Search Results (334)

Background/Objectives: Recent advances in immunotherapy have revolutionized cancer treatment, as exemplified by multiple monoclonal antibodies against programmed cell death protein 1 (PD-1) or programmed death-ligand 1 (PD-L1). Nevertheless, immunotherapeutic antibodies exhibit certain limitations, which drives the development of alternative approaches, such as small interfering RNA (siRNA)-based therapeutics. The aim of this study was to design and produce new biological PD-L1 siRNA (BioRNA/PD-L1-siRNA) molecules and further define their immunotherapeutic efficacy against non-small cell lung cancer (NSCLC) in vitro. Methods: A novel RNA molecular bioengineering platform was employed to produce new BioRNA/PD-L1-siRNA agents. The functions of BioRNA/PD-L1-siRNAs were determined by quantitative PCR, Western blot, immunofluorescence confocal imaging, flow cytometry, and PD-1/PD-L1 blockade assays in human NSCLC cells, alone and co-cultured with human peripheral blood mononuclear cells (PBMCs). Results: After heterologous overexpression and purification of five BioRNA molecules, one siRNA named BioRNA/PD-L1-siRNA-1 was identified as the most effective to selectively suppress human PD-L1 mRNA and protein levels in H460 and H1975 cells. Disruption of PD-1/PD-L1 interactions by BioRNA/PD-L1-siRNA-1 was further demonstrated via a PD-1/PD-L1 blockade bioassay. In addition, the immunomodulatory effectiveness of BioRNA/PD-L1-siRNA-1 was established in co-culture models, as indicated by the induction of T-cell and natural killer cell populations and an increase in specific cytokines and cytotoxic granules, and subsequent enhancement of apoptosis and greater inhibition of NSCLC cell viability. Conclusions: Overall, these findings demonstrate the potential of bioengineered PD-L1 siRNA entities for NSCLC immunotherapy. Full article

Verticillium dahliae is a devastating soil-borne fungal pathogen that causes severe yield losses in melon (Cucumis melo L.) and other crops. Identifying novel resistance genes is crucial for sustainable disease management. In this study, we characterized the function of CmVQ23, a candidate gene previously identified through QTL mapping, in mediating defense against V. dahliae using heterologous expression in Arabidopsis thaliana. Subcellular localization assays revealed that the CmVQ23-eGFP fusion protein predominantly localized to the nucleus, consistent with its predicted role as a co-factor of transcription factor. Upon V. dahliae inoculation, CmVQ23-overexpressing Arabidopsis lines exhibited significantly reduced disease indices and restricted fungal proliferation compared with wild-type and mutant plants, although these lines displayed altered vegetative growth, including delayed bolting and reduced plant height. Mechanistically, CmVQ23 overexpression promoted reactive oxygen species (ROS) accumulation and hypersensitive response (HR)-mediated cell death at infection sites, as evidenced by intensified DAB and trypan blue staining. Furthermore, transgenic lines maintained higher photosynthetic efficiency, enhanced antioxidant enzyme activities, and increased lignin deposition via upregulation of phenylalanine ammonia-lyase (PAL) and polyphenol oxidase (PPO). Notably, CmVQ23 overexpression markedly upregulated both salicylic acid (SA)- and jasmonic acid/ethylene (JA/ET)-responsive marker genes, including AtPR1, AtPR2, AtPR5, AtPAD4, AtPDF1.2, and AtVSP2 upon infection. Collectively, these findings demonstrate that CmVQ23 functions as a positive regulator of resistance to Verticillium dahliae by orchestrating ROS/HR-mediated cell death, antioxidant defense, phenylpropanoid pathway activation, and phytohormone signaling crosstalk, offering a promising genetic resource for improving Verticillium wilt resistance in crops. Full article

Efficient saccharification of lignocellulose, the most abundant renewable carbon reservoir resource, is of great industrial importance. Trichoderma reesei is a premier cellulase producer, but its fermentation efficiency is often constrained by dual challenges: dissolved oxygen limitation and intrinsic oxidative stress. To address this, we engineered T. reesei to heterologously express a robust catalase gene (cat-3) from Neurospora crassa. The recombinant strain Tr-cNcat3 exhibited a 7.4-fold increase in extracellular catalase activity. Tr-cNcat3 showed an increase in total extracellular protein, resulting in markedly enhanced filter paper activity (FPA) and β-glucosidase activity compared to the control. Strikingly, this intervention specifically triggered a significantly higher expression of β-glucosidase, a known bottleneck in T. reesei’s cellulase system, particularly on bagasse and straw as the carbon source. Moreover, the ability of the supernatant to degrade cellulose substrates was improved. Our results reveal that overexpression of cat-3 in T. reesei could modify the cellulase cocktail by triggering a higher level of β-glucosidase. This study provides a novel and effective genetic engineering strategy to unlock the full industrial potential of T. reesei for cost-effective lignocellulosic biorefining. Full article

