WRKY transcription factors (TFs) participate in plant defense mechanisms against biological and abiotic stresses. However, their regulatory role in heat resistance is still unclear in non-heading Chinese cabbage. Here, we identified the WRKY-IIe gene BcWRKY22(BraC09g001080.1), which is activated under high temperatures and plays an active role in regulating thermal stability, through transcriptome analysis. We further discovered that the BcWRKY22 protein is located in the nucleus and demonstrates transactivation activity in both the yeast and plant. Additionally, our studies showed that the transient overexpression of BcWRKY22 in non-heading Chinese cabbage activates the expression of catalase 2 (BcCAT2), enhances CAT enzyme activity, and reduces Hydrogen Peroxide (H₂O₂) accumulation under heat stress conditions. In addition, compared to its wild-type (WT) counterparts, Arabidopsis thaliana heterologously overexpresses BcWRKY22, improving thermotolerance. When the BcWRKY22 transgenic root was obtained, under heat stress, the accumulation of H₂O₂ was reduced, while the expression of catalase 2 (BcCAT2) was upregulated, thereby enhancing CAT enzyme activity. Further analysis revealed that BcWRKY22 directly activates the expression of BcCAT2 (BraC08g016240.1) by binding to the W-box element distributed within the promoter region of BcCAT2. Collectively, our findings suggest that BcWRKY22 may serve as a novel regulator of the heat stress response in non-heading Chinese cabbage, actively contributing to the establishment of thermal tolerance by upregulating catalase (CAT) activity and downregulating H₂O₂ accumulation via BcCAT2 expression.

As one of the most important vegetables globally, peppers have garnered significant attention from breeders due to their diverse agronomic traits, including plant type, leaf shape, and maturity. Understanding the genetic mechanisms underlying these traits is crucial for systematic advancements in sweet pepper breeding. In this study, leveraging the PepperSNP50K liquid breeding chip, we conducted a comprehensive analysis of horticultural traits and genetic diversity using sweet pepper germplasm samples. Initially, the sweet pepper populations were analyzed using SNP-based liquid chip technology. Subsequently, phenotypic surveys were performed on 217 sweet pepper samples, and the collected phenotypic data were integrated with SNP markers to conduct a genome-wide association study (GWAS) of key agronomic traits. Among the 25 horticultural traits evaluated, 11 exhibited significant associations with 54 SNP polymerization regions and 193 candidate genes. These findings provide a robust foundation for the utilization of sweet pepper germplasm resources and the development of new, improved varieties. Furthermore, in this study, we identified Caz06g05770 (Lycopene beta-cyclase) as a candidate gene responsible for the color of mature ripe fruits. This research not only enhances our understanding of the genetic basis of sweet pepper traits but also offers a practical roadmap for advancing breeding programs and boosting agricultural productivity. By bridging the gap between genetic research and practical breeding applications, this study paves the way for the development of high-yield, high-quality sweet pepper varieties tailored to meet the growing demands of global agriculture.

Leaves are the primary organs for plant photosynthesis, and their flat, symmetric morphology is crucial for plant growth and development. The LBD family transcription factor ASYMMETRIC LEAVES 2 (AS2) plays a central role in the establishment of leaf polarity. In this study, we cloned the BcAS2 gene from the non-heading Chinese cabbage cultivar “NHCC001” and successfully generated overexpression strains through genetic transformation. Phenotypic analysis revealed that overexpression of BcAS2 led to significant upward curling of leaves in non-heading Chinese cabbage. Additionally, we found that the expression of BcPHB, a gene associated with leaf adaxial polarity development, was significantly up-regulated in BcAS2-overexpressing plants compared to controls. This interaction was further confirmed through yeast one-hybridization (Y1H), dual-luciferase reporter assays, and electrophoretic mobility shift assay (EMSA), all of which demonstrated that BcAS2 directly binds to the GATA-motif site of the BcPHB promoter and promotes its transcription. Functional validation via overexpression and silencing of BcPHB confirmed its role in regulating adaxial polarity development in non-heading Chinese cabbage leaves. This study elucidates the molecular mechanism of the BcAS2-BcPHB pathway in regulating leaf polarity in non-heading Chinese cabbage, providing a theoretical foundation for morphological improvement breeding.

