Search Results (2,178)

Plants face an array of environmental stresses, including both abiotic and biotic stresses. These stresses significantly impact plant lifespan and reduce agricultural crop productivity. Abiotic stresses, such as ultraviolet (UV) radiation, high and low temperatures, salinity, drought, floods, heavy metal toxicity, etc., contribute to widespread crop losses globally. On the other hand, biotic stresses, such as those caused by insects, fungi, and weeds, further exacerbate these challenges. These stressors can hinder plant systems at various levels, including molecular, cellular, and development processes. To overcome these challenges, multi-omics computational approaches offer a significant tool for characterizing the plant’s biomolecular pool, which is crucial for maintaining homeostasis and signaling response to environmental changes. Integrating multiple layers of omics data, such as proteomics, metabolomics, ionomics, interactomics, and phenomics, simplifies the study of plant resistance mechanisms. This comprehensive approach enables the development of regulatory networks and pathway maps, identifying potential targets for improving resistance through genetic engineering or breeding strategies. This review highlights the valuable insights from integrating multi-omics approaches to unravel plant stress responses to both biotic and abiotic factors. By decoding gene regulation and transcriptional networks, these techniques reveal critical mechanisms underlying stress tolerance. Furthermore, the role of secondary metabolites in bio-based products in enhancing plant stress mitigation is discussed. Genome editing tools offer promising strategies for improving plant resilience, as evidenced by successful case studies combating various stressors. On the whole, this review extensively discusses an advanced multi-omics approach that aids in understanding the molecular basis of resistance and developing novel strategies to improve crops’ or organisms’ resilience to abiotic and biotic stresses. Full article

Since the advent of the clustered regularly interspaced short palindromic repeats (CRISPR) system in the gene editing field, diverse CRISPR-based gene editing tools have been developed for treating genetic diseases. Of these, base editors (BEs) are promising because they can carry out precise gene editing at single-nucleotide resolution without inducing DNA double-strand breaks (DSBs), which pose significant risks of genomic instability. Despite their outstanding advantages, the clinical application of BEs remains challenging due to their large size, which limits their efficient delivery, particularly in adeno-associated virus (AAV)-based systems. To address this issue, various strategies have been explored to reduce the size of BEs. These approaches include truncating the nonessential domains and replacing the bulky components with smaller substitutes without compromising the editing efficiency. In this review, we highlight the importance of downsizing BEs for therapeutic applications and introduce recent advances in size-reduction strategies. Additionally, we introduce the ongoing efforts to overcome other limitations of BEs, providing insights into their potential for improving in vivo gene editing. Full article

A20/Tnfaip3, an early NF-κB response gene and key negative regulator of NF-κB signaling, suppresses proinflammatory responses. Its ubiquitinase and deubiquitinase activities mediate proteasomal degradation within the NF-κB pathway. This study investigated the involvement of A20 signaling alterations in podocytes in the development of kidney injury. The phenotypes of A20Δpodocyte (podocyte-specific knockout of A20) mice were compared with those of control mice at 6 months of age to identify spontaneous changes in kidney function. A20Δpodocyte mice presented elevated serum urea nitrogen and creatinine levels, along with increased accumulation of inflammatory cells—neutrophils and macrophages—within the glomeruli. Additionally, A20Δpodocyte mice displayed significant podocyte loss. Ultrastructural analysis of A20 podocyte-knockout mouse glomeruli revealed hypocellularity of the glomerular tuft, expansion of the extracellular matrix, podocytopenia associated with foot process effacement, karyopyknosis, micronuclei, and podocyte detachment. In addition to podocyte death, we also observed damage to intracapillary endothelial cells with vacuolation of the cytoplasm and condensation of nuclear chromatin. A20 expression downregulation and CRISPR-Cas9 genome editing targeting A20 in a podocyte cell line confirmed these findings in vitro, highlighting the significant contribution of A20 activity in podocytes to glomerular injury pathogenesis. Finally, we analyzed TNFAIP3 transcription levels alongside genes involved in apoptosis, anoikis, NF-κB regulation, and cell attachment in glomerular and tubular compartments of kidney biopsies of patients with various renal diseases. Full article