China’s edible oil self-sufficiency rate is only about 30%, and approximately 100 million acres of saline-alkali land are potentially available for rapeseed cultivation. However, salt stress severely inhibits rapeseed growth and yield, making the breeding of salt-tolerant varieties and the study of underlying molecular mechanisms urgent priorities. In this study, 1609 Brassica napus accessions were screened for salt tolerance, and the highly salt-tolerant material ‘Xiangnong Saline-Alkali Oil No. 1’ (XNSA01) was identified. Subsequent physiological, hormonal, transcriptomic, and functional analyses were conducted to characterize its salt-tolerance mechanisms. Salt stress significantly activated antioxidant defense and stress-related hormone responses in XNSA01, accompanied by significant accumulation of proline, abscisic acid, and salicylic acid. Transcriptome analysis identified five candidate salt-responsive genes, among which BnaA10g15320D, BnaA02g04730D, and BnaC07g08360D were closely associated with antioxidant enzyme activities and endogenous hormone contents. In particular, BnaA02g04730D showed a significant negative correlation with MDA content. Yeast heterologous expression and transgenic validation further demonstrated that BnaA02g04730D enhanced salt tolerance by increasing antioxidant enzyme activities and reducing membrane lipid peroxidation. These findings provide a useful basis for the breeding of salt-tolerant rapeseed varieties and for further investigation of salt-tolerance mechanisms in B. napus. Full article

Drought stress severely limits grapevine (Vitis spp.) productivity, yet the regulatory mechanisms distinguishing drought-tolerant and drought-sensitive cultivars remain insufficiently understood. In this study, we integrated transcriptomic, metabolomic, physiological, and functional analyses to compare drought responses between the tolerant cultivar ‘Miguang’ and the sensitive cultivar ‘Red Globe’, together with previously generated datasets from ‘Shine Muscat’ and ‘Thompson Seedless’. Compared with sensitive cultivars, tolerant cultivars showed stronger antioxidant capacity, lower lipid peroxidation, and more coordinated changes in pathways related to redox balance, osmotic adjustment, and energy metabolism. In ‘Miguang’, drought responses were associated with activation of the pentose phosphate pathway, accumulation of tricarboxylic acid cycle intermediates, and genotype-specific alternative splicing events affecting metabolic and signaling genes. Comparative analysis across four cultivars identified VvGRIK1 as a conserved drought-responsive regulator associated with redox and bioenergetic homeostasis. Heterologous overexpression of VvGRIK1 in tobacco enhanced drought-related physiological performance by increasing antioxidant enzyme activities and reducing membrane damage. Yeast two-hybrid assays and molecular docking further suggested a potential interaction between VvGRIK1 and VvKING1, a SnRK1-related energy sensor. Together, these findings suggest that the VvGRIK1-SnRK1 module may contribute to drought adaptation by coordinating redox protection and energy homeostasis, providing a candidate regulatory target for future functional studies and grapevine molecular breeding. Full article

Maize (Zea mays L.) productivity is severely constrained by drought stress. Although the maize SET domain-containing gene family has previously been investigated, the earlier analysis was based on the incomplete B73 RefGen_v2 genome assembly, and the drought-responsive functions of individual ZmSET members remain largely uncharacterized. In this study, 47 ZmSET genes were identified using the updated B73 RefGen_v5 genome and systematically analyzed for their physicochemical properties, chromosomal distribution, gene structures, conserved motifs, and promoter cis-acting elements. The ZmSET family exhibited substantial evolutionary conservation, while its promoters contained numerous stress- and hormone-responsive elements. Transcriptome analysis identified ZmSET9 as a drought-responsive gene, and RT-qPCR showed that it maintained relatively high expression throughout drought treatment. Heterologous overexpression of ZmSET9 in Arabidopsis thaliana enhanced drought tolerance and supported plant growth under drought stress. Compared with wild type plants, the transgenic lines exhibited higher superoxide dismutase (SOD), catalase (CAT), and peroxidase (POD) activities and lower malondialdehyde (MDA) contents, indicating enhanced antioxidant capacity and reduced membrane lipid peroxidation. Under PEG induced osmotic stress, AtCAT1 and AtMYC2 were more strongly induced in the transgenic lines. Protein–protein interaction prediction and yeast two-hybrid assays further demonstrated that ZmSET9 physically interacts with FERTILIZATION-INDEPENDENT ENDOSPERM 1 (FIE1), a core component of Polycomb repressive complex 2. These findings update the genomic characterization of the maize SET family and suggest that ZmSET9 contributes to drought tolerance by enhancing antioxidant defense and regulating stress-responsive gene expression. Full article

