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Gel-free proteomic analysis of soybean root proteins affected by calcium under flooding stress.

Oh M, Nanjo Y, Komatsu S - Front Plant Sci (2014)

Bottom Line: Soybean is sensitive to flooding stress and exhibits reduced growth under flooding conditions.Proteins involved in protein degradation/synthesis/posttranslational modification, hormone/cell wall metabolisms, and DNA synthesis were decreased by flooding stress; however, their reductions were recovered by calcium treatment.Fermentation and glycolysis-related proteins were increased in response to flooding; however, these proteins were not affected by calcium supplementation.

View Article: PubMed Central - PubMed

Affiliation: Life Sciences and Bioengineering, Graduate School of Life and Environmental Sciences, University of Tsukuba Tsukuba, Japan ; National Institute of Crop Science, National Agriculture and Food Research Organization Tsukuba, Japan.

ABSTRACT
Soybean is sensitive to flooding stress and exhibits reduced growth under flooding conditions. To better understand the flooding-responsive mechanisms of soybean, the effect of exogenous calcium on flooding-stressed soybeans was analyzed using proteomic technique. An increase in exogenous calcium levels enhanced soybean root elongation and suppressed the cell death of root tip under flooding stress. Proteins were extracted from the roots of 4-day-old soybean seedlings exposed to flooding stress without or with calcium for 2 days and analyzed using gel-free proteomic technique. Proteins involved in protein degradation/synthesis/posttranslational modification, hormone/cell wall metabolisms, and DNA synthesis were decreased by flooding stress; however, their reductions were recovered by calcium treatment. Development, lipid metabolism, and signaling-related proteins were increased in soybean roots when calcium was supplied under flooding stress. Fermentation and glycolysis-related proteins were increased in response to flooding; however, these proteins were not affected by calcium supplementation. Furthermore, urease and copper chaperone proteins exhibited similar profiles in 4-day-old untreated soybeans and 4-day-old soybeans exposed to flooding for 2 days in the presence of calcium. These results suggest that calcium might affect the cell wall/hormone metabolisms, protein degradation/synthesis, and DNA synthesis in soybean roots under flooding stress.

No MeSH data available.


Related in: MedlinePlus

Effects of calcium on the mRNA expression levels of urease and copper chaperone proteins in different organs of soybean under flooding stress. Two-day-old soybeans were flooded without or with 50 mM CaCl2 for 1 and 2 days. Untreated soybeans were used as a control. RNAs extracted from roots, hypocotyls, and cotyledons of the soybeans were analyzed by qRT-PCR with specific primers for urease and copper chaperone (Supplemental Table 1). Relative mRNA abundance was normalized against that of 18S rRNA. Data are shown as means ± SD from three independent biological replicates. Means with the same letter are not significantly different according to ANOVA (P < 0.05).
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Figure 5: Effects of calcium on the mRNA expression levels of urease and copper chaperone proteins in different organs of soybean under flooding stress. Two-day-old soybeans were flooded without or with 50 mM CaCl2 for 1 and 2 days. Untreated soybeans were used as a control. RNAs extracted from roots, hypocotyls, and cotyledons of the soybeans were analyzed by qRT-PCR with specific primers for urease and copper chaperone (Supplemental Table 1). Relative mRNA abundance was normalized against that of 18S rRNA. Data are shown as means ± SD from three independent biological replicates. Means with the same letter are not significantly different according to ANOVA (P < 0.05).

Mentions: Six differentially changed proteins were common between 4-day-old soybeans and 2-day-flooded soybeans treated with calcium (Figure 3). The abundance of these proteins was calculated among 4-day-old soybeans, 2-day-flooded soybeans, and 2-day-flooded soybeans treated with 50 mM CaCl2 (Supplemental Figure 5). The analysis of protein abundance indicated that urease (Glyma05g27840.1) and two copper chaperones (Glyma10g14110.1 and Glyma02g19380.1) exhibited the same profiles in 4-day-old soybeans and 2-day-flooded soybeans treated with 50 mM CaCl2. To determine whether the changes in protein abundance were regulated at the transcriptional level, the mRNA expression levels of these proteins were analyzed in root, hypocotyl, and cotyledon under flooding for 0, 1, and 2 days (Figure 5). Total RNAs extracted from roots, hypocotyls, and cotyledons of soybeans were analyzed using qRT-PCR. In roots, the mRNA level of urease was down-regulated by 1 and 2 days flooding; however, the level was not changed by calcium supplementation. In hypocotyls, the mRNA level of urease was down-regulated by 1 day of flooding and significantly up-regulated by 1 day flooding with calcium; however, there was no significant between 2 days flooding without and with calcium. The mRNA expression level of copper chaperone was similar to their protein level in the root and hypocotyl under 2 days flooding with calcium. In roots and hypocotyls, the level of copper chaperone was down-regulated by 1 day flooding without and with calcium; whereas the level was significantly up-regulated by 2 days flooding with calcium. In cotyledons, the mRNA levels of urease and copper chaperone were significantly up-regulated by 1 and 2 day flooding when calcium was exogenously added (Figure 5).