CRISPR/Cas9, as a well-established gene editing technology, has been applied in numerous model organisms, but its application in wild-type E. coli remains limited. Pathogenic wild-type E. coli, a major cause of foodborne illnesses and intestinal inflammation in humans and animals, poses a significant global public health threat. The valine-glycine repeat protein G (VgrG) is a key virulence factor that enhances E. coli pathogenicity. In this study, PCR was used to identify 50 strains carrying the virulence gene VgrG2 out of 83 wild pathogenic E. coli strains, with only one strain sensitive to kanamycin and spectinomycin. A homologous repair template for VgrG2 was constructed using overlap PCR. A dual-plasmid CRISPR/Cas9 system, combining pTarget (spectinomycin resistance) and pCas (kanamycin resistance) with Red homologous recombination, was then used to induce genomic cleavage and knock out VgrG2. PCR and sequencing confirmed the deletion of a 1708 bp fragment of the VgrG2 gene in wild-type E. coli. IPEC-J2 cells were infected with E. coli-WT and E. coli ∆VgrG2, and treated with the mTOR inhibitor rapamycin to study the effects of VgrG2 on the mTOR signaling pathway. The qPCR results showed that VgrG2 activated the mTOR pathway, suppressed mTOR and p62 mRNA levels, and upregulated the autophagy-related genes and LC3-II protein expression. In conclusion, we utilized CRISPR/Cas9 technology to achieve large-fragment deletions in wild-type E. coli, revealing that VgrG2 activates the mTOR signaling pathway and upregulates autophagy markers. These findings offer new insights into E. coli genome editing and clarifies the pathogenic mechanisms through which VgrG2 induces cellular damage. Full article

Rice blast, caused by Magnaporthe oryzae, is a major threat to global rice production, necessitating the development of resistant cultivars through genetic improvement. Breakthroughs in rice genomics, including the complete genome sequencing of japonica and indica subspecies and the availability of various sequence-based molecular markers, have greatly advanced the genetic analysis of blast resistance. To date, approximately 122 blast-resistance genes have been identified, with 39 of these genes cloned and molecularly characterized. The application of these findings in marker-assisted selection (MAS) has significantly improved rice breeding, allowing for the efficient integration of multiple resistance genes into elite cultivars, enhancing both the durability and spectrum of resistance. Pangenomic studies, along with AI-driven tools like AlphaFold2, RoseTTAFold, and AlphaFold3, have further accelerated the identification and functional characterization of resistance genes, expediting the breeding process. Future rice blast disease management will depend on leveraging these advanced genomic and computational technologies. Emphasis should be placed on enhancing computational tools for the large-scale screening of resistance genes and utilizing gene editing technologies such as CRISPR-Cas9 for functional validation and targeted resistance enhancement and deployment. These approaches will be crucial for advancing rice blast resistance, ensuring food security, and promoting agricultural sustainability. Full article

The global nutraceutical industry is experiencing a paradigm shift, driven by an increasing demand for functional foods and dietary supplements that address malnutrition and chronic diseases such as obesity, diabetes, cardiovascular conditions, and cancer. Traditional plant- and animal-derived nutraceuticals face limitations in scalability, cost, and environmental impact, paving the way for microbial biotechnology as a sustainable alternative. Microbial cells act as bio-factories, converting nutrients like glucose and amino acids into valuable nutraceutical products such as polyunsaturated fatty acids (PUFAs), peptides, and other bioactive compounds. By harnessing their natural metabolic capabilities, microorganisms efficiently synthesize these bioactive compounds, making microbial production a sustainable and effective approach for nutraceutical development. This review explores the transformative role of microbial platforms in the production of nutraceuticals, emphasizing advanced fermentation techniques, synthetic biology, and metabolic engineering. It addresses the challenges of optimizing microbial strains, ensuring product quality, and scaling production while navigating regulatory frameworks. Furthermore, the review highlights cutting-edge technologies such as CRISPR/Cas9 for genome editing, adaptive evolution for strain enhancement, and bioreactor innovations to enhance yield and efficiency. With a focus on sustainability and precision, microbial production is positioned as a game-changer in the nutraceutical industry, offering eco-friendly and scalable solutions to meet global health needs. The integration of omics technologies and the exploration of novel microbial sources hold the potential to revolutionize this field, aligning with the growing consumer demand for innovative and functional bioactive products. Full article