Salt stress represents a major constraint on the productivity of lotus (Nelumbo nucifera), an aquatic crop of significant ornamental, nutritional, and medicinal value. Although aquaporins (AQPs) are key regulators of plant water and solute homeostasis, the genomic organization and stress-responsive functions of this gene family in lotus remain largely uncharacterized. In this study, a total of 32 NnAQP genes were identified in the lotus genome and classified into five subfamilies. Segmental duplication was identified as the primary driver of family expansion under strong purifying selection. Expression profiling revealed broad transcriptional activity across various tissues, and qRT-PCR analysis demonstrated that NnPIP2-7 was significantly and sustainedly induced under salt stress. Subcellular localization confirmed that NnPIP2-7 is targeted to the plasma membrane. Heterologous overexpression of NnPIP2-7 in Arabidopsis thaliana significantly improved seed germination and primary root elongation under saline conditions. Crucially, transgenic plants exhibited enhanced osmotic adjustment and elevated antioxidant capacity, resulting in reduced oxidative damage compared to wild-type plants. These findings establish the evolutionary trajectory of the lotus AQP family and demonstrate the core regulatory role of NnPIP2-7 in plant salt stress adaptation. Full article

Zinc/iron-regulated transporter proteins (ZIPs) are involved in the transport of zinc and iron, maintenance of ion homeostasis, and regulation of plant responses to abiotic stresses. Although previous studies have identified members of the ZIP gene family and screened for alkaline-responsive ZIP genes in wild soybean (Glycine soja), the functional implications of these genes under alkaline stress conditions remain to be fully elucidated. The present study aimed to identify the GsZIP7 gene and investigate its regulatory role in Arabidopsis and wild soybean under alkaline stress. The results showed that GsZIP7 was highly expressed in roots and young stems. GsZIP7 exhibited differential expression in response to alkaline stress, zinc deficiency, or iron deficiency in wild soybean. In addition, heterologous expression of GsZIP7 in yeast mutants increased tolerance to iron- and zinc-deficient conditions. Overexpression of GsZIP7 in Arabidopsis resulted in reduced root growth, decreased fresh weight, and a weakened antioxidant defense system. Furthermore, GsZIP7-overexpressing soybean hairy roots showed increased sensitivity to alkaline stress, whereas GsZIP7-RNAi lines exhibited enhanced tolerance. Together, these findings indicate that GsZIP7 negatively regulates alkaline tolerance in plants. Full article

CONSTANS-LIKE (COL) genes are pivotal regulatory components in the photoperiodic flowering pathway of plants. These genes can be regulated by both photoreceptors and the circadian clock, modulating plant flowering responses under specific day lengths by regulating florigen levels. However, the COL gene family in Litchi chinensis Sonn. has not yet been characterized. In this study, we identified eight COL family members in litchi and classified them into three subgroups based on phylogenetic analysis. The analysis of cis-regulatory elements within the promoters of LcCOLs revealed a wide distribution of elements associated with light, hormone, and stress responses. Transcript expression profiling indicated that most LcCOLs exhibited relatively high expression levels in leaf buds, leaves, and young fruits. Diurnal expression analysis under natural photoperiod conditions revealed that the expression peaks of all LcCOLs, with the exception of LcCOL1, occurred during the nighttime. The heterologous overexpression of LcCOL7 and LcCOL8, the closest homologs to AtCO, in Arabidopsis thaliana significantly delayed the flowering time under both long-day (LD) and short-day (SD) conditions, indicating that these genes act as repressors of flowering. This study provides a foundational basis for elucidating the molecular mechanisms underlying litchi flowering regulation and identifies promising candidate genes for the molecular breeding of litchi flowering-related agronomic traits. Full article

