| 摘要: |
| 为探究芥菜MYB28家族成员在生物胁迫防御中的功能，该研究利用从叶用芥菜中克隆获得的6个BjuMYB28s同源基因，明确了其亚细胞定位和转录激活活性，并重点研究了BjuBMYB28-2的表达模式及其对甜菜夜蛾和核盘菌的响应特征。结果表明：（1）BjuAMYB28-1、BjuBMYB28-1和BjuBMYB28-3与前期研究的BjuAMYB28-2和BjuAMYB28-3一致，均定位于细胞核和细胞质，而BjuBMYB28-2仅定位于细胞核，6个BjuMYB28s均具有转录激活活性。（2）BjuBMYB28-2主要在维管组织中特异性表达，甜菜夜蛾取食和核盘菌侵染后12 ~ 24 h叶脉中GUS信号减弱，RT-qPCR进一步验证了核盘菌侵染后其整体表达水平下降。（3）异源表达BjuBMYB28-2可使拟南芥对甜菜夜蛾与核盘菌的抗性显著提高，表现为取食幼虫体重降低，核盘菌侵染病斑缩小。（4）转录组分析显示，过表达株系中硫苷生物合成3个阶段的多个关键基因均上调表达。综上，BjuBMYB28-2可能通过协同上调硫苷生物合成通路关键基因的表达进而提高硫苷含量，从而增强对甜菜夜蛾和核盘菌的抗性。该研究为理解芥菜MYB28家族成员的功能分化及其在抗虫抗病中的作用提供了理论依据。 |
| 关键词: 芥菜，BjuBMYB28-2，硫苷，甜菜夜蛾，核盘菌，生物胁迫 |
| DOI：10.11931/guihaia.gxzw202603046 |
| 分类号: |
| 基金项目:：国家自然科学基金面上项目(32072557)，浙江省自然科学基金项目（LY24C150003）。 |
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| Expression of BjuBMYB28-2 in mustard (Brassica juncea) and its functional analysis in pest and disease resistance |
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WANG Shanyi, WANG Jingjing, ZHANG Lili, YU Youjian, ZHU Zhujun, XU Liai*
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College of Horticulture, Zhejiang A&F University, Hangzhou 311300, China
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| Abstract: |
| To investigate the functional roles of the MYB28 family members in biotic stress defense in mustard (Brassica juncea), this study utilized six BjuMYB28s homologous genes cloned from leaf mustard. Their subcellular localization and transcriptional activation activities were systematically characterized, with particular emphasis on the expression patterns of BjuBMYB28-2 and its responsiveness to Spodoptera exigua (beet armyworm) herbivory and Sclerotinia sclerotiorum infection. Our experimental results revealed several key findings. (1) Regarding subcellular distribution, BjuAMYB28-1, BjuBMYB28-1, and BjuBMYB28-3 were localized in both the nucleus and the cytoplasm, consistent with previously reported patterns for BjuAMYB28-2 and BjuAMYB28-3. In contrast, BjuBMYB28-2 exhibited an exclusive nuclear localization, suggesting potential functional specialization among these homologs. Notably, all six BjuMYB28 proteins demonstrated transcriptional activation activity in yeast assays, indicating their capacity to function as transcriptional regulators. (2) Tissue-specific expression analysis showed that BjuBMYB28-2 was predominantly expressed in vascular tissues. Interestingly, GUS staining intensity in leaf veins markedly decreased at 12-24 h post-treatment with either insect feeding or fungal inoculation, implying a rapid down-regulation of promoter activity under stress conditions. This observation was further corroborated by RT-qPCR analysis, which confirmed a significant reduction in overall transcript abundance of BjuBMYB28-2 following S. sclerotiorum infection. (3) Heterologous overexpression of BjuBMYB28-2 in Arabidopsis thaliana conferred significantly enhanced resistance to both insect pests and fungal pathogens. Transgenic lines exhibited reduced larval body weights of S. exigua fed on overexpression plants, alongside diminished lesion areas resulting from S. sclerotiorum inoculation, compared to wild-type controls. (4) Transcriptomic profiling of overexpression lines revealed coordinated up-regulation of multiple key genes involved in all three stages of glucosinolate biosynthesis, including core structural and regulatory genes. Collectively, these findings strongly suggest that BjuBMYB28-2 reinforces plant defense by synergistically up-regulating the glucosinolate biosynthetic pathway, leading to increased glucosinolate accumulation, which in turn contributes to enhanced resistance against both chewing insects and necrotrophic fungi. This comprehensive investigation not only provides mechanistic insights into how BjuBMYB28-2 modulates biotic stress tolerance but also establishes a theoretical foundation for understanding the functional divergence among MYB28 family members in B. juncea, with potential implications for breeding programs aimed at improving pest and disease resistance in Brassica crops. |
| Key words: Brassica juncea, BjuBMYB28-2, glucosinolate, Spodoptera exigua, Sclerotinia sclerotiorum, biotic stress |
