Laboratory of Plant Molecular Physiology, Graduate School of Agricultural and Life Sciences, The University of Tokyo, Tokyo, Japan.
Graduate School of Industrial Science and Technology, Tokyo University of Science, Tokyo, Japan.
Methods Mol Biol. 2022;2462:181-189. doi: 10.1007/978-1-0716-2156-1_14.
Abscisic acid (ABA) is a key phytohormone involved in plant development, seed germination and responses to osmotic stresses, such as drought and high salinity. SNF1-related protein kinases (SnRK2s) play important roles in ABA-dependent and ABA-independent osmotic stress signaling. SnRK2s phosphorylate transcription factors and ion channels in response to ABA or osmotic stress to induce the expression of stress-responsive genes and stomatal closure, respectively, to confer osmotic stress tolerance. The activity of SnRK2s is directly or indirectly regulated by several protein factors. Identification of downstream substrates or upstream regulators of SnRK2s is very useful for elucidating protein components that regulate ABA and osmotic stress signaling. Here, we describe the use of affinity purification by coimmunoprecipitation and liquid chromatography-tandem mass spectrometry to identify protein complexes involved in ABA and osmotic stress signaling in plants. We previously identified several protein factors that regulate ABA and osmotic stress signaling by using this method.
脱落酸（ABA）是一种参与植物发育、种子萌发和应对渗透胁迫（如干旱和高盐）的关键植物激素。SNF1 相关蛋白激酶（SnRK2s）在 ABA 依赖和非依赖的渗透胁迫信号转导中发挥重要作用。SnRK2s 磷酸化转录因子和离子通道，以响应 ABA 或渗透胁迫，分别诱导应激响应基因的表达和气孔关闭，从而赋予渗透胁迫耐受性。SnRK2s 的活性直接或间接受到几种蛋白因子的调节。鉴定 SnRK2s 的下游底物或上游调节剂对于阐明调节 ABA 和渗透胁迫信号转导的蛋白成分非常有用。在这里，我们描述了使用共免疫沉淀和液相色谱-串联质谱联用技术进行亲和纯化，以鉴定参与植物中 ABA 和渗透胁迫信号转导的蛋白质复合物。我们之前使用这种方法鉴定了几种调节 ABA 和渗透胁迫信号转导的蛋白因子。
