番茄斑萎病毒基因组RNA m6A修饰调控病毒侵染的分子机制研究

番茄斑萎病毒（TSWV）是一种在农业生产上危害极为严重的植物负链RNA病毒。前期研究发现，TSWV基因组RNA存在高丰度的N6-甲基腺嘌呤(m6A)修饰，沉默m6A去甲基酶NbALKBH9可促进TSWV的侵染，这与已报道的敲低m6A去甲基酶抑制植物正链RNA病毒侵染的作用机制相反，暗示m6A修饰对植物负链RNA病毒的复制和侵染存在全新的分子调控机制。m6A作为一种新的病毒调控因子，对植物负链RNA病毒的表观遗传修饰作用及其调控机制尚待深入研究。本项目拟继续鉴定TSWV基因组的m6A修饰位点，探究突变m6A位点对TSWV侵染的影响，阐明m6A甲基转移酶和去甲基酶对TSWV侵染的调控作用，揭示病毒基因组m6A修饰在TSWV侵染时抵抗寄主植物的反防御机制，从而全面解析病毒基因组m6A甲基化的表观遗传调控机制。本项目的开展对揭示TSWV和寄主植物之间防御和反防御机理具有重要意义。

Tomato spotted wilt virus (TSWV) is one of the most serious plant segmented negative-stranded RNA (NSR) viruses in the world. Our preliminary studies found that the TSWV genomic RNAs has a high level of N6-methyladenosine (m6A) methylation. Moreover, the silencing of m6A demethylase NbALKBH9 promoted the TSWV infection in Nicotiana benthamiana, which is contrary to the report that knockdown of demethylase AtALKBH9B inhibited the infection of plant positive-stranded RNA viruses in Arabidopsis thaliana, suggesting that m6A modification utilizes a novel molecular regulation mechanism for the replication and infection in plant NSR viruses. As a new regulatory factor of viral infection, it remains unclear how m6A methylation regulating the replication and infection of plant NSR viruses. In this project, we intend to identify the m6A methylation sites of TSWV genomic RNA segments, and to investigate the role of m6A methylation in TSWV infection. Furthermore, we will explore the regulatory mechanism of m6A demethylases and methyltransferases in TSWV infection based on reverse genetics system to reveal the epigenetic regulation mechanism of viral genomic m6A methylation, and eventually illustrate the counter-defense mechanism of viral genomic m6A modification against host plants upon TSWV infection. The implement of this project will provide a great significance in revealing the defense and counter-defense mechanisms between TSWV and host plants.