Genome editing based on CRISPR-derived technologies has become a cornerstone in both fundamental research and clinical applications. Accurately measuring editing efficiency is crucial for developing and applying genome editing strategies. This study offers a detailed comparison of widely used techniques for evaluating on-target gene editing efficiency, including T7 Endonuclease I (T7EI), Tracking of Indels by Decomposition (TIDE), Inference of CRISPR Edits (ICE), droplet digital PCR (ddPCR), and live-cell assays involving engineered fluorescent reporter cells. Through a comparative analysis, this study highlights the unique strengths and limitations of each method, aiding researchers in choosing the most appropriate method for their specific needs, ensuring more tailored monitoring of genome editing outcomes in a precise and reliable manner. Full article

The genetic transformation of plants has provided fundamental insights into plant biology. However, the genetic transformation systems for most horticultural plants remain incomplete. Genome editing has significantly contributed to the improvement of crop traits, but it heavily relies on effective genetic transformation. Currently, reducing costs and improving the efficiency of genetic transformation are crucial for promoting the widespread application of genome editing in plants. Here, we review the advances in plant genetic transformation research, performing analysis of three methods for plant gene function analysis that bypass tissue culture: Agrobacterium rhizogenes-mediated root genetic transformation, developmental regulators (DRs)-mediated genetic transformation, and virus-mediated genome editing. We analyzed transformation efficiency in strawberry and citrus using the A. rhizogenes infiltration method, employing GFP to label different subcellular locations to investigate the morphology of microfilaments, nuclei, and peroxisomes in strawberry cells. Sequence analysis revealed that a series of developmental regulators critical for enhancing genetic transformation efficiency in specific species are highly conserved across different plant species. Additionally, we successfully edited the endogenous Pds gene in Cas9-overexpressing transgenic tobacco using TRV and CLBV containing the gRNA module. These three methods offer the benefits of being cost-effective and time-efficient, providing valuable technical insights for the application of plant genome editing. Full article

Thymus mongolicus (Lamiaceae) is a plant commonly found throughout China, in which it is widely used in chemical products for daily use, traditional medicinal preparations, ecological management, and cooking. In this study, we have assembled and annotated for the first time the entire mitochondrial genome (mitogenome) of T. mongolicus. The mitochondrial genome of T. mongolicus is composed in a monocyclic structure, with an overall size of 450,543 base pairs (bp) and a GC composition of 45.63%. It contains 32 unique protein-encoding genes. The repetitive sequences of the T. mongolicus mitogenome include 165 forward repetitive sequences and 200 palindromic repetitive sequences, in addition to 88 simple sequence repeats, of which tetramers accounted for the highest proportion (40.91%). An analysis of the mitogenome codons revealed that synonymous codons generally end with A/U. With the exception of nad4L, which uses ACG/ATG as an initiation codon, all other genes begin with the ATG start codon. Codon analysis of the mitogenome also showed that leucine (909) are the most abundant amino acid, while tryptophan (134) are the least prevalent. In total, 374 RNA editing sites were detected. Moreover, 180 homologous segments totaling 105,901 bp were found when the mitochondrial and chloroplast genomes of T. mongolicus were compared. Phylogenetic analysis further indicated that T. mongolicus is most closely related to Prunella vulgaris in the Lamiaceae family. Our findings offer important genetic insights for further research on this Lamiaceae species. To the best of our knowledge, this study is the first description of the entire mitogenome of T. mongolicus. Full article

