Yin Xiaojian, Sakata Katsumi, Komatsu Setsuko
Graduate School of Life and Environmental Sciences, University of Tsukuba , Tsukuba 305-8572, Japan.
J Proteome Res. 2014 Dec 5;13(12):5618-34. doi: 10.1021/pr500621c. Epub 2014 Oct 24.
Flooding has severe negative effects on soybean growth. To explore the flooding-responsive mechanisms in early-stage soybean, a phosphoproteomic approach was used. Two-day-old soybean plants were treated without or with flooding for 3, 6, 12, and 24 h, and root tip proteins were then extracted and analyzed at each time point. After 3 h of flooding exposure, the fresh weight of soybeans increased, whereas the ATP content of soybean root tips decreased. Using a gel-free proteomic technique, a total of 114 phosphoproteins were identified in the root tip samples, and 34 of the phosphoproteins were significantly changed with respect to phosphorylation status after 3 h of flooding stress. Among these phosphoproteins, eukaryotic translation initiation factors were dephosphorylated, whereas several protein synthesis-related proteins were phosphorylated. The mRNA expression levels of sucrose phosphate synthase 1F and eukaryotic translation initiation factor 4 G were down-regulated, whereas UDP-glucose 6-dehydrogenase mRNA expression was up-regulated during growth but down-regulated under flooding stress. Furthermore, bioinformatic protein interaction analysis of flooding-responsive proteins based on temporal phosphorylation patterns indicated that eukaryotic translation initiation factor 4 G was located in the center of the network during flooding. Soybean eukaryotic translation initiation factor 4 G has homology to programmed cell death 4 protein and is implicated in ethylene signaling. The weight of soybeans was increased with treatment by an ethylene-releasing agent under flooding condition, but it was decreased when plants were exposed to an ethylene receptor antagonist. These results suggest that the ethylene signaling pathway plays an important role, via the protein phosphorylation, in mechanisms of plant tolerance to the initial stages of flooding stress in soybean root tips.
洪水对大豆生长具有严重的负面影响。为了探究大豆早期阶段对洪水的响应机制，采用了磷酸化蛋白质组学方法。对两天大的大豆植株进行无洪水处理或分别进行3、6、12和24小时的洪水处理，然后在每个时间点提取并分析根尖蛋白质。洪水处理3小时后，大豆鲜重增加，而根尖ATP含量下降。使用无凝胶蛋白质组学技术，在根尖样品中总共鉴定出114种磷酸化蛋白质，其中34种磷酸化蛋白质在洪水胁迫3小时后其磷酸化状态发生了显著变化。在这些磷酸化蛋白质中，真核生物翻译起始因子发生去磷酸化，而几种与蛋白质合成相关的蛋白质发生磷酸化。蔗糖磷酸合酶1F和真核生物翻译起始因子4G的mRNA表达水平下调，而UDP-葡萄糖6-脱氢酶mRNA表达在生长过程中上调，但在洪水胁迫下下调。此外，基于时间磷酸化模式对洪水响应蛋白进行的生物信息学蛋白质相互作用分析表明，在洪水期间真核生物翻译起始因子4G位于网络中心。大豆真核生物翻译起始因子4G与程序性细胞死亡4蛋白具有同源性，并参与乙烯信号传导。在洪水条件下，用乙烯释放剂处理可增加大豆重量，但当植株暴露于乙烯受体拮抗剂时重量会降低。这些结果表明，乙烯信号通路通过蛋白质磷酸化在大豆根尖对洪水胁迫初期的植物耐受性机制中起重要作用。