Gel-free proteomic analysis of soybean root proteins affected by calcium under flooding stress.

Oh M, Nanjo Y, Komatsu S - Front Plant Sci (2014)

Effects of calcium on the mRNA expression levels of urease and copper chaperone proteins in different organs of soybean under flooding stress. Two-day-old soybeans were flooded without or with 50 mM CaCl2 for 1 and 2 days. Untreated soybeans were used as a control. RNAs extracted from roots, hypocotyls, and cotyledons of the soybeans were analyzed by qRT-PCR with specific primers for urease and copper chaperone (Supplemental Table 1). Relative mRNA abundance was normalized against that of 18S rRNA. Data are shown as means ± SD from three independent biological replicates. Means with the same letter are not significantly different according to ANOVA (P < 0.05).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4202786&req=5

Figure 5: Effects of calcium on the mRNA expression levels of urease and copper chaperone proteins in different organs of soybean under flooding stress. Two-day-old soybeans were flooded without or with 50 mM CaCl2 for 1 and 2 days. Untreated soybeans were used as a control. RNAs extracted from roots, hypocotyls, and cotyledons of the soybeans were analyzed by qRT-PCR with specific primers for urease and copper chaperone (Supplemental Table 1). Relative mRNA abundance was normalized against that of 18S rRNA. Data are shown as means ± SD from three independent biological replicates. Means with the same letter are not significantly different according to ANOVA (P < 0.05).
Mentions: Six differentially changed proteins were common between 4-day-old soybeans and 2-day-flooded soybeans treated with calcium (Figure 3). The abundance of these proteins was calculated among 4-day-old soybeans, 2-day-flooded soybeans, and 2-day-flooded soybeans treated with 50 mM CaCl2 (Supplemental Figure 5). The analysis of protein abundance indicated that urease (Glyma05g27840.1) and two copper chaperones (Glyma10g14110.1 and Glyma02g19380.1) exhibited the same profiles in 4-day-old soybeans and 2-day-flooded soybeans treated with 50 mM CaCl2. To determine whether the changes in protein abundance were regulated at the transcriptional level, the mRNA expression levels of these proteins were analyzed in root, hypocotyl, and cotyledon under flooding for 0, 1, and 2 days (Figure 5). Total RNAs extracted from roots, hypocotyls, and cotyledons of soybeans were analyzed using qRT-PCR. In roots, the mRNA level of urease was down-regulated by 1 and 2 days flooding; however, the level was not changed by calcium supplementation. In hypocotyls, the mRNA level of urease was down-regulated by 1 day of flooding and significantly up-regulated by 1 day flooding with calcium; however, there was no significant between 2 days flooding without and with calcium. The mRNA expression level of copper chaperone was similar to their protein level in the root and hypocotyl under 2 days flooding with calcium. In roots and hypocotyls, the level of copper chaperone was down-regulated by 1 day flooding without and with calcium; whereas the level was significantly up-regulated by 2 days flooding with calcium. In cotyledons, the mRNA levels of urease and copper chaperone were significantly up-regulated by 1 and 2 day flooding when calcium was exogenously added (Figure 5).

Bottom Line: Soybean is sensitive to flooding stress and exhibits reduced growth under flooding conditions.Proteins involved in protein degradation/synthesis/posttranslational modification, hormone/cell wall metabolisms, and DNA synthesis were decreased by flooding stress; however, their reductions were recovered by calcium treatment.Fermentation and glycolysis-related proteins were increased in response to flooding; however, these proteins were not affected by calcium supplementation.

View Article: PubMed Central - PubMed

Affiliation: Life Sciences and Bioengineering, Graduate School of Life and Environmental Sciences, University of Tsukuba Tsukuba, Japan ; National Institute of Crop Science, National Agriculture and Food Research Organization Tsukuba, Japan.

ABSTRACT
Soybean is sensitive to flooding stress and exhibits reduced growth under flooding conditions. To better understand the flooding-responsive mechanisms of soybean, the effect of exogenous calcium on flooding-stressed soybeans was analyzed using proteomic technique. An increase in exogenous calcium levels enhanced soybean root elongation and suppressed the cell death of root tip under flooding stress. Proteins were extracted from the roots of 4-day-old soybean seedlings exposed to flooding stress without or with calcium for 2 days and analyzed using gel-free proteomic technique. Proteins involved in protein degradation/synthesis/posttranslational modification, hormone/cell wall metabolisms, and DNA synthesis were decreased by flooding stress; however, their reductions were recovered by calcium treatment. Development, lipid metabolism, and signaling-related proteins were increased in soybean roots when calcium was supplied under flooding stress. Fermentation and glycolysis-related proteins were increased in response to flooding; however, these proteins were not affected by calcium supplementation. Furthermore, urease and copper chaperone proteins exhibited similar profiles in 4-day-old untreated soybeans and 4-day-old soybeans exposed to flooding for 2 days in the presence of calcium. These results suggest that calcium might affect the cell wall/hormone metabolisms, protein degradation/synthesis, and DNA synthesis in soybean roots under flooding stress.

No MeSH data available.


Related in: MedlinePlus