Search Results (550)

Drought seriously affects wheat yield; it is therefore important to study the molecular mechanism of wheat resistance to drought stress to ensure national food security. Plants can remove harmful substances through autophagy, thus improving their drought resistance. The results of previous studies have shown that autophagy is involved in the drought stress response; however, the molecular mechanism of autophagy in response to drought stress has yet to be elucidated. In this study, molecular biological methods such as immunohistochemistry, Co-Immunoprecipitation (Co-IP), and pull-down were used to explain the molecular mechanism of autophagy in response to drought stress at the protein level. We found that a dehydrin protein called cold-regulated 410 (TaCOR410) interacts with autophagy-related 8 (TaATG8, a key protein of wheat autophagy). TaCOR410 interacted with TaATG8 through its ATG8-interacting motif (AIM), and interaction was inhibited after mutation of the AIM. Interference with TaCOR410 inhibited autophagy and reduced the drought resistance of wheat. In contrast, transient transfection of TaCOR410 promoted autophagy. In wheat, overexpression of TaATG8 improved the drought resistance of wheat. Following interference with TaATG5, TaATG7 inhibited autophagy and reduced the drought resistance of wheat. From the above results, it is evident that autophagy can improve the drought resistance of wheat and can respond to drought stress through the interaction of TaCOR410 with TaATG8. Full article

Rosa roxburghii Tratt., a fruit crop known for its high Vitamin C content and other nutritional compounds, has not yet been studied for its auxin response factor (ARF) family members. ARFs are important proteins in auxin-mediated pathways, playing a vital role in plant physiological and biochemical processes such as plant development, and flower and fruit maturation. In the present study, we identified 14 ARF genes (designated as RrARFs) in R. roxburghii, which are distributed across seven chromosomes and grouped into four subfamilies. An analysis of cis-acting elements revealed that these genes might be involved in various biological processes, including plant development, flower development, light responses, cell cycle regulation, phytohormone responses, and responses to abiotic and biotic stresses. A gene expression analysis demonstrated differential expression of RrARF genes across different tissues and stages of fruit development, with four members showing higher expression during the fruit ripening stages. Furthermore, a coexpression analysis identified that RrARF5 was highly coexpressed with RrMDHAR1, a key enzyme involved in Vitamin C biosynthesis. Moreover, transactivation assays and transient overexpression experiments confirmed that RrARF5 activates the transcription of RrMDHAR1. The findings of this study suggest a potential role of the ARF gene family in Vitamin C accumulation in R. roxburghii and enhance our understanding of the diverse regulatory function of the ARF gene family in plants. Full article

The β-glucosidases (BGLUs) of Glycoside Hydrolase Family 1 (GH1) exhibit essential functions in plant secondary metabolism and stress responses, mediated by their dual catalytic capabilities in hydrolysis and transglycosylation. This study identified 149 BGLU family members within B. napus (Brassica napus L.), which were systematically categorized into 10 distinct subgroups. Subsequent characterization encompassed detailed examination of their motif composition, chromosomal distribution, gene collinearity, selection pressure, and expression profiling. Transient overexpression of BnBGLU77 in N. benthamiana (Nicotiana benthamiana), combined with untargeted metabolomics analysis, revealed pronounced modulatory effects on the degradation and accumulation of β-glucosidic compounds, suggesting potential roles of the protein encoded by BnBGLU77 in metabolic homeostasis and stress response mechanisms. These experimental results first validated the bidirectional catalytic activity of a BGLU enzyme in B. napus, while simultaneously advancing fundamental understanding of BnBGLU gene functions and providing new insights for developing stress-resistant rapeseed cultivars through targeted genetic improvement. Full article

