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Background: Cardamine violifolia is a significant Brassicaceae plant known for its high selenium (Se) accumulation capacity, serving as an essential source of Se for both humans and animals. WRKY transcription factors play crucial roles in plant responses to various biotic and abiotic stresses, including cadmium stress, iron deficiency, and Se tolerance. However, the molecular mechanism of CvWRKY in Se accumulation is not completely clear., Results: In this study, 120 WRKYs with conserved domains were identified from C. violifolia and classified into three groups based on phylogenetic relationships, with Group II further subdivided into five subgroups. Gene structure analysis revealed WRKY variations and mutations within the CvWRKYs. Segmental duplication events were identified as the primary driving force behind the expansion of the CvWRKY family, with numerous stress-responsive cis-acting elements found in the promoters of CvWRKYs. Transcriptome analysis of plants treated with exogenous Se and determination of Se levels revealed a strong positive correlation between the expression levels of CvWRKY034 and the Se content. Moreover, CvWRKY021 and CvWRKY099 exhibited high homology with AtWRKY47, a gene involved in regulating Se accumulation in Arabidopsis thaliana. The WRKY domains of CvWRKY021 and AtWRKY47 were highly conserved, and transcriptome data analysis revealed that CvWRKY021 responded to Na 2 SeO 4 induction, showing a positive correlation with the concentration of Na 2 SeO 4 treatment. Under the induction of Na 2 SeO 3 , CvWRKY021 and CvWRKY034 were significantly upregulated in the roots but downregulated in the shoots, and the Se content in the roots increased significantly and was mainly concentrated in the roots. CvWRKY021 and CvWRKY034 may be involved in the accumulation of Se in roots., Conclusions: The results of this study elucidate the evolution of CvWRKYs in the C. violifolia genome and provide valuable resources for further understanding the functional characteristics of WRKYs related to Se hyperaccumulation in C. violifolia., (© 2024. The Author(s).)

Background: Cardamine violifolia, native to China, is one of the selenium (Se) hyperaccumulators. The mechanism of Se metabolism and tolerance remains unclear, and only limited genetic information is currently available. Therefore, we combined a PacBio single-molecule real-time (SMRT) transcriptome library and the Illumina RNA-seq data of sodium selenate (Na 2 SeO 4 )-treated C. violifolia to further reveal the molecular mechanism of Se metabolism., Results: The concentrations of the total, inorganic, and organic Se in C. violifolia seedlings significantly increased as the Na 2 SeO 4 treatment concentration increased. From SMRT full-length transcriptome of C. violifolia, we obtained 26,745 annotated nonredundant transcripts, 14,269 simple sequence repeats, 283 alternative splices, and 3407 transcription factors. Fifty-one genes from 134 transcripts were identified to be involved in Se metabolism, including transporter, assimilatory enzyme, and several specific genes. Analysis of Illumina RNA-Seq data showed that a total of 948 differentially expressed genes (DEGs) were filtered from the four groups with Na 2 SeO 4 treatment, among which 11 DEGs were related to Se metabolism. The enrichment analysis of KEGG pathways of all the DEGs showed that they were significantly enriched in five pathways, such as hormone signal transduction and plant-pathogen interaction pathways. Four genes related to Se metabolism, adenosine triphosphate sulfurase 1, adenosine 5'-phosphosulfate reductase 3, cysteine (Cys) desulfurase 1, and serine acetyltransferase 2, were regulated by lncRNAs. Twenty potential hub genes (e.g., sulfate transporter 1;1, Cys synthase, methionine gamma-lyase, and Se-binding protein 1) were screened and identified to play important roles in Se accumulation and tolerance in C. violifolia as concluded by weighted gene correlation network analysis. Based on combinative analysis of expression profiling and annotation of genes as well as Se speciation and concentration in C. violifolia under the treatments with different Na 2 SeO 4 concentrations, a putative Se metabolism and assimilation pathway in C. violifolia was proposed., Conclusion: Our data provide abundant information on putative gene transcriptions and pathway involved in Se metabolism of C. violifolia. The findings present a genetic resource and provide novel insights into the mechanism of Se hyperaccumulation in C. violifolia.