锌指蛋白NCA1通过MAPK信号通路参与调控植物逆境响应的分子机制研究

Abiotic stresses impact plant growth severely and lead to a reduction in grain yield. Multiple abiotic stresses lead to accumulation of Reactive Oxygen Species, which activates MAPK signaling pathway and finally responds to multiple stresses. ROS-induced activation of MAPK cascades plays an important role in response to multiple abiotic stresses. However, it is not clear whether other regulators work in ROS-induced MAPK activation. Our previous work found an Arabidopsis mutant nca1-3, in which the gene encodes a RING finger protein NCA1 is mutated, shows sensitive phenotype to saline stress, alkaline stress, oxidative stress and cold stress. Besides, the hydrogen peroxide content in nca1-3 mutant is higher than that in wildtype. NCA1 interacting protein MPK3 and MPK6 were identified by Mass spectrum. Loss function of NCA1 leads to a delay of MPK3/MPK6 kinase activation under ROS treatment. We will further study on the mechanism of NCA1 regulation on ROS-activated MPK3/MPK6 activity. Our study will further illuminate the regulation of ROS-activated MAPK signaling pathway and consummate the mechanism of MAPK-involved response to multiple abiotic stresses in plant. Therefore, this work will provide theoretical basis for breeding development of stress-resistant crops.

非生物胁迫会严重影响植物生长，造成作物减产。多种非生物胁迫都会引起活性氧的积累，而活性氧会激活MAPK信号级联途径，使植物快速响应非生物胁迫。活性氧引发的MAPK信号通路的激活对于植物响应多种胁迫起到重要作用，但活性氧激活MAPK通路的过程中是否存在其他因子的精细调控，这一问题还有待深入研究。本项目的前期工作发现一个拟南芥RING型锌指蛋白突变体nca1-3对盐碱胁迫、氧化胁迫、低温胁迫多种非生物胁迫具有敏感表型，并且突变体中过氧化氢含量高于野生型；质谱检测发现NCA1与MPK3、MPK6相互作用；NCA1的缺失使活性氧引起的MPK3、MPK6激活发生延迟。本项目拟深入研究NCA1调控活性氧激活MPK3、MPK6的分子机制，进一步阐明活性氧激活的MAPK信号通路的调节方式，完善植物通过MAPK通路调控多种逆境响应的机制，为耐逆境农作物新品种培育提供理论基础。

由于植物具有固着生长的特性，植物在生长过程中面临着包括盐碱、低温等多种非生物胁迫，这些胁迫对于植物的生长发育造成严重影响。多种非生物胁迫都会引起植物体内活性氧积累，MAPK信号级联系统能够被活性氧激活，参与植物对多种胁迫的响应，但活性氧激活MAPK的过程中是否存在其他因子的精细调控，这一问题还有待进一步研究。本项目研究发现编码拟南芥RING型锌指蛋白的NCA1基因突变体对盐碱胁迫、氧化胁迫、低温胁迫多种非生物胁迫具有敏感表型，并且突变体中过氧化氢含量高于野生型；互作分析发现NCA1与MPK3、MPK6存在相互作用，体外磷酸化实验证明NCA1增强持续激活形式的MPKK5（即MPKK5DD）激活MPK3、MPK6，体内磷酸化实验证明NCA1对活性氧诱导的MPK3、MPK6快速激活具有重要作用。进一步研究发现，NCA1作为分子伴侣维持MPK3、MPK6的构象，并且影响MAPK信号通路下游ROS应答基因的表达。综上所述，本项目阐明了NCA1通过调控活性氧诱发的MPK3、MPK6激活，进而调节植物通过MAPK信号通路应答多种非生物胁迫的分子机制，为后续通过现代分子生物学技术改造植物耐逆性奠定了理论基础。