The ABI3 transcription factor is a key regulator in plant growth and development. Through transcriptome analysis of the resistant soybean cultivar ‘Dongnong L10′ and the susceptible cultivar ‘Heinong 37′ exposed to soybean cyst nematode race 3 (SCN 3) stress, the differentially expressed gene GmABI3VP1 was identified. The GmABI3VP1 gene was then cloned and analyzed through bioinformatics, subcellular localization, and qRT-PCR analysis of resistant and susceptible soybean germplasms, as well as overexpression and gene editing of soybean hairy roots followed by SCN 3 identification analysis. It was found that the protein encoded by GmABI3VP1 is an acidic and hydrophilic protein with transmembrane domains. It has a collinear relationship with Arabidopsis and is widely distributed in plants. Through the analysis of promoter elements, it was shown that this gene contains multiple hormone-responsive promoter elements like ABRE/ABRE3a/ABRE/4a/as-1 and stress-responsive elements such as Myb/MYC/MYc. Transient expression in tobacco indicated that the GmABI3VP1 gene is located in the nucleus. The transcription of GmABI3VP1 responds to the stress of SCN, and its transcriptional level is relatively high in the roots of resistant materials. Genetic transformation mediated by Agrobacterium rhizogenes was used to obtain GmABI3VP1 gene overexpressed and CRISPR-Cas9 gene-edited soybean hairy roots. In comparison to the wild type (WT), the density of nematodes per area was notably lower in hairy roots overexpressing (OX) the gene, whereas the density of SCN per unit area (per cm of lateral root length) significantly increased in gene-edited (KO) soybean hairy roots. Through SCN phenotyping, GmABI3VP1 was identified as a contributor to SCN 3 resistance. This study provides initial insights into the role of the GmABI3VP1 gene in SCN resistance, establishing a robust basis for future research on the mechanisms underlying SCN disease resistance and offering valuable genetic reservoirs for SCN 3 resistance. Full article

Squamosa Promoter-Binding Protein Like (SPL) is a critical transcription factor that plays a significant role in regulating plant growth and development. Mining the coconut SPL family offers valuable insights into the regulation of important agronomic traits, including the length of the juvenile phase. In this study, 25 CnSPLs were identified and were classified into eight subfamilies. Analysis of gene structure and conserved protein motifs indicated a high conservation of CnSPLs within the same subfamilies; however, variations in protein structure and gene length were observed across different subfamilies. Gene expansion analysis indicated that most gene members within subfamilies originated from duplications of the same genomic segment, and transposable element insertion contributed to the divergence of gene sequences within these subfamilies. Characterization of the miR156 target sequence in SPL transcripts revealed that subfamilies IV to VIII contained these sequences, while subfamilies I to III did not. In both coconut and 14 other plant species, some SPLs lost their miR156-binding loci due to gene structure variations. The gene expression profiles revealed significant divergence between miR156-targeted and non-targeted CnSPLs; the former exhibited low expression levels in the endosperm, while the latter showed comparable expression across all tissues. Notably, CnSPL15A demonstrated steadily increasing expression levels in leaves throughout successive leaf primordia and significantly promoted flowering when overexpressed in Arabidopsis. Transient expression assays and 5′ RACE confirmed that CnSPLs are targeted by miR156. This study establishes a foundation for investigating the evolutionary characteristics of CnSPLs and provides a theoretical framework for analyzing the functions of key CnSPLs involved in the coconut flowering control pathway. Full article

Paeonia ostii is an economically significant species serving as an ornamental, medicinal herb, and woody oilseed crop. Gene function elucidation and molecular breeding are hindered by the lack of efficient, stable transformation methods due to tissue culture challenges. To enable year-round functional studies without material constraints, we established a novel transient transformation system mediated by Agrobacterium using in vitro embryo-derived seedlings (TTAES) in P. ostii. By optimizing embryo germination media, we achieved consistent seedling production. Orthogonal experiments with a GUS reporter identified optimal conditions: OD₆₀₀ = 1.0, 200 μM of acetosyringone, six negative-pressure treatments, and 2 h infection. Under this optimized system, maximum transformation efficiency was achieved at 35 days after germination. With this system, we demonstrated its application in investigating transcription factor-mediated regulation of target gene promoters using GUS as a reporter gene. To achieve non-destructive identification of transiently transformed plants, we employed GFP as a reporter gene. Using transient expression of VIGS (knockdown) and 35S constructs (overexpression), we characterized gene functions, thereby confirming the system’s effectiveness for functional analysis. This system facilitates the acquisition of plant experimental materials and significantly improves research efficiency for year-round gene function elucidation in P. ostii. Full article

