热激蛋白AtHSP70X在拟南芥萼片开裂中的功能及机理研究

Sepal open, the result of the combined effect of both interior and exterior factors, is the critical developmental stage in plants that is closely associated with pollination and fertilization, thus, unraveling its regulatory mechanisms including the interaction between interior and exterior factors is essential not only to the understanding of the process of plant reproductive development, but also to its application in agriculture. Our preliminary results showed the involvement of an Arabidopsis heat shock protein 70 (AtHSP70X) in the process of sepal open. The mutant showed abnormal sepal epidermis lacking ridge-like cutin, which resulted in the fusion between two lateral sepals, leading to the failure of sepal open and flower opening and the reduction in fertility without affecting the function of other reproductive organs. In this proposal, we will first to define the genetic function of AtHSP70X in sepal open using genetic approaches, then to explore the possible ATPase activity and potential interaction proteins of AtHSP70X by biochemical approaches, and to screen the AtHSP70X mediated pathways and gene networks via integrated metabolomic and transcriptomic methods, with the aim to unravel the functions and underlying mechanisms of AtHSP70X in sepal open. The result of this study is important not only to improve understandings of the regulatory mechanisms of flower opening, but also to facilitate the mechanism studies on the involvement of HSP and cuticular lipids in plant development and stress response.

萼片开裂是决定植物能否正常授粉受精的关键生物学步骤，是内部信号和外部环境共同作用所致，其调控机理包括内外影响因子互作机理的研究对了解植物生殖发育过程及农业应用具有重要的价值。我们前期研究发现热激蛋白AtHSP70X参与调控拟南芥萼片开裂。突变体两个旁侧萼片因表面脊状角质缺失发生融合，阻碍了萼片开裂和花朵开放，导致育性下降，但其它花器官功能正常。在此基础上，本项目拟以athsp70x为试材，进一步采用遗传学方法明确AtHSP70X在萼片开裂中的遗传功能，并尝试用生化手段分析AtHSP70X的蛋白定位、ATP酶活性和互作蛋白等生化功能，结合芯片分析和角质层脂及膜脂的代谢测定解析AtHSP70X可能影响的代谢通路和基因网络，期望系统阐明AtHSP70X在萼片开裂过程中的功能及机理。该研究成果将完善对植物花朵开放调控机理的认识，为HSP和角质层脂参与植物生长发育和逆境应答机制的研究提供理论借鉴。

萼片开裂是决定植物能否正常授粉受精的关键生物学步骤，是内部信号和外部环境共同作用的结果，研究其调控机理对了解植物生殖发育过程及农业应用具有重要的价值。我们前期研究发现，拟南芥热激蛋白70基因（AtHSP70-16）的突变导致了部分花蕾萼片发生生殖后融合，这些花朵不能正常开放，这种表型在轻度高温逆境下更严重。本研究从遗传、生化和代谢三个方面系统研究了AtHSP70-16在常温（22oC）及轻度高温（27oC）逆境下拟南芥花朵萼片开裂中的功能及分子机理。我们的研究结果表明，常温下，作为分子伴侣，AtHSP70-16通过维持特定类型蛋白的稳态（包括蛋白激酶、转录因子、HSFs和HSPs等）及相关代谢稳态（激素、ROS、脂质/角质层和细胞壁等），促进萼片正常角质层的形成，维持花朵的正常开放；AtHSP70-16功能一旦缺失，萼片无法感知并正确传导上述信号，导致角质层代谢失调，减少了萼片表面蜡质合成，诱发萼片融合，限制了花朵的正常开放、授粉受精和结实。轻度高温逆境下，AtHSP70-16通过影响HSFs和其它HSPs影响萼片的热激反应，并通过影响蜡质合成和调控基因的表达影响角质层代谢；AtHSP70-16的缺失不仅影响到萼片的高温胁迫反应网络，也影响到角质层的正常形成，加重了对植物花朵开放的负面影响。本研究第一次全面揭示了萼片在协调植物花器官生长发育信号和外界温度变化信号方面的作用及分子机制，丰富和完善了对植物花朵开放调控分子机理的认识，拓展了对HSP蛋白功能的认识，搭建了HSP和角质层脂类代谢之间的桥梁，为植物生长发育和逆境应答分子机制的研究提供重要借鉴。