Functional & Integrative Genomics最新文献

Oenothera species are increasingly valued for their medicinal and ornamental qualities and serve as important models in classical cytoplasmic genetics research. The genus Oenothera L., one of the largest in the Onagraceae family, includes 18 subsections and two deep phylogenetic lineages, Clade A and Clade B. Analyzing high-quality chloroplast genomes can provide crucial insights into species classification and genus-level evolution. In this study, we report the complete chloroplast genome of Oenothera drummondii Hook., the first species from subsection Raimannia, with a total length of 167,177 bp and a GC content of 39.3%. This genome contains 129 genes and displays a typical quadripartite structure. Combining this genome with data from 16 publicly available chloroplast genomes, we conducted a comprehensive comparative and evolutionary analysis. Our results indicate that Clade B species diverged independently from Clade A species. Within Clade A, species from subsection Muniza form a distinct branch, while O. drummondii clusters closely with species from subsection Oenothera. Phylogenetic analysis correlates well with chloroplast genome structural differences, such as the loss of the infA gene in Clade B species, the expansion of the IR regions in Muniza, and a shared large inversion in the LSC region among Raimannia and Oenothera species. We also identified repeat sequences, six highly variable genes, and positively selected genes among the 17 chloroplast genomes analyzed. These findings offer valuable insights into the evolutionary processes of Oenothera species and provide a foundation for the development of future molecular markers based on the identified genes and structural variations.

Graphical Abstract

The breakthrough progress in gene editing technology has established a brand-new paradigm for gene function analysis and crop genetic improvement, and has already become the core driving force in modern agricultural biotechnology. However, traditional methods for screening edited individual plants without T-DNA insertion in their offspring are time-consuming and labor-intensive. To accelerate the identification of T-DNA-free edited plants, researchers have continuously explored various strategies to improve screening efficiency, achieving some success. Despite this progress, these strategies also exhibit varying degrees of limitations. Recently, research teams led by Cheng and Sun (2025) and Zhu et al. (2025) constructed gamete elimination systems, successfully achieving a 100% non-transgenic status in the mutant plants. This innovative approach has transformed traditional molecular testing from a "laboratory-dependent" model to "field-based real-time decision-making" providing an efficient and cost-effective solution for precision breeding.

Eukaryotic translation initiation factor 3 (eIF3), the largest eukaryotic initiation factor, regulates mRNA translation initiation. While eIF3 has been extensively studied in various plant species, research on eIF3 in upland cotton remains limited. A total of 60 GheIF3s were identified using a bioinformatics approach, and comprehensive characteristic analysis was conducted in upland cotton. The results revealed that the different subunits of the GheIF3s were strongly conserved in both structural and evolutionary relationships. RNA-seq and qRT‒PCR results revealed that the expression levels of GheIF3L.1 and GheIF3M.2 in early‒maturing varieties were significantly greater than those in late-maturing varieties. Furthermore, no single-nucleotide polymorphisms (SNPs) were detected in GheIF3L.1, and a SNP (A05:105181831, T/G) significantly related to the flowering period within the coding sequence of GheIF3M.2 was identified. The flowering time of the varieties carrying the GG allele was significantly earlier than that of the varieties carrying the TT allele, which was verified in both early − flowering and late-flowering varieties via Sanger sequencing. Genetic diversity analysis revealed that the gene region differed between early − and late − maturing varieties. Virus-induced gene silencing (VIGS) results indicated that the silenced plants (TRV:GheIF3M.2) presented a significant decrease in plant height; inhibited flower bud differentiation; and significantly decreased IAA, GA₃, and BR contents. These analyses elucidated the functions of GheIF3s in upland cotton and provided genetic resources related to plant morphology and early maturity.

Clustered regularly interspaced short palindromic repeats (CRISPR)-associated nuclease protein 9 (Cas9) is a powerful tool used for genome engineering, but concerns remain about off-target effects. Here we evaluate potential deleterious effects of CRISPR-Cas9 by combining sequence analysis and the genetics of the male X chromosome in a Drosophila model. Since males have only one X chromosome deleterious mutations on the X chromosome will manifest in reducing viability or result in visible phenotypes and thus provide sensitive readouts of off-target activity. Our data do not support large scale off-target effects in Drosophila. To optimize sgRNA selection, we incorporated off-target evaluation into the PlatinumCRISPr sgRNA selection tool for a broad range of organisms.