Chloroplast-genome variation below the species level remains insufficiently characterized in Brassica, because most previous studies have relied on only one or a few representative plastomes. To determine whether dense within-subspecies sampling could reveal localized variation obscured by broader comparisons, we sequenced and assembled complete chloroplast genomes from 10 morphologically diverse varieties of Non-heading Chinese cabbage (NHCC; Brassica campestris [syn. Brassica rapa] ssp. chinensis) and placed their codon-usage patterns in the context of 15 Brassica plastomes. The NHCC plastomes were highly conserved in genome size (153,480–153,533 bp), GC content (36.36–36.39%), gene content, gene order, and quadripartite structure. Nevertheless, locus-level comparisons showed that the limited sequence variation was concentrated in a small number of regions. Six protein-coding loci and four intergenic spacers exhibited relatively elevated nucleotide diversity, and explicit counts of variable sites and internal indel blocks further characterized the sequence differences underlying these values. These loci represent candidate regions for population-level validation and potential molecular-marker development rather than confirmed variety-specific markers. In the broader comparison, mean ENC varied only from 45.15 to 45.49 among the 15 plastomes, indicating a highly conserved codon-usage background characterized by a general preference for A/U-ending synonymous codons. Eight codons—ACU, AGA, CAA, CUA, GCU, GGU, GUA, and UGU—were consistently recovered as putative optimal codons, although statistical support for ACU and GGU was restricted to a subset of accessions. PR2 analysis revealed imbalances within complementary nucleotide pairs rather than an overall preference for G/C-ending codons. Thus, the principal contribution of this study is the fine-scale localization of plastome variation through dense sampling within NHCC, providing candidate regions for subsequent diversity assessment and marker validation in independently sampled individuals.

Background/Objectives: Fruit quality in melon is determined by complex traits including sugar accumulation, aroma, flesh color, ripening behavior and disease resistance. Conventional genomic selection provides limited mechanistic interpretation, especially for regulatory and de novo variants. We adapted the EVEE interpretable embedding–probing framework to melon using PlantCAD2 as a frozen DNA language model backbone. Methods: The model was not fine-tuned or further pre-trained. Instead, lightweight probes were trained on fixed embeddings derived from sequence windows surrounding genetic variants. The pipeline integrated supervised variant-effect prediction, functional perturbation profiling, training-free masked log-likelihood ratio scoring, and cis-regulatory element generation. We applied the framework to 32,268 public genotyping-by-sequencing (GBS) SNPs from a melon diversity panel. A total of 30,052 variants were successfully encoded and analyzed. Results: Of these, 31.9% were classified as high predicted functional effect under the GWAS-proximal probe. We adopted a rigorous independent split based on chromosome and locus blocks to ensure test variants and training variants occupy distinct linkage-disequilibrium blocks. In this setting, the covariance probe yields an AUROC of approximately 0.51 with a 95% confidence interval covering 0.50, indicating no predictive advantage over random guessing. The full-cohort AUROC value of 0.748 reported earlier is attributed to linkage-disequilibrium leakage and overlapping positive samples between datasets, and thus does not reflect true independent generalization. The 17-class structural annotation probe reached 95.97% per-position accuracy. Functional perturbation signals were enriched in intronic, coding, and proximal promoter regions, consistent with known regulatory architectures in melon. Using the masked language model head with Gibbs sampling, we generated 76 candidate 400 bp cis-regulatory elements associated with sugar metabolism pathways. A posterior consistency check against 18 published GWAS/QTL loci showed effect-score patterns broadly concordant with reported loci; this is a consistency analysis against known loci, not an independent prospective validation. Variants in the CmTST2 region exhibited elevated mean effect scores of 0.7922 compared with the genome-wide background of 0.406. Conclusions: Overall, this study provides a computational, hypothesis-generating framework in which a frozen plant DNA language model combined with lightweight probes prioritizes melon variants and generates candidate regulatory sequences; all outputs are unverified in-silico predictions requiring experimental validation before any functional or breeding use. Because every variant is scored independently from its local sequence context, the framework necessarily treats genetically linked, network-dependent traits as single-locus effects and does not model epistasis, pathway-level interactions, or genotype-by-environment effects; the outputs are sequence-level priors, not phenotype predictions. The results support a generalizable framework for model-guided functional genomics and precision breeding in Cucurbitaceae crops.