拟南芥HSB1基因参与调控茉莉酸积累和植物抗病性的作用机制

Developing and planting disease-resistant varieties is one of the effective ways of combating crop losses caused by diseases and insects. Oriented towards the identification of key components and molecular mechanisms involved in plant defense responses against fungal infections, a focused genetic screening was carried out which led to the isolation of an Arabidopsis mutant, hsb1 (hyper-susceptible to botrytis 1) which shows more susceptible to infections by Botrytis cinerea as well as to attacks by herbivorous insect Spodoptera exigua than the wild type plants. But response to bacterial pathogen Pst DC3000 inoculations and inhibitory action of auxin on root elongation are not compromised by HSB1 mutations. HSB1 mutations cause increased sensitiveness to JA-inhibited seedling growth and JA-induced leaf senescence. Over-accumulations of jasmonates, H2O2 and NO and increased sensitivity of seed germination to supplied oxalic acid were also observed in hsb1 mutants. These results suggest the involvement of HSB1 in modulating cellular redox homeostasis, jasmonates accumulation and plant defense responses. To define the mechanisms of HSB1 functions, we propose to employ biochemical, transgenomic, genetic, and physiological approaches to: 1. study the expression pattern and subcellular location of HSB1 and its responsiveness to applied jasmonates, H2O2 and to fungal infections; 2. identify the interacting partners and co-regulated genes of HSB1; 3. observe whether the induced H2O2 and NO productions and calcium signaling by applied MeJA, H2O2 and fungal infections are altered by hsb1 mutations; 4. examine the possible impact on the biosynthesis of ethylene and its precursor ACC by hsb1 mutations and 5. to understand how HSB1 coordinates changes of cellular redox state and the interplay of JA, ethylene signaling pathways in response to biotic stresses.

理解植物抗病性的分子机制，是抗病育种和保障粮食安全的需要。在未发表的前期研究中，我们发现HSB1基因功能缺失导致植物对灰霉菌侵染和甜菜夜蛾幼虫取食的抗性降低；突变体对草酸抑制种子萌发、茉莉酸阻遏幼苗发育、茉莉酸诱导叶片衰老的敏感性均显著提高；hsb1突变在引起JA、JA-Ile、H2O2和NO等显著积累的同时可能下调乙烯的生物合成但不影响生长素抑制主根伸长反应。初步推测，HSB1参与调节茉莉酸合成、细胞氧化还原平衡以及植物的防御反应。在本申请项目中，我们将分析HSB1的表达特点及其对外源茉莉酸、H2O2、真菌侵染的响应能力，通过基因定点突变和转化截短片段等鉴定核心功能单元，定量乙烯及其合成前体的含量以及PDF1.2等抗病基因的表达，检测细胞氧化还原防护系统状态及相关基因的表达，分离HSB1的互作蛋白和共调控基因；明确HSB1在H2O2、NO和JA信号介导的植物防御反应中的作用及其分子机制。

茉莉酸及其衍生产物作为一类具有具有环戊烷酮结构的植物激素，在植物防御病虫害中起至关重要的作用。通过开展本项目，我们发现HSB1/BIG基因突变后，导致植物对病原细菌和病原真菌的抗性显著降低以及植物生长缓慢、开花延迟、种子显著变小和主根变短等多重表型。深入研究后发现，外源施加或者通过机械伤害诱导的内源茉莉酸类物质能非常特异地下调HSB1/BIG基因的转录水平，而HSB1/BIG的功能突变体中茉莉酸及其衍生产物具有较高的本底积累水平，突变体对茉莉酸甲酯诱导的植物生长抑制和叶片衰老的敏感性增强。当在突变体中引入coi1突变则可以显著回复主根短和种子小等茉莉酸的抑制效应。相比于野生型对照植物，在病原菌侵染和机械伤害等处理下，突变体显著积累更多的茉莉酸类物质。 与此观察相一致的是，突变体的水杨酸信号途径被显著抑制，并导致依赖于该途径的气孔免疫和植物防御丁香假单胞杆菌的能力严重受损。我们的工作表明，HSB1/BIG是茉莉酸信号通路的负调控子，其表达受茉莉酸积累的下调，从而使得植物既可以防止茉莉酸过量积累抑制生长，也可以在发育和逆境因子的诱导下迅速积累茉莉酸启动适应性反应。有意思地是，我们的研究还发现HSB1/BIG基因突变在促进茉莉酸信号合成的同时却显著抑制乙烯的合成，HSB1/BIG对JA和乙烯的协同过程中不可或缺， 其功能缺失破坏了茉莉酸和乙烯信号途径在植物发育和抗逆过程的协同作用，但是具体的作用机制还有待深入解析。简而言之， 在本项目中我们发现HSB1/BIG作为一个全新的茉莉酸信号调控元件在植物调控茉莉酸稳态及其与乙烯相互作用中发挥重要作用，深化了我们对植物防御反应机制的理解，对农作物的抗病育种有促进作用。另外，共有8名研究生的科研训练受益于该项目的资助，包括已经毕业的4名博士生和2名硕士生。我们取得的研究发现，已经分别发表在New Phytologist、Plant Physiology and Biochemistry和Journal of Plant Physiology等杂志上，并应邀撰写《Stomata》专题综述文章一篇。