GATA3 is expressed in the outer cells of the morula stage during embryonic development and is considered a key driver of the regulation of early lineage development in bovines. This research presents an optimised somatic cell validation resource, successfully generating GATA3 knockout (KO) Bovine Fetal Fibroblasts and MDBK cells using CRISPR/Cas9-mediated genome editing for their future implications in vivo studies designed to definitively understand the role of GATA3 in cell lineage specification and bovine embryo development. This involved designing single-guide RNAs (sgRNAs) targeting different regions of the GATA3 gene, cloning them into the px459 plasmid, delivering the CRISPR clone into bovine fibroblast cells and the MDBK cell line, screening for successful targeting and knockouts, and MiSeq analysis to verify successful disruption of the GATA3 gene. A total of eleven guides were designed targeting the functional domains in Exons 4 and 5 and the transcription initiation site in Exon 2. Designed guides were first optimized and screened using an in vitro cleavage assay. The guides with the best cutting efficiencies were then tested in vivo by targeting bovine fetal fibroblast (BFFs) and MDBK cell line followed by MiSeq analysis to verify the successful knockouts. A total of two effective guides were identified targeting the zinc-finger (ZnF) functional domains of the GATA3 gene (sgRNA#5 and sgRNA#8 in Exon 4 and Exon 5, respectively) and one in Exon 2 (sgRNA#1) targeting the transcription initiation site of the GATA3 gene. MiSeq data from targeted bovine cells showed indel frequency of 47.40%, 55.5%, and 42.4% in bovine fetal fibroblasts, 11.03%, 28.9% and 7.3% for MDBK cells for top three sgRNAs. Overall, MiSeq data for 3 selected sgRNAs showed successful disruption of the GATA3 gene, inserting a base pair 2–3 bp upstream of the PAM site, ultimately resulting in a premature stop codon TAA in the downstream region. This study established and validated highly efficient sgRNAs targeting the GATA3 gene, forming a molecular basis for forthcoming functional investigations in bovine embryos to explore gene function and protein-level effects.

This study aimed to investigate the effects of paeonol (pae) on blood pressure, tissue damage, and the fetuses and placentas in a preeclampsia (PE) rat model induced by L-NAME, and to explore the potential regulatory mechanism involving the NEDD4L-ACSL4 axis. By quantitatively analyzing urinary protein, blood pressure, and sFlt-1 levels, the role of pae in the PE rat model was evaluated. The effects of pae on HTR-8/SVneo and primary trophoblast cells were investigated using a cell model. The role of pae in the process of NEDD4L-mediated ACSL4 ubiquitination was determined through experiments such as Co-IP and ubiquitination assays. Pae reduced blood pressure, urinary protein, and sFlt-1 levels in a PE rat model, increased the fetal survival rate, and improved placental and renal damage. Mechanistically, pae mediated the ubiquitination level of ACSL4 by promoting the interaction between ACSL4 and NEDD4L, thereby reversing hypoxia-induced ferroptosis. In brief, pae further inhibited ferroptosis by facilitating NEDD4L-mediated ACSL4 ubiquitination, exerting a significant improving effect on blood pressure and tissue damage in PE. Thus, it provided a new strategy for the treatment of PE and laid a foundation for further research on the mechanism of action of pae and its related compounds.

Recent studies have highlighted the efficacy of platelet-rich plasma-derived exosomes (PRP-Exo) in managing osteoarthritis (OA). This research aimed at exploring the effect and mechanism of PRP-Exo-mediated intervention in chondrocyte pyroptosis and knee osteoarthritis. In this study, PRP was collected from human volunteers and rats, and the PRP-Exo was extracted, which was employed in the in vitro and in vivo OA model. The molecular interaction was analyzed by luciferase reporter assay, Chromatin immunoprecipitation and Electrophoretic Mobility Shift Assay experiment. Cell pyroptosis was analyzed by flow cytometry and biomarkers detection. Degrees of cartilage injury were detected using Alcian Blue, Safranin O Fast Green, and Toluidine Blue O staining. Our results showed that PRP-Exo had a similar effect with PRP on alleviating chondrocyte pyroptosis and OA progression. PRP-Exo functioned by transferring miR-25-3p to chondrocyte, which targeting to MAP4K2 and regulating Hippo pathway. MiR-25-3p inhibition weaken the effect of PRP-Exo, while MAP4K2 overexpression reversed the effect of miR-25-3p overexpression. Besides, TNFAIP3 was confirmed as a downstream molecular of MAP4K2/Hippo pathway. The effect of miR-25-3p was also confirmed in a rat OA model. Collectively, this study elucidated that PRP-Exo derived miR-25-3p alleviated chondrocyte pyroptosis and OA progression by regulating the MAP4K2/Hippo/TNFAIP3 pathway.

