SnRKs蛋白激酶对高温强光下番茄气孔运动的调节机制

Tomato is often expose to high temperature and light stress during the summer cultivation. Stomatal movement directly affects the photosynthetic and transpiration of tomato, thus restricting its quality and yield. Therefore, it is very important to explore the mechanism of stomatal movement under high temperature and light stress in tomato. Currently, we have obtained SnRKs overexpressed and silenced plants in tomato through 35S overexpression and CRISPR-Cas 9 gene editing techniques, respectively. Firstly, we are planning to understand the effect of SnRKs on stomatal movement by observing the stomata parameters in SnRKs silencing, overexpressing and wild-type plants based on our previously results. Secondly, the key genes or proteins and its phosphorylation sites regulated by SnRKs under high temperature and light stress will obtained through comprehensive analysis of transcriptome and phosphoproteome data. Thirdly, to verify the interaction between SnRKs and these key proteins, we employed Yeast two hybrid assay (Y2H), Pull down, and BiFC. Then, the specific interactive sites between SnRKs and these key proteins verified by site-directed mutagenesis. Finally, the mechanism of SnRKs-regulated stomatal movement under high temperature and light stress in tomato will be lay out, This result help to provide a theoretical basis for improving the yield and quality of tomato cultivation in summer facilities.

番茄在设施越夏栽培中常遭受高温强光胁迫，其气孔运动直接影响光合和蒸腾作用，进而影响其品质和产量，因此，探明番茄高温强光下气孔运动机制极为重要。申请者已通过35S过表达和CRISPR-Cas9基因编辑技术在番茄中分别获得了SnRKs过表达和沉默植株。本项目拟以番茄SnRKs沉默、过表达和野生型为试材，首先，通过对三种植株对照组和高温强光组的气孔参数测定，探明SnRKs对气孔运动的影响作用；其次，综合分析转录组和磷酸化蛋白质组数据，挖掘SnRKs在高温强光下调节气孔运动的下游关键基因/蛋白及其磷酸化位点；再次，通过酵母双杂、Pull down、BiFC实验验证SnRKs与上述关键蛋白的互作关系，并通过对磷酸化位点的定点突变，验证SnRKs与这些关键蛋白的具体作用位点。研究结果有助于解析番茄SnRKs对高温强光下气孔运动的调节机制，为提高设施番茄越夏栽培的产量和品质提供理论依据。

番茄在设施越夏栽培中常遭受高温胁迫，气孔运动直接影响光合和蒸腾作用，进而影响其品质和产量，因此，探明番茄高温下气孔运动机制极为重要。基于此，本研究通过CRISPR-Cas9基因编辑技术在番茄中获得了ABA诱导的调节气孔运动的关键基因SlSnRKs（蔗糖非发酵1相关蛋白激酶）沉默的T2代植株。通过对气孔参数的观察，发现SlSnRK2.3调节高温胁迫下气孔关闭与ABA（脱落酸）积累量、气孔发育相关基因的表达和ROS（活性氧）信号通路的激活有关。为了筛选与SlSnRK2.3相互作用的蛋白质，我们构建了筛选cDNA文库，发现SUI（蛋白质翻译因子SUI1同源物）与SlSnRK2.3相互作用。通过Y2H、LUC和BiFC实验进一步验证SlSnRK2.3和SlSUI1存在互作关系。为进一步验证SlSUI1的功能，采用农杆菌侵染和瞬时转化的方式，沉默了SlSUI1，发现SlSnRK2.3/SlSUI1在气孔运动中的功能类似，SlSUI1影响高温下的ROS平衡，进而影响高温耐受性。研究结果有助于解析番茄SnRK2.3对高温下气孔运动的调节机制，为提高设施番茄越夏栽培的产量和品质提供理论依据。