番茄Tm-22蛋白是一种CC-NBS-LRR（卷曲螺旋结构域-含核苷酸结合结构域-富亮氨酸重复结构域）类抗性（Resistance，R）蛋白，它能识别烟草花叶病毒（Tobacco mosaic virus，TMV）的运动蛋白（Movement protein，MP）并激活植物产生强烈的免疫反应。植物需要迅速、精确地对参与抗病反应的基因进行调控，该过程通常涉及表观遗传调控。真核生物中，组蛋白乙酰化修饰作为一种表观遗传调控的方式，对染色质结构修饰和基因表达调控至关重要，但该类修饰在R蛋白介导的抗病毒反应中的作用及其机制尚不清楚。本研究发现，本生烟中组蛋白乙酰化酶NbHAT1和NbHAT2是Tm-22介导的抗TMV免疫反应的正调控因子。并且，NbHAT2能够与Trihelix转录因子家族的NbGT1（Trihelix transcription factor-1）直接互作，协同参与Tm-22介导的抗病毒反应。同时，二者响应TMV的侵染，表现为TMV侵染Tm-22转基因植物后，NbHAT2和NbGT1基因的转录水平上调以及二者编码的蛋白之间的互作增强。进一步研究发现，沉默NbHAT2或NbGT1基因导致植物组蛋白H4（Histone 4）整体的乙酰化水平降低；过表达NbHAT2或NbGT1的转基因植物中，组蛋白H4整体的的乙酰化水平升高，尤其对组蛋白H4第16位赖氨酸（H4K16）的乙酰化水平影响显著。本研究还发现高迁移率族蛋白NbHMGB3（High mobility group B3）同样也正向调控Tm-22介导的抗TMV过程。NbGT1可结合NbHMGB3基因的启动子区，它可能招募NbHAT2调节该基因启动子区域的组蛋白H4的乙酰化水平来共同调节NbHMGB3基因的转录水平，从而影响Tm-22转基因植物对TMV的抗性。综上所述，本研究首次报道了NbHAT2、NbGT1和NbHMGB3是Tm-22抗TMV信号通路的重要调控蛋白，揭示了组蛋白乙酰化酶在Tm-22介导的抗病毒反应中的分子机制，探索了乙酰化修饰调控在植物R基因介导的抗病毒机制中的作用，为作物抗病毒提供新思路。
Tomato Tm-22 is a representative example of resistance （R） proteins and confers resistance against the Tobacco mosaic virus （TMV）. Histone acetylation is essential for chromatin structure modification and gene expression regulation in eukaryotes. It participates in rapid and massive reprogramming of transcription to precisely regulate and fine-turn defense responses after pathogen infection. However, the underlying mechanism of histone acetylation in the R protein-mediated antiviral response is poorly understood. This study showed that the histone acetylase NbHAT2 directly interacted with NbGT1, a member of Trihelix transcription factor-1, both of which are positive regulators of Tm-22-mediated resistance against TMV. Once TMV infected transgenic Tm-22 plants, the transcription levels of the NbHAT2 and NbGT1 gene were up-regulated, and the interaction between NbHAT2 and NbGT1 was enhanced. It was found that NbHAT2 and NbGT1 modulated the overall acetylation level of histone H4, especially in the H4K16. The high mobility group protein NbHMGB3 also positively regulated Tm-22-mediated resistance against TMV, which was shown to be co-regulated by NbHAT2 and NbGT1. NbGT1 specifically bound to the GT element in the promoter region of the NbHMGB3 gene and probably recruited NbHAT2 to modify histone H4 acetylation in the promoter region of the gene, which activated the transcription of the NbHMGB3 gene to enhance TMV resistance. In conclusion, an uncharacterized epigenetic complex was verified to function in Tm-22-mediated TMV resistance, which provides mechanistic insights into how a histone acetylase along with a transcription factor contributes to R protein-mediated immune responses.