Trophinin-associated protein (TROAP) is a proline-rich cytoplasmic protein exclusively on the apical side of syncytiotrophoblasts and is associated with the microtubular cytoskeleton. We analyzed the expression, promoter methylation, and relevant pathways of TROAP in colorectal cancers (CRCs) using bioinformatics, validated TROAP expression with RT-PCR, western blot and immunohistochemistry, and examined its clinical implications. Its biological processes and molecular mechanisms were investigated by tumor xenograft models, TUNEL, CCK-8, flow cytometry, wound healing and transwell assays, Nile red staining, western blot, proteomic and bioinformatics analysis. TROAP expression was significantly elevated in CRC compared to that in normal mucosa (p < 0.05). TROAP mRNA expression was positively correlated with p53 mutation, poor clinical outcome and favorable prognosis in CRC (p < 0.05). TROAP methylation was inversely correlated with its mRNA expression, lower clinicopathological staging, and non-mutant p53 expression (p < 0.05). TROAP expression was positively associated with younger age, distal metastasis, TNM stage, differentiation, and poor prognosis in CRC patients (p < 0.05). TROAP expression was closely linked to cell cycle, nuclear division, chromatid segregation, calcium and Wnt signaling pathway, ECM regulators and glycoproteins, cell senescence, CPCR-ligand, iron ion and heparin binding in CRCs (p < 0.05). TROAP promoted cell proliferation, resistance to apoptosis and pyroptosis, as well as cell migration, invasion, and epithelial-mesenchymal transition in CRC cells. TROAP aggravated lipid droplet formation and subsequent chemoresistance via de novo lipogenesis via histone acetylation and PI3K/Akt pathway. Aberrant TROAP expression could serve as a biomarker for aggressive behavior and poor prognosis in CRCs, as well as a molecular target for gene therapy.

Excessive adipose tissue accumulation adversely impacts the health of both humans and livestock. Adenylyl cyclase 3 (ADCY3) is a promising anti-obesity target, yet its regulatory role in adipogenesis remains incompletely understood. Our findings revealed a dynamic pattern of ADCY3 expression during adipogenesis and lipid droplet (LDs) accumulation. Functional analyses demonstrated that ADCY3 overexpression impaired adipogenesis by downregulating adipogenic transcription factors CEBPα and PPARγ. Furthermore, it reduced both the number and size of LDs through suppressing triglyceride synthesis and fatty acid metabolism, concomitantly downregulating key genes involved in LDs formation (PLIN1, CIDEC, FIT2, and Seipin), as well as factors mediating glycerol ester synthesis and fatty acid metabolism (DGAT1, DGAT2, ACC, SCD, FASN, and ACSL1). Transcriptomic profiling revealed that ADCY3 overexpression suppressed PPARγ signaling, leading to the downregulation of oxidative phosphorylation genes encoded by both the nuclear and mitochondrial genomes. Our results implicate ADCY3 in the regulation of lipid metabolism, with the speculative involvement of mitochondrial metabolic remodeling. This perspective offers a framework for developing future interventions against excessive lipid deposition.

The rapid expansion of transcriptomic data has necessitated the development of efficient and scalable analytical frameworks for Differential Gene Expression (DGE) Analysis. We present a web-based tool implementing the DGEAR (Differential Gene Expression Analysis with R), designed using a three-tier architecture to enhance usability, performance, and modularity. The system comprises a frontend user interface (UI) for seamless data submission and visualization, a middleware processing layer handling computational tasks with an Application Programming Interface, and a backend data layer for secure data management and efficient execution of statistical analyses. The DGEAR algorithm integrates multiple statistical methods and an ensemble model with a cut-off-based majority voting strategy, ensuring robustness, flexibility, and accuracy in identifying differentially expressed genes from microarray and RNA-seq datasets. Furthermore, DGEAR integrates gene set enrichment analysis and PPI network construction, giving a major head start in downstream analysis. The web tool supports custom parameter selection, data visualization, and end-to-end encryption, making it securely accessible to researchers and clinicians. Generated output data can be directly accessible and can be downloaded by the user. This platform significantly streamlines transcriptomic analysis, providing an intuitive, high-performance environment for bioinformatics investigations, and is publicly available at https://dgear.compbiosysnbu.in/.