稻曲病菌分泌蛋白UvScd1在致病过程中的作用及其机制研究

Rice false smut disease caused by Ustilaginoidea virens is one of the most destructive rice diseases worldwide, causing severe economic loss for rice production, and also produce toxins that threaten health of human and animals. U. virens usually secretes number of proteins to manipulate host immunity during infection. Previous studies indicated that a secretory protein UvScd1 can suppress the hypersensitive cell death induced by elicitor and can interact with the rice protein EXP, one of the expansins which play important roles for cell wall loosening. Rice filament elongation, pollen germination and ovary development were affected by U. virens infection. We speculate that the UvScd1 can regulate rice immunity and the development of reproduction organ through interacting with EXP. Thus, this project was proposed to analyze the biological function of UvScd1. Further, the biological significance of interaction will be demonstrated by investigating the influence of the interaction on fungal pathogenicity, as well as the resistance and development of reproduction organs in rice. The results will enhance the understanding of mechanisms through which U. virens manipulates the development of rice reproduction organs, thus benefits the virulence, and provide theoretical bases for the management of rice false smut and the molecular breeding of resistant cultivars.

稻曲病是由稻曲病菌引起的一种世界性分布的水稻穗部病害，该病不仅引起水稻产量的巨大损失，而且产生的毒素严重影响稻米食用安全。稻曲病菌在侵染过程中会分泌大量的蛋白来帮助侵染。申请人前期研究发现稻曲病菌一个分泌蛋白UvScd1可以抑制植物过敏性坏死反应，并且可与水稻中参与调控细胞壁伸展和松弛的扩展蛋白EXP发生互作。水稻被稻曲病菌侵染之后，花丝伸长、花粉萌发以及子房膨大受到影响。因此推测UvScd1通过和EXP发生互作来影响细胞壁功能，进而对水稻的生殖器官发育和免疫系统进行调控，从而有利于病菌致病。本项目拟解析UvScd1在稻曲病菌侵染过程中的生物学功能，并研究UvScd1与EXP互作对稻曲病菌致病、水稻抗病和生殖器官发育等方面的影响，明确互作的生物学意义。研究结果有助于揭示稻曲病菌调控水稻生殖器官发育和抗病性的分子机理，为稻曲病防控及抗病育种提供新的思路和理论依据。

稻曲病菌侵染水稻引起的稻曲病在我国发生日趋严重，对粮食安全生产造成严重威胁。而且在侵染过程中能够产生毒素，对人和动物的健康产生危害。稻曲病菌在侵染过程中会分泌大量的效应蛋白来调控水稻的抗病反应，从而帮助其侵染和定殖。本项目以UvScd1为研究对象，对其相关的功能进行了分析。实时定量分析发现UvScd1侵染3天时具有较高的表达量。通过多种方法对其信号肽功能进行分析发现其信号肽确实具有分泌功能。UvScd1可以抑制BAX、XEG1以及INF1引起的细胞坏死，对含有的6个半胱氨酸位点逐一进行点突变，并对其抑制坏死的功能进行分析，发现除127位外，其他位点半胱氨酸突变之后丧失了抑制坏死的功能。酵母双杂交发现，UvScd1可以和自身发生互作，说明该蛋白可能通过形成二聚体或多聚体发挥功能。在稻曲病菌中进行UvScd1的敲除和过表达，发现对致病性有影响，说明其参与了病原菌的致病性。异源表达UvScd1的水稻突变体出现了类病斑的症状，叶片上观察到了更多的活性氧积累，抗病相关基因表达量发生变化，对稻曲病菌的抗性降低，但是对稻瘟病菌的抗性增强，说明该蛋白在促进病原菌侵染具有特异性。通过酵母双杂交以及GST-Pull Down等进行验证，表明UvScd1可以特异性的和水稻EXP蛋白发生互作。研究结果加深了对稻曲病菌致病机理的理解，并为水稻抗病育种提供新的研究思路和材料。