Liu Tiantian, Zhou Zehua, Luo Changwei, Luo Hua, Tang Jun, Shi Xiaojiang, Li Diping, Zhang Qiong, Li Jin, Xia Yonggang, Song Na, Yi Tuyong
Hunan Provincial Key Laboratory of Plant Diseases and Pests, College of Plant Protection, Hunan Agricultural University, Changsha, Hunan, China.
Shaoyang Academy of Agricultural Sciences, Shaoyang, Hunan, China.
Front Microbiol. 2024 Aug 29;15:1425441. doi: 10.3389/fmicb.2024.1425441. eCollection 2024.
Reactive oxygen species (ROS) generation is a common disease defense mechanism in plants. However, it is unclear whether host activates defense response against causing citrus melanose disease by producing ROS, and the underlying molecular mechanisms are unknown.
DAB staining and RNA-Seq technology were used to compare the active oxygen burst and differential gene expression, respectively, in uninfected and infected leaves at different time points during infection . The functions of (a significant DEG) were confirmed in through the -mediated transient expression system.
DAB staining indicated that initiated defense against infection within 24 h by generating ROS. Illumina sequencing revealed 25,557 expressed genes of . The most upregulated DEGs ( = 1,570) were identified 72 h after fungal inoculation (sample denoted as CD72). In the CD72 vs. Cs (samples at 0 h after fungal inoculation) comparison, the KEGG pathway category with the highest number of genes ( = 62) and most significant enrichment was Protein processing in endoplasmic reticulum, followed by Glutathione metabolism and MAPK signaling pathway-plant. GO analysis revealed that the DEGs of CD72 vs. Cs related to active oxygen burst and chitin recognition were significantly grouped into the regulation of biological processes and molecular functions, with GO terms including response to ROS, response to fungus, and oxidoreductase activity. Remarkably, was significantly enriched in the GO and KEGG analyses, and its expression pattern in qRT-PCR and DAB staining results were consistent. Among the 63 ROS-related DEGs, HSP genes and genes associated with the peroxidase family were highly significant as revealed by protein-protein interaction networks. Furthermore, ROS accumulation, cell death, and upregulation of defense-related genes were observed in leaves with expressed through the -mediated transient expression system.
Our findings suggested that activates and ROS-related genes, leading to ROS accumulation to resist the invasion by . This study laid the foundation for future research on molecular mechanisms and breeding of cultivars resistant to citrus melanose.
活性氧（ROS）的产生是植物中常见的病害防御机制。然而，尚不清楚寄主是否通过产生活性氧来激活针对引起柑橘黑星病的病原菌的防御反应，其潜在的分子机制也尚不明确。
分别采用DAB染色和RNA测序技术，比较感染过程中不同时间点未感染和感染叶片中的活性氧爆发和差异基因表达。通过农杆菌介导的瞬时表达系统在柑橘中证实了一个显著差异表达基因（DEG）的功能。
DAB染色表明，柑橘在24小时内通过产生活性氧启动了对病原菌感染的防御。Illumina测序揭示了柑橘的25557个表达基因。在真菌接种后72小时鉴定出上调最显著的差异表达基因（n = 1570）（样本记为CD72）。在CD72与Cs（真菌接种后0小时的样本）的比较中，基因数量最多（n = 62）且富集最显著的KEGG通路类别是内质网中的蛋白质加工，其次是谷胱甘肽代谢和植物中的MAPK信号通路。GO分析表明，CD72与Cs中与活性氧爆发和几丁质识别相关的差异表达基因显著聚集在生物过程和分子功能的调控中，GO术语包括对活性氧的反应、对真菌的反应和氧化还原酶活性。值得注意的是，该基因在GO和KEGG分析中显著富集，其在qRT-PCR中的表达模式与DAB染色结果一致。蛋白质-蛋白质相互作用网络显示，在63个与活性氧相关的差异表达基因中，热休克蛋白（HSP）基因和与过氧化物酶家族相关的基因高度显著。此外，在通过农杆菌介导的瞬时表达系统表达该基因的柑橘叶片中观察到活性氧积累、细胞死亡和防御相关基因的上调。
我们的研究结果表明，柑橘激活了病程相关基因和与活性氧相关的基因，导致活性氧积累以抵御病原菌的入侵。本研究为今后柑橘抗黑星病品种的分子机制研究和育种奠定了基础。
