Phosphoproteomics Reveals the Effect of Ethylene in Soybean Root under Flooding Stress

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 4G 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 4G was located in the center of the network during flooding. Soybean eukaryotic translation initiation factor 4G 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 mechanismsof plant tolerance to the initial stages of flooding stress in soybean root tips