Chrysanthemum morifolium Ramat. is an important ornamental plant, holding dual economic value as a medicinal and edible plant. Jinsi Huangju is a popular healthy tea drink prepared from the large and elegant shaped flowers of C. morifolium. However, the suboptimal accumulation of bioactive flavonoids during conventional harvest (full bloom stage) limits its commercial potential. To elucidate the molecular mechanisms governing flavonoid biosynthesis in Jinsi Huangju flowers and identify key genetic regulators for metabolic engineering, we performed integrated metabolomic and transcriptomic analyses of flowers at distinct developmental stages using ultra-performance liquid chromatography–tandem mass spectrometry (UPLC-MS/MS) and RNA-seq. Differential metabolites were screened, and candidate genes were validated via transient transformation assays. Among 2146 identified metabolites, flavonoids were the predominant differential compounds, with accumulation patterns being strongly stage dependent. Thirty-eight flavonoid biosynthetic genes and key transcription factors from the MYB, bHLH, and WD40 families exhibited dynamic expression. The CmMYB8a was confirmed as a positive regulator of flavonoid biosynthesis through transient overexpression. This study deciphers the stage-specific flavonoid accumulation in Jinsi Huangju and identifies CmMYB8a as a pivotal regulatory target. Our findings provide genetic resources for breeding high-flavonoid cultivars via molecular design. Full article

Gray mold disease, caused by Botrytis cinerea, severely impacts grape production worldwide. Although proanthocyanidins (PAs) contribute to fungal pathogen resistance, their role in grape defense against B. cinerea remains unclear. Here, we demonstrate that VvMYBPA1, a key transcriptional regulator of PA biosynthesis, negatively modulates B. cinerea resistance in grape berries. While infection suppressed endogenous VvMYBPA1, its agroinfiltration-mediated transient overexpression in berries elevated susceptibility, paralleling reduced β-1,3-glucanase (BGL) and polyphenol oxidase (PPO) activities. Additionally, VvMYBPA1 overexpression elevated VvRBOHs’ expression and reduced peroxidase (POD) activity, resulting in excessive hydrogen peroxide (H₂O₂) accumulation and more cell death. Our results reveal that VvMYBPA1 negatively regulates B. cinerea resistance by disrupting antioxidant enzyme activity and ROS homeostasis, providing new insights into the interplay between PA biosynthesis and fungal defense mechanisms. Full article

Pinus massoniana Lamb. is an economically important conifer native to China. However, it is highly susceptible to the pine wood nematode (Bursaphelenchus xylophilus, PWN), the causal agent of pine wilt disease (PWD), resulting in substantial ecological and economic losses. To elucidate potential molecular defense mechanisms, 50 NAC (NAM, ATAF1/2, and CUC2) transcription factors (PmNACs) were identified in the P. massoniana genome. Phylogenetic analysis divided these PmNACs into seven subfamilies, and motif analysis identified ten conserved motifs associated with stress responses. Twenty-three genes were selected for expression analysis in various tissues and under exogenous salicylic acid (SA), methyl jasmonate (MeJA), and PWN infection. Six genes (PmNAC1, PmNAC8, PmNAC9, PmNAC17, PmNAC18, and PmNAC20) were significantly up-regulated by both hormonal treatment and PWN infection, implying their involvement in JA/SA-mediated immune pathways. Functional characterization showed PmNAC8 is a nuclear-localized transcription factor with autoactivation activity. Furthermore, transient overexpression of PmNAC8 in Nicotiana benthamiana induced reactive oxygen species (ROS) accumulation and necrotic lesions. Collectively, these results elucidate NAC-mediated defense responses to PWN infection in P. massoniana and identify candidate genes for developing PWD-resistant pine varieties. Full article

