SlMAPKs在番茄防御TYLCV中的作用机理

Tomato Yellow Leaf Curl Virus (TYLCV) is one of the most serious pathogens harming tomato, and its variety has broken through the existing resistance of cultivated tomato. This indicates that the response mechanism of tomato’s defense to TYLCV needs to be further studied in order to enrich the theoretical basis of tomato breeding with TYLCV resistance. Through transcriptome data analysis, the applicant finds that SlMAPKs participate in the response of tomato against TYLCV. Further detection of the expression of SlMAPKs confirm three genes with significant differences, but the mechanism is not clear. In this project, susceptible and Ty-2-containing tomato plants which is the second NB-LRR type disease-resistant gene in plant response to geminivirus were used as experimental materials. VIGS technology will be used to quickly identify candidate SlMAPKs. Overexpression and gene knockout will be used to validate the resistance function of identified candidate SlMAPKs to TYLCV. Finally, yeast hybrid and BiFC technology will be used to analyze the mechanism of SlMAPKs participating in the defense of TYLCV. The purpose of this project is to analyze the response of SlMAPKs to TYLCV resistance in tomato and elucidate its regulation mechanism, so as to provide theoretical reference for breeding of tomato resistant to TYLCV.

番茄黄化曲叶病毒（TYLCV）是危害番茄最严重的病原之一，其变种已突破栽培番茄的现有抗性。这表明亟需对番茄防御TYLCV的响应机制做深入研究，以丰富番茄抗TYLCV育种的理论基础。申请人通过对转录组数据分析发现SlMAPKs参与番茄防御TYLCV的应答响应，进一步对SlMAPKs表达检测后确认3个差异显著基因，但其作用机制尚不清楚。本项目以感病和含Ty-2这一植物应对双生病毒响应中的第二个NB-LRR类型抗病基因的番茄植株为实验材料，利用VIGS技术快速确认候选SlMAPKs，采用过表达和基因敲除对已确认候选SlMAPKs对TYLCV抗性的功能进行验证，最后利用酵母双杂、BiFC等技术剖析SlMAPKs参与防御TYLCV的作用机理。本项目旨在分析SlMAPKs在番茄对TYLCV抗性应答中的响应规律，并阐明其作用机理，为番茄抗TYLCV育种工作提供理论参考。

番茄黄化曲叶病毒病是世界性的番茄生产限制性因素。因TYLCV较易发生变异，生产中长期使用含有特定抗病基因的抗性品种会加速病毒的变异，并突破抗病基因的防御。因此，我们对番茄抗TYLCV机理开展了进一步探讨。在本研究中我们主要明确了在TYLCV侵染初期病毒会通过影响ROS的平衡状态、植物激素的合成与信号转导等的变化进而影响植株抗性和生长状态。植物激素在番茄抗TYLCV过程中扮演重要角色，尤以水杨酸（SA）在抗性防御中作用明显；在感病材料Moneymaker和抗性材料CLN2777A的RNA-seq数据比较分析中发现20个共同调控的DEGs，这些基因可能是番茄对TYLCV侵染响应的枢纽。同时通过ATAC-seq发现共有的关键转录因子对应的显著增强结构域主要有BZIP60, TGA6, SEP3，WRKY59。后续仍需深入挖掘TYLCV侵染后番茄植株体内上游差开放染色质区域差异性与下游差异基因表达之间的调控关联，为全面解析番茄对TYLCV防御机制提供新思路；TYLCV侵染后抗感材料中MAPK信号通路相关基因均呈现多样化表达模式，这可能是两种材料平衡和适应病毒侵染能力的部分原因。我们通过不同方式进一步剖析了番茄防御TYLCV的信号转导通路及其作用机理，为今后调控番茄对TYLCV抗性提供理论指导，进而培育抗TYLCV的番茄新品种，具有十分重要的科学与现实意义。