Sea Island cotton (Gossypium barbadense L.) is a premium raw material for high-end textiles due to its excellent fiber quality. The AP2/ERF transcription factor family plays critical roles in plant growth and hormone signaling. Here, 161 GbERF family members were identified in Sea Island cotton and classified into nine subgroups, with GbERF13 belonging to Group V. Expression analysis revealed that GbERF13 was specifically and highly expressed in fibers, with transcript abundance peaking at 15–30 days post-anthesis (DPA), coinciding with the transition from fiber elongation to secondary wall thickening. Exogenous abscisic acid (ABA) treatment significantly induced GbERF13 expression and inhibited fiber elongation. Heterologous overexpression of GbERF13 in Arabidopsis increased trichome and root hair numbers while suppressing primary root growth, confirming its role in cell elongation and development. A nonsynonymous SNP (A/C) at the 117th base pair of the GbERF13 coding region (GbERF13-117SNP) was identified in 213 Sea Island cotton accessions. Association analysis showed the C allele was significantly and positively associated with fiber length, strength, and uniformity. An allele-specific PCR marker was further developed for molecular breeding. Collectively, GbERF13 acts as a key ABA-responsive transcription factor regulating fiber development, and its functional SNP marker provides a valuable tool for improving Sea Island cotton fiber quality. Full article

Maize (Zea mays L.), a critical global food crop, suffers severe yield losses from drought and salt stresses. Xyloglucan endotransglucosylase/hydrolases (XTHs) are cell wall-modifying enzymes regulating plant growth and abiotic stress responses, but the role of -ZmXTH23 in drought and salt tolerance remains unclear. Here, we heterologously expressed ZmXTH23 in Nicotiana benthamiana (N. benthamiana) and confirmed its extracellular localization. ZmXTH23-overexpressing (OE) lines showed significantly increased plant height, root length, and shoot fresh weight under normal conditions. Under 350 mM NaCl or drought stress, the OE lines exhibited enhanced tolerance, with less leaf wilting, higher biomass, and larger leaf area. Physiologically, the OE lines had higher peroxidase (POD) and superoxide dismutase (SOD) activities and relative water content (RWC), but lower malondialdehyde (MDA) content. Additionally, OE seeds maintained ≥50% germination under 120 mM NaCl (WT: 0%) and nearly 100% under 200 mM mannitol. ZmXTH23 improves drought and salt tolerance in N. benthamiana by upregulating antioxidant enzymes and enhancing water retention, making it a promising candidate for maize stress-resistance breeding. Full article

Ulva prolifera is the main causative species of marine green tides and exhibits extreme tolerance to intertidal abiotic stress. However, the underlying molecular mechanisms remain largely unclear. In this study, we cloned and characterized an MYB-related transcription factor, UpMYB-PHL, from U. prolifera. Expression analysis showed that UpMYB-PHL is rapidly and significantly induced by high-salt stress. Furthermore, heterologous overexpression of UpMYB-PHL in the model microalga Chlamydomonas reinhardtii significantly improved its salt tolerance and biomass. By yeast one-hybrid and dual-luciferase assays, we demonstrated that UpMYB-PHL directly binds to and activates the promoter of the phosphate transporter gene UpPHT1, which is a typical target gene of phosphate response (PHR) transcription factor and participates in salt stress responses of plants. Interestingly, yeast two-hybrid assays revealed that UpMYB-PHL physically interacts with UpGAPDH, a core enzyme in energy metabolism. Taken together, our findings reveal a novel regulatory network in which UpMYB-PHL coordinates phosphorus transporter and energy metabolism in response to salt stress in U. prolifera. This study provides a vital molecular explanation for the rapid adaptation and massive growth of U. prolifera under severe intertidal salt stress. Full article

