低温是植物遭受最普遍的环境胁迫，严重损害作物生长并导致减产，研究植物响应低温胁迫的机制具有重要科学意义和运用价值。Ca2+信号是植物响应低温的主要形式，产生Ca2+信号的内向转运机制得到较多关注，但外向转运知之甚少。研究发现，Ca2+外向转运的Ca2+-ATPase ACA8表达受低温激烈诱导，负调节CBFs及下游一系列冷响应基因的表达。为研究ACA8调节植物响应低温的途径，本项目拟以野生型、缺失型和回补型拟南芥为研究对象：1）分析低温下ACA8缺失对Ca2+信号及质膜Ca2+-ATPase活性的影响；2）通过转录组测序筛选ACA8调节的下游基因和信号通路；3）利用融合蛋白沉淀技术钓取ACA8的互作蛋白；4）进行体内互作验证，确定互作结构域。由此探明ACA8调节植物响应低温的机制，为认识Ca2+外向转运系统对Ca2+信号调节的分子机理提供新知识，为改良作物低温抗性提供新线索。

Cold stress is one of the most significant abiotic stresses of plants, affecting both plant growth and yield potential of agricultural crops. The researches on the mechanism of plants in response to cold stress have important scientific significance and application value. Ca2+ signalling is essential for plant responses to low temperature. Influx mechanisms have received considerable attention because their regulation is of primary importance to initiate a Ca2+ signal. By contrast, less consideration has been given to the role of efflux systems in Ca2+ signalling. In previous research, we found that the expression of ACA8 was quickly responded to low temperature. ACA8 participates in the cold-acclimation response by negatively modulating [Ca2+]cyt and controlling the induction of CBFs and downstream genes, probably by ensuring the proper control of Ca2+ homeostasis in response to low temperature. In this project, we choose the wild-type, the insertion mutant and the rescued line to study the mechanism of ACA8 in response to cold stress. And then we analyze (1) the impact of Ca2+ signalling and Ca2+-ATPase activity by the disruption of ACA8, (2) conducting the transcriptome sequencing to identify potential downstream genes and pathways of ACA8, (3) using the Pulldown-MS to screen interacting proteins of ACA8, and (4) verifying the interactions in vivo and determining the domains of interaction. Finally, we want to reveal the molecular mechanism of ACA8 in response to cold stress and fill gaps in understanding the role of efflux systems in Ca2+ signalling and provide a new candidate gene for improving crop freezing tolerance.