Proteomic and Biochemical Analyses of the Mechanism of Tolerance in Mutant Soybean Responding to Flooding Stress

Abstract

To investigate the mechanism of flooding tolerance of soybean, flooding-tolerant mutants derived from gamma-ray irradiated soybean were crossed with parent cultivar Enrei for removal of other factors besides the genes related to flooding tolerance in primary generated mutant soybean. Although the growth of the wild type was significantly suppressed by flooding compared with the non-flooding condition, that of the mutant lines was better than that of the wild type even if it was treated with flooding. A two-day-old mutant line was subjected to flooding for 2 days and proteins were analyzed using a gel-free/label-free proteomic technique. Oppositely changed proteins in abundance between the wild type and mutant line under flooding stress were associated in endoplasmic reticulum according to gene-ontology categorization. Immunoblot analysis confirmed that calnexin accumulation increased in both the wild type and mutant line; however, calreticulin accumulated in only the mutant line under flooding stress. Furthermore, although glycoproteins in the wild type decreased by flooding compared with the non-flooding condition, those in the mutant line increased even if it was under flooding stress. Alcohol dehydrogenase accumulated in the wild type and mutant line; however, this enzyme activity significantly increased and mildly increased in the wild type and mutant line, respectively, under flooding stress compared with the non-flooding condition. Cell death increased and decreased in the wild type and mutant line, respectively, by flooding stress. These results suggest that the regulation of cell death through the fermentation system and glycoprotein folding might be an important factor for the acquisition of flooding tolerance in mutant soybean.