12 phenotypes, while CRIF overexpression ameliorated them. Further, we found that CRIF was also required for heterochromatin formation, as loss of CRIF suppressed position-effect variegation (PEV) and reduced the levels of Heterochromatin Proteins 1 (HP1) and Histone H3 Lysine 9 trimethylation (H3K9me3). CRIF physically interacted with HP1, suggesting a role in recruiting HP1 to heterochromatin. Notably, CRIF did not regulate HP1 transcription or total protein levels but influenced HP1 localization. Our findings demonstrate that CRIF functions as a tumor suppressor by negatively regulating cell proliferation and maintaining heterochromatin stability. CRIF's interaction with HP1 and its role in heterochromatin regulation suggest a novel mechanism linking heterochromatin to tumor suppression in Ras-driven cancers. These results highlight CRIF as a potential therapeutic target and provide new insights into the interplay between chromatin regulation and oncogenic signaling.