Radish (Raphanus sativus), a commonly grown root vegetable prized for its nutrition and culinary use, is particularly vulnerable to lead (Pb) stress, which mainly results in Pb accumulation in the roots. However, the molecular mechanisms underlying Pb accumulation in radish remain largely unknown. In this study, we investigated the role of BASIC PENTACYSTEINE (BPC) genes in radish’s response to Pb stress. Phylogenetic analysis revealed that radish contains 10 BPC genes, which are distinctly clustered in Cluster III. Expression analysis revealed that, except for RsBPC2, RsBPC4, and RsBPC7, the expression of most RsBPC genes was significantly altered under Pb stress. Notably, the expression of RsBPC5 gradually decreased with prolonged Pb exposure. Subcellular localization analysis confirmed that RsBPC5 is localized in the nucleus and acts as a transcriptional repressor. Functional assays demonstrated that transient overexpression of RsBPC5 enhanced the tolerance of radish plants to Pb stress via reducing Pb accumulation and activating the antioxidant defense system. Collectively, our findings suggest that RsBPC5 plays a key role in radish’s response to Pb stress, potentially improving Pb tolerance by modulating Pb uptake and strengthening antioxidant defense mechanisms. Full article

Adoptive cell therapies have transformed the treatment landscape for hematologic malignancies. Yet, translation to solid tumors remains constrained by antigen heterogeneity, an immunosuppressive tumor microenvironment (TME), and poor persistence of conventional CAR-T cells. In response, innate immune cell platforms, particularly chimeric antigen receptor–engineered natural killer (CAR-NK) cells and chimeric antigen receptor–macrophages (CAR-MΦ), have emerged as promising alternatives. This review summarizes recent advances in the design and application of CAR-NK and CAR-MΦ therapies for solid tumors. We highlight key innovations, including the use of lineage-specific intracellular signaling domains (e.g., DAP12, 2B4, FcRγ), novel effector constructs (e.g., NKG7-overexpressing CARs, TME-responsive CARs), and scalable induced pluripotent stem cell (iPSC)-derived platforms. Preclinical data support enhanced antitumor activity through mechanisms such as major histocompatibility complex (MHC)-unrestricted cytotoxicity, phagocytosis, trogocytosis, cytokine secretion, and cross-talk with adaptive immunity. Early-phase clinical studies (e.g., CT-0508) demonstrate feasibility and TME remodeling with CAR-MΦ. However, persistent challenges remain, including transient in vivo survival, manufacturing complexity, and risks of off-target inflammation. Emerging combinatorial strategies, such as dual-effector regimens (CAR-NK⁺ CAR-MΦ), cytokine-modulated cross-support, and bispecific or logic-gated CARs, may overcome these barriers and provide more durable, tumor-selective responses. Taken together, CAR-NK and CAR-MΦ platforms are poised to expand the reach of engineered cell therapy into the solid tumor domain. Full article

The establishment of a transient expression system in petals is significant for elucidating gene functions in flowering trees characterized by a prolonged juvenile phase. Genetic improvements in Camellia japonica have been hindered due to the absence of a functional validation platform. In this study, we explored an Agrobacterium-mediated and readily observable transient expression system in camellia petals to systematically optimize four critical factors affecting transformation efficiency. As a result, the bud stage, ‘Banliuxiang’ genotype, OD600 of 1.0, and 1-day co-cultivation achieved the highest intensity of transient expression, and overexpression of the Ruby1 reporter gene induced substantial anthocyanin synthesis, manifested as distinct red pigmentation. Furthermore, the optimized transient expression system revealed that the R2R3-MYB transcription factor CjMYB5, which interacted with CjGL3, promoted anthocyanin biosynthesis in camellia petals by transactivating key DFR structural genes. This transient expression platform not only advances functional genomics studies in ornamental woody species but also lays a foundation for molecular breeding programs in C. japonica. Full article