CRISPR-Cas is an adaptive immune system found in bacteria and archaea that provides resistance against invading nucleic acids. Elements of this natural system have been harnessed to develop several genome editing tools, including CRISPR-Cas9. This technology relies on the ability of the nuclease Cas9 to cut DNA at specific locations directed by a guide RNA. In addition, the nuclease activity of Cas9 requires the presence of a short nucleotide motif (5′-NGG-3′ for Cas9 from Streptococcus pyogenes) called PAM, flanking the targeted region. As the reliance on this PAM is typically strict, diverse Cas9 variants recognising different PAM motifs have been studied to target a broader range of genomic sites. In this study, we assessed the potential of Cas9 from Streptococcus mutans strain P42S (SmutCas9) in gene editing. SmutCas9 recognises the rarely targeted 5′-NAA-3′ and 5′-NGAA-3′ PAMs. To test its efficacy, two genes of the virulent lactococcal phage p2 were edited, thereby demonstrating the potential of SmutCas9 for gene editing purposes, particularly in AT-rich genomes. Sequencing of total RNA also revealed the RNA components of this system, allowing further molecular characterisation of the type II-A CRISPR-Cas system of S. mutans. Full article

Background: An emerging trend of mutual convergence between drug-resistant and highly virulent strains of K. pneumoniae has been identified, highlighting the urgent need for the development of novel vaccines. Methods: To delete the target genes and eliminate the plasmids carrying antibiotic resistance genes, CRISPR-Cas9 technology was employed to perform genome editing on a clinically isolated O2 serotype of K. pneumoniae. Subsequently, this strain was utilized as a host to express genes associated with the synthesis of O1 serotype LPSs to construct the recombinant strain capable of simultaneously expressing LPSs of both O1 and O2 serotypes. This recombinant strain was then used as the production strain for the preparation of vaccines based on GMMAs (Generalized Modules for Membrane Antigens), and its biological characteristics were characterized. Finally, the safety and immunogenicity of the vaccine were evaluated using mice as the model animals. Result: a GMMA vaccine characterized by a high yield and low toxicity was gained. Importantly, the lipopolysaccharides (LPSs) of both O1 and O2 serotypes of K. pneumoniae were successfully expressed on the surface of the outer membrane vesicles. Following immunization with the GMMA vaccine, mice were capable of producing antibodies against the GMMA and demonstrated resistance to the invasion of both serotypes of clinically isolated K. pneumoniae. Conclusions: The GMMA vaccine showed significant promise as a bivalent vaccine against K. pneumoniae. Full article

Background: Breast cancer (BrCa) patients with tumors expressing high interleukin-6 (IL6) levels have poor clinical outcomes. In BrCa, altered occupancy of CCCTC-binding factor (CTCF) within its DNA binding sites deregulates the expression of its targeted genes. In this study, we investigated whether CTCF contributes to the altered IL6 expression in BrCa. Methods/Results: We performed CTCF gain- and loss-of-function assays in BrCa cell lines and observed an inverse correlation between CTCF and IL6 expression levels. To understand how CTCF negatively regulates IL6 gene expression, we performed luciferase gene reporter assays, site-directed mutagenesis assays, and chromatin immunoprecipitation assays. Our findings revealed that CTCF interacted with the IL6 promoter, a form of regulation disrupted in a CpG methylation-independent fashion in MDA-MB-231 and Tamoxifen-resistant MCF7 cells. Data from TCGA and GEO databases allowed us to explore the clinical implications of our results. An inverse correlation between CTCF and IL6 expression levels was seen in disease-free survival BrCa patients but not in patients who experienced cancer recurrence. Conclusions: Our findings provide evidence that the CTCF-mediated negative regulation of the IL6 gene is lost in highly tumorigenic BrCa cells. Full article

