转录因子NTO1对尖镰孢寄生致病作用调控机理研究

Fusarium oxysporum is a world-wide soil-borne fungus that induces wilt and root rot on a variety of plants. Regulation mechanisms of Fusarium oxysporum on parasitism and pathogenicity in vascular tissue is the key to screen the molecular targets on disease prevention and control. Transcription factors play important roles on growth, development and virulence of pathogen, which come to be the new molecular targets on disease prevention and control. We have got the NTO1 deletion mutants and complementation mutants by gene knockout and complementation based on the genome of F. oxysporum f. sp. conglutinans in this research. Transcription factor NTO1 is one of the subunits of histone acetylation transferase NuA3 complex. The differences on conidia development, oxygen stress resistance, secondary metabolites and virulence are significant between wild type isolates and deletion mutants. The differentially expressed genes have been identified by RNA-Seq analysis. Furthermore, the binding site of NTO1 on the genome of F.oxysporum f. sp. conglutinans, the genes and pathways which are regulated by transcription factors NTO1 will be analyzed by ChIP-Seq and RNA-Seq. Based on the results, the predicted key genes will be knockout and their biological functions will be confirmed. The results contribute to clarify the regulation mechanisms of transcription factor NTO1 on parasitism and pathogenicity in Fusarium oxysporum. It is significant to analysis the molecular mechanisms on growth, development and virulence of F. oxysporum from transcription factors. This will enrich the pathogenic mechanisms of F. oxysporum and provide the new targets and strategies for the controls of cabbage Fusarium wilt.

尖镰孢菌是世界性的土传病原真菌，给多种经济作物和农作物造成重大经济损失。解析尖镰孢菌的寄生致病机理是筛选其防控分子靶标的关键。转录因子在病原菌生长发育和致病中发挥重要作用，成为新的防控分子靶标。前期获得NuA3组蛋白乙酰化转移酶复合体中转录因子NTO1敲除及回补菌株，发现缺失突变菌株在孢子产量、抗氧胁迫能力、次级代谢产物和致病力上与野生菌株有显著差异。对RNA-Seq数据进行分析，获得受NTO1调控的显著差异表达基因。本项目拟利用RNA-Seq和ChIP-Seq分析NTO1在尖镰孢基因组DNA上的结合位置，预测其调控的基因和代谢途径；对关键基因通过遗传操作进行功能分析，确认其所调控的下游基因和参与的代谢途径；阐明NTO1对尖镰孢菌寄生致病作用的调控机理。本项目为甘蓝枯萎病的防控提供了新靶标和新策略。

尖镰孢菌是世界性的土传病原真菌，引起维管束病害，造成植物枯萎病，可侵染的寄主多达300余种，给多种经济作物和农作物造成重大经济损失。转录因子在病原物的生长发育和致病中发挥重要作用，其功能的缺失可能会引起生物体一系列性状的改变。申请人前期研究结果显示，转录因子NTO1的缺失引起小型分生孢子产量降低，抗氧胁迫能力下降，次级代谢产物发生改变，并且对寄主甘蓝的致病力降低，表明转录因子NTO1在调控尖镰孢生长发育、抗氧胁迫能力、次级代谢产物以及对寄主的致病力等多个表型中发挥作用。本研究在前期基础上对野生菌株、NTO1敲除突变菌株和回补菌株进行比较转录组分析，发现差异表达基因涉及分泌蛋白类如糖苷水解酶、次级代谢产物类、蛋白酶和DNA结合蛋白，并对关键基因通过敲除等遗传操作手段来进行功能分析，表型结果揭示其参与病原物生长过程、产孢过程以及细胞壁和细胞膜对胁迫的响应过程，最终反馈到与寄主植物早期互作能力减弱，发生不亲和互作。研究结果表明，NTO1作为乙酰转移酶复合体NuA3中的一个亚基参与染色质组蛋白乙酰化。组蛋白乙酰化水平对于细胞行使正常的功能和生物体的发育过程十分必要，而且在病原菌与寄主植物互作过程中发挥着重要作用。NTO1亚基基因的缺失，可能引起NuA3功能受损，从而导致组蛋白乙酰化水平降低，引起基因转录发生改变，导致敲除突变菌株表型影响效应涉及营养生长、生殖生长及致病力等多个方面。此研究为进一步丰富尖镰孢菌的致病机理和甘蓝枯萎病的防控提供理论依据。