Analysis of chromium (Cr)-responses genes and miRNAs in brassica juncea (L.)

Brassica juncea, commonly known as Indian or brown mustard, is a significant industrial oil crop worldwide. It is commercially valuable because it provides raw materials for various industries, including animal feed, biodiesel, cooking oil, and medicine. B. juncea demonstrates substantial potential for the phytoremediation of chromium (Cr)-contaminated soils owing to its high biomass, rapid growth, moderate metal accumulation, ease of harvesting, and metal tolerance. Recently, microRNAs are recognized for their crucial regulatory roles in plant responses to different metal stresses. Nevertheless, there is limited information on microRNAs in B. juncea, particularly those involved to chromium (Cr) stress response. In this study, the transcriptomic response of B. juncea Early raya, which is tolerant to chromium toxicity, and B. juncea Sindh raya, which is sensitive to chromium toxicity, was investigated using RNA sequencing. The expression of specific genes be regulated by chromium toxicity tolerance was further examined through RT-qPCR. In the physiological growth analysis, Early raya showed less impact from high chromium (1000 µM) on all growth parameters, including root-shoot length, shoot fresh weight, root fresh weight, and root-shoot dry weight, compared to other B. juncea varieties. Conversely, Sindh raya exhibited more significant effects from high chromium (1000 µM) on the same growth parameters. Additionally, after chromium application, the highest chromium uptake was observed in the roots, with the root chromium content being significantly higher than in plant's above-ground parts. Early raya accumulated high chromium in the roots when treated with Cr³⁺ (1000 µM) for two weeks (15 days) and showed high chromium tolerance compared to the other Brassica juncea varieties. A total of 70 genes were differentially expressed in the roots of Early raya when treated with 1000 µM Cr, compared to the control plants (without Cr). Of these genes, the expression of 27 were statistically significant (p<0.05). Among the significantly different genes, 19 were upregulated and 8 were downregulated. On the other hand, the roots of Sindh raya when treated with 1000 µM Cr for two weeks showed low chromium tolerance; 80 genes in variety were differentially expressed compared to the control. Of these genes, the expression of 50 were statistically significant (p<0.05). Among them, 28 were upregulated, and 22 were downregulated. For RT-qPCR analysis, three genes involved in different mechanisms and showing significant differential expression were selected according to transcriptome sequencing results. Among the genes studied in the Cr-treated experimental groups compared to the control, the APX, Fe-SOD, and RBOH genes were upregulated. This thesis identifies potentially significant miRNAs, genes, and pathways related to Cr³⁺ stress response through miRNA and mRNA profiling in B. juncea. The identified pathways, genes, and miRNAs are significant targets for future omics studies to enhance chromium tolerance in B. juncea. This represents the first comprehensive study on chromium stress tolerance and transcriptomic changes in B. juncea. This study supported by the Council of Higher Education's (YOK) 100/2000 (Plant Genetics and Agricultural Biotechnology Program) Doctoral Scholarship Program and the Scientific and Technological Research Council of Turkey (TUBITAK) (2211-A Ph.D. Program). Additionally, the support from BAP Project given to Prof. Dr. Abdülrezzak Memon (project no: 2024/MF002) is appreciated.