Trans-kingdom conjugation (TKC)/inter-domain conjugation is a horizontal gene transfer phenomenon that transfers DNA from eubacteria to eukaryotes and archaebacteria via a type IV secretion system encoded in IncP1-type broad-host-range plasmids. Although TKC is considered a potential gene introduction tool, donor chromosomal genes that influence TKC efficiency have rarely been analyzed, hindering targeted donor breeding. To identify potential TKC-related genes on a donor chromosome, a genome-wide screening of TKC-deficient mutants was performed using a comprehensive collection of Escherichia coli gene knockout mutants (Keio collection) as donors and a Saccharomyces cerevisiae strain as a recipient. Out of 3884 mutants, two mutants (∆aceE, ∆priA) showed a severe decrease in TKC efficiency by more than two orders of magnitude but not in bacterial conjugation. The effect on TKC efficiency by the two mutants was partly recovered by a preculture with a fresh culture medium before the TKC reaction, regardless of the presence of antibiotics. These results suggest that no single chromosomal target gene is solely responsible for universally blocking IncP1-type conjugation by impeding its function. The results also suggest the existence of an unidentified recognition or transfer mechanism distinct from bacterial conjugation, highlighting the novel roles of aceE and priA. Full article

Prunus cerasoides D.-Don is a rare winter-blooming species and a distinctive and potential germplasm resource for cherry blossoms. We have characterized the mitochondrial genome (mitogenome) of P. cerasoides and acquired a monocyclic molecule measuring 421,258 bp. A total of 58 unique genes were annotated, comprising 36 protein-coding genes, 19 tRNAs, and three rRNAs. In the mitochondrial genome of P. cerasoides, we detected 86 simple sequence repeats, 727 dispersed repeats, and 21 tandem repeats. We detected 456 RNA editing sites from 34 unique protein-coding genes, leading to the cytosine to uracil transitions. Collinear analysis revealed that the mitogenome of P. cerasoides is quite conservative among species of the subgenus Cerasus. Moreover, our study detected 26 segments of plastid genomic DNA that had transferred from the plastome to the mitogenome. Six genes were found to be completely transferred from these fragments. The maximum likelihood phylogenetic analysis utilizing the mitogenomes of 29 distinct Rosaceae species supports the classification of P. cerasoides into separate branches. Comprehending the mitochondrial genomic characterization of P. cerasoides is crucial for elucidating its genetic foundation and offers insights into evolutionary relationships within the Prunus species. Full article

Peanut allergies affect millions of people worldwide, often causing life-threatening reactions and necessitating strict avoidance. Recent advancements in oral immunotherapy, such as Palforzia™, and IgE-mediated treatments (e.g., Xolair), have improved care options; however, their high costs limit accessibility and widespread utility. To address these challenges, researchers are employing conventional breeding and advanced molecular tools, such as CRISPR editing, to develop peanut lines with reduced levels of major allergenic proteins (Ara h1, Ara h2, Ara h3, and Ara h6). These reduced-immunogenicity genotypes retain their agronomic viability, flavor, and nutritional quality to some extent, offering the potential for cost-effective oral immunotherapy and safe food options for use in public spaces by non-allergic individuals. Rigorous evaluation, including immunological assays and human feeding trials, is essential to confirm their effectiveness in reducing allergic reactions. Adoption will depend on the establishment of clear regulatory guidelines, stakeholder education, and transparent communication of the benefits and risks. With sustained research, public trust, and supportive policies, reduced-immunogenicity peanuts could substantially lower the global burden of peanut allergies. This communication examined the impact of peanut allergies worldwide and explored strategies to develop peanut genotypes with reduced allergen content, including conventional breeding and advanced genetic engineering. It also addressed the challenges associated with these approaches, such as policy and regulatory hurdles, and outlined key requirements for their successful adoption by farmers and consumers. Full article