免疫共沉淀与质谱联用检测拟南芥SIK1/Hippo互作蛋白

Detection of Arabidopsis SIK1/Hippo Interacting Proteins by Immunoprecipitation and Mass Spectrometry

Hippo通路是迄今为止发现的最重要的负调控动物器官大小的信号通路之一,其核心由两对级联的激酶-脚手架蛋白复合物构成.本实验室在前期工作中首次发现了拟南芥Hippo(Hpo)的同功能基因SIK1,并建立了与Hpo-Mats对应的SIK1-MOB1模块,发现其通过促进退出细胞分裂与细胞延展,参与植物侧生器官大小的调控.为进一步探究SIK1调控器官大小的分子机制,构建了SIK1-GFP转基因株系,用GFP-Trap免疫共沉淀方法捕获与SIK1内源互作的蛋白,并进行质谱分析,获得了SIK1的候选互作蛋白.研究结果将为后续建立完整的植物Hippo信号通路打下基础.

The Hippo pathway is a core signaling pathway that negatively regulates organ sizes in animals.Central to the pathway is a kinase cascade composed of two protein kinases,Hippo and Warts,and their scaffold proteins,Mats and Salvador.Our lab had previously identified the Arabidopsis protein kinase SIK1 as a functional orthologue for Hippo(Hpo),and had established the plant SIK1-MOB1 module as parallel to the animal Hpo-Mats module.By promoting cell cycle exit and cell expansion,SIK1 controls lateral organ size in plants.In order to explore the molecular function of SIK1,transgenic lines carrying SIK1-GFP were generated in this study for immunoprecipitation and subsequent mass spectrometry analysis.A list of candidate proteins was obtained as potential interacting partners for SIK1.The study will facilitate the establishment of a complete plant Hippo signaling pathway.