拟南芥气孔应答低空气湿度相关基因的鉴定和作用机理研究

Plant stomata will close in response to reduced atmospheric humidity to maintain the cellular homeostasis in terms of water and physiological status, by which they can enhance the ability to tolerate the stress. There have been extensive studies on ABA signaling pathways induced by soil drought, however there remains elusive on the pathway underlying the response of stomata to reduced atmospheric humidity. We previously identified two components involved in the atmospheric humidity signaling pathway, OST1 and ABA2, and proved that these two components are shared by humidity- and ABA-signaling pathways. We have recently identified 2 new stomatal mutants, one of which is a mutant of protein kinase gene, by screening SALK T-DNA population based on the advanced infrared camera system. In this proposal, we plan to undertake further study by investigating stomatal conductance, the level of ABA synthesis, the change of stomatal aperture after challenged by ABA and expression profiling, identification of interactive proteins and other related studies. Furthermore, we will utilize our newly developed and patented chamber with well-controlled environments to screen for more mututant candidates from Arabidopsis SALK T-DNA population. This project will complete the extensive characterization of the candidate mumtant and deepen our understanding in the mechanism of stomatal response to atmospheric humidity.

植物气孔在大气湿度降低后关闭，保持了细胞的水分和生理状态稳定，提高了植株抗逆性。当前对土壤干旱诱导的气孔ABA信号通路研究较为深入，而对气孔在大气低湿度下关闭的机理还知之甚少。本课题组前期鉴定了气孔湿度信号通路的元件OST1和ABA2，并首次证明二者是湿度信号通路和ABA信号通路的共用元件。为进一步解析气孔湿度信号通路，我们使用高精度红外热成像仪，筛选到两个新气孔湿度突变体，其一为蛋白激酶基因突变体,本项目拟在此基础上，对该突变体展开深入研究，包括测定低湿度下气孔导度、ABA合成水平、气孔对ABA信号应答等生理分析，表达谱测定、互作蛋白质鉴定、与已知湿度信号元件的关系等分子遗传学分析。此外，我们将应用自主设计发明的精密环控舱系统，对拟南芥SALK T-DNA突变群体进行筛选，获得更多湿度突变体并确定基因位点。本项目通过对候选突变体的系统鉴定和分析，来深化对气孔湿度信号转导机制的认识。

早在110多年前，人们就发现植物气孔会在大气湿度降低后关闭，但是对其中的分子机制，目前仍知之甚少。为解析这一生物学过程，本课题组对前期筛选的新气孔湿度突变体c2进行了研究。结果表明，c2突变体的表型是由于T-DNA插入到At2g14160基因的5’端启动子区。该基因编码蛋白质由85个氨基酸组成，含有RNA结合域(RRM/RBD/RNP motifs)，属于RNA结合蛋白家族成员。采用光合仪、红外热成像仪和水分散失测定等方法，确认C2基因是气孔湿度信号通路的重要元件。c2突变体对ABA和第二信使Ca2+不敏感，表明C2蛋白可能也是ABA信号通路的元件其在信号通路中位于Ca2+的下游。此外，C2基因在野生型植株失水处理后表达量逐渐增加，而c2突变体中，C2基因的表达量下降，ABA信号通路关键基因的表达水平发生显著改变，进一步表明C2可能通过ABA信号元件来对湿度信号进行应答。这些研究结果加深了对C2基因功能的认识，为解析气孔湿度应答机制奠定了很好的基础。