The BAHD acyltransferase family plays a critical role in plant secondary metabolism by catalyzing acyl transfer reactions that are essential for synthesizing metabolites involved in environmental adaptation. However, systematic investigation of this superfamily in Brassica napus has not been reported. In this study, 158 BnaBAHD genes were identified by comprehensive analyses of evolutionary relationships, motif structures, chromosomal distribution, gene collinearity, and selection pressures, and these genes were phylogenetically classified into five clades harboring conserved catalytic domains (HXXXD and DFGWG). Transient overexpression combined with metabolomic profiling demonstrated that two homologous seed-specific Clade V members, BnaBAHD040 and BnaBAHD120, which exhibited elevated expression during late seed development, significantly enhanced the accumulation of acylated metabolites contributing to biotic/abiotic stress resistance. This study provides the first experimental validation of the catalytic functions of BAHD enzymes in B. napus, establishing a theoretical foundation for leveraging this gene family in genetic improvement to develop novel rapeseed cultivars with enhanced stress tolerance and yield. Full article

Fusarium wilt, caused by Fusarium solani, is a devastating disease that leads to significant losses in ginger (Zingiber officinale) crops worldwide. To explore the molecular mechanisms underlying F. solani infection and disease progression, we performed a comparative transcriptome analysis of ginger rhizomes during storage, comparing inoculated and non-inoculated samples. A total of 647 and 6398 DEGs were identified in the 1.5- and 2-day infection groups, respectively. KEGG analysis revealed that most DEGs were enriched in the plant–pathogen interaction pathway, with both PTI and ETI being activated. Six DEGs in this pathway were validated by qRT-PCR at two time points, showing a strong correlation with FPKM values from the transcriptome data. Furthermore, transient expression analysis in Nicotiana benthamiana leaves demonstrated that overexpressing ZoCEBiP1 helped scavenge excess ROS, thereby reducing disease severity. Transcriptional profiling of DEGs in the plant–pathogen interaction pathway revealed significant changes in genes involved in ROS and NO metabolism. In F. solani-infected ginger rhizomes, levels of H₂O₂ and O₂⁻ were elevated, along with increased activities of antioxidant enzymes (POD, CAT, SOD, and APX) and higher NO content and NOS activity. These findings elucidated the early defense response of ginger rhizomes to F. solani infection and provided insights for developing effective strategies to manage fungal diseases. Full article

Introduction: The interface between T cells and the tumor microenvironment, termed the ‘immunological synapse’, consists of multiple checkpoint protein pairs co-expressed on both sides of the synapse. mir-16, a microRNA from a widely known tumor-suppressor family of miRNAs, was previously shown by us to be downregulated in melanoma. As other miRNAs from this family have been shown to directly target checkpoint proteins, here we investigated whether miR-16 influences the expression patterns of checkpoint proteins in melanoma. Methods: Single-cell gene expression data from the melanoma microenvironment were retrieved from a public database. Melanoma cell lines were established from metastatic lesions and transiently transfected with an hsa-miR-16-5p-mimic RNA or a mir-16-expressing plasmid. The mRNA expression profiles were analyzed using an Affymetrix microarray. Direct targets of miR-16 were identified by luciferase reporter assays. Protein levels were assessed by Western blotting. Results: Bioinformatic analysis revealed that the expression levels of eight checkpoint mRNAs, known to be present on the melanoma side of the immunological synapse, were highly correlated. Four of these mRNAs contained putative binding sites for the miR-15/16 family. miR-16 expression was significantly reduced in melanoma cells, compared to normal melanocytes. Luciferase reporter assays demonstrated that miR-16 directly targets the 3′ untranslated regions (3′UTRs) of CD40, CD80. The mRNAs downregulated following miR-16 overexpression were highly enriched for genes involved in autophagy, vesicle-mediated transport, and the regulation of protein membrane localization. Among these, VTI1B and SMPD1 were confirmed to be direct targets of miR-16. Transient overexpression of miR-16 resulted in a significant reduction in SMPD1 and VTI1B levels in melanoma cell lines. Conclusions: Our findings suggest that miR-16 potentially modulates melanoma tumorigenesis, metastasis and immunogenicity by altering the composition of checkpoint proteins at the immunological synapse and by regulating cellular pathways associated with intracellular trafficking and transmembrane protein presentation. Full article