Glycogen synthase kinase 3 (GSK3/Shaggy-like) belongs to evolutionarily conserved serine/threonine protein kinases that regulate plant morphological development, multi-hormone crosstalk and adaptation to abiotic stresses. However, systematic genome-wide characterization of CmGSK3 is still absent in melon (Cucumis melo L.). This study identified six CmGSK3 members on a whole-genome level, unevenly distributed among four chromosomes. Combined phylogenetic and synteny profiling separated these six genes into four conserved subclades; orthologous links were discovered between melon, Arabidopsis, and rice, revealing evolutionary conservation between monocot and dicot crops. Prediction of promoter cis-regulatory motifs combined with transcriptome datasets suggested that CmGSK3 genes participate in hormone transduction and environmental stress adaptation. Quantitative real-time PCR further verified that exogenous brassinosteroid (BR) application dramatically induced transcriptional accumulation of CmSK21 and CmSK22. Heterologous overexpression of these two genes in wild-type Arabidopsis significantly lowered plant sensitivity to BR, confirming they may function as negative modulators of the BR signaling cascade. Collectively, CmGSK3 members coordinate multiple metabolic routes, dominated by BR-related signal transduction, to manipulate melon growth and stress adaptability. This study establishes the first systematic research on the melon GSK3 family and supplies elite candidate genes for molecular breeding targeting fruit quality and stress resistance improvement in melon. Full article

Drought stress is one of the most prevalent abiotic stressors and severely impairs plant growth and productivity. Therefore, identifying functional genes associated with drought tolerance is essential for the molecular breeding of drought-resistant crops. The RD22 (Responsive to Desiccation 22) gene family encodes conserved BURP domain-containing proteins that participate in plant responses to drought stress. In this study, two RD22 homologs, DsRD22a and DsRD22b, were isolated and characterized from the drought-tolerant ornamental species Dianthus spiculifolius. Sequence analysis showed that both proteins contain a conserved BURP domain and are typical members of the RD22 family. Tissue-specific expression analysis revealed that both genes were predominantly expressed in leaves and stems. Abiotic stress assays demonstrated that the expression levels of DsRD22a and DsRD22b were significantly induced by abscisic acid (ABA), osmotic stress, and salt stress, whereas their transcriptional responses to relatively low-temperature and oxidative stress were relatively weak. Subcellular localization analysis indicated that DsRD22a and DsRD22b proteins are localized in the cytoplasm. Heterologous overexpression assays showed that transgenic Arabidopsis thaliana lines overexpressing DsRD22a or DsRD22b exhibited significantly enhanced tolerance to salt and osmotic stresses compared with wild-type (WT) plants. Soil drought assays further confirmed that the transgenic lines had higher soluble protein contents and improved drought tolerance than WT plants. These findings suggest that DsRD22a and DsRD22b positively regulate plant responses to drought stress, potentially by promoting soluble protein accumulation. Collectively, DsRD22a and DsRD22b represent valuable candidate genes for the genetic improvement of drought tolerance in plants. Full article

1-Deoxy-D-xylulose-5-phosphate synthase (DXS) serves as the initial rate-limiting enzyme in the methylerythritol phosphate (MEP) pathway, governing the biosynthesis of precursors for photosynthetic pigments and terpenoids. In this study, the LaDXS2-2 gene was cloned and functionally characterized in lavender (Lavandula angustifolia). The full-length coding sequence (CDS) of LaDXS2-2 spans 2178 base pairs, encoding a protein of 725 amino acids. Phylogenetic analysis revealed that LaDXS2-2 is most closely related to the DXS from Salvia miltiorrhiza. Expression profiling demonstrated that LaDXS2-2 was highly expressed in flower buds, and its transcript levels were significantly upregulated (p < 0.05) in response to ethephon, high light intensity, and low temperature, while exhibiting tissue-specific responses to gibberellin application. Subcellular localization assays confirmed LaDXS2-2 is targeted to the chloroplast. Heterologous overexpression of LaDXS2-2 in tobacco resulted in a marked increase in photosynthetic pigment content, enhanced the actual photochemical efficiency of photosystem II [Y(II)], and reduced non-photochemical quenching (NPQ). Integrated transcriptomic and metabolomic analyses further revealed that LaDXS2-2 overexpression activated the diterpenoid biosynthesis pathway and upregulated amino acid metabolism as well as the TCA cycle, while competitively suppressing phenylpropanoid and flavonoid biosynthesis pathways. These findings indicate that LaDXS2-2 not only enhances photosynthetic efficiency by promoting the synthesis of photosynthetic pigments but also suggests a potential role in influencing primary carbon and nitrogen metabolism, as inferred from transcriptomic and metabolomic data. This functionality may ultimately influence plant growth and metabolic homeostasis. Overall, this study provides a theoretical foundation for the synergistic improvement of photosynthetic efficiency and secondary metabolism in crops. Full article