Fig 1.
Growth of bean (Phaseolus vulgaris L. var. soisson nain hâtif) seedlings germinated in the presence of distilled water or 200 μM CuCl2.
Fig 2.
Time courses of (A, B) germination rate and (C, D, E, F) seedling length in bean seeds in the absence (CTR) and in the presence of CuCl2 (concentrations 20, 50, 100, 200, 500 μM).
Letters indicate significant differences compared with the respective control sample (a: p < 0.05, b: p < 0.01 and c: p < 0.001).
Table 1.
Level of H2O2 and activities of antioxidant enzymes in the seedlings after 3 days (a) and cotyledons after 9 days (b) of germinated bean seeds in the presence of H2O (CTR) or 200 μM Cu.
Fig 3.
Enzymatic activities of SOD, CAT and peroxidases (APX, GPX and POX) in (A, C) seedlings and (B, D) cotyledons of bean seeds during germination in the presence of distilled water (CTR) or 200 μM Cu.
Values are means ± SE (n = 5). Letters indicate significant differences compared with the respective control sample (a: p < 0.05, b: p < 0.01 and c: p < 0.001).
Table 2.
Levels of protein carbonyl and thiol groups in the seedlings (3 days-old) and the cotyledons (9 days-old) of germinated bean seeds in the presence of H2O (CTR) or 200 μM Cu.
Table 3.
Activities of redox enzymes in the seedlings (3 days-old) and the cotyledons (9 days-old) of germinated bean seeds in the presence of H2O (CTR) or 200 μM Cu.
Table 4.
Activities of NAD(P)H oxidases and redox ratios of coenzymes in the seedlings (3 days-old) and the cotyledons (9 days-old) of germinated bean seeds in the presence of H2O (CTR) or 200 μM Cu.
Fig 4.
Representative images of 1DE gels of proteins (100 μg) in seedlings of bean seeds germinated for 3 days. (A) In the presence of distilled water (CTR) or (B) 200 μM Cu. Gels were stained with Coomassie G-250 (scanned with GS-800 calibrated densitometer) and with IAF labeling (scanned with Typhoon 9400 scanner).
Total optical densities for each lane obtained from IAF staining were normalized with those from Coomassie G-250 staining of the same gel. (C, D) Levels of proteins containing thiol groups. Values shown are (C) means of 4 biological replicates (±SD) numbered from 1 to 4, and (D) means of 4 technical replicates (±SD). Each measurement was performed in an extract obtained from several seedlings. Analyses were performed using ANOVA, student’s T test; *p<0.05, **p<0.01, ***p<0.001.
Fig 5.
Representative images of 1DE gels of proteins (150 μg) in cotyledons of bean seeds germinated for 9 days in the presence of (A) distilled water (CTR) or (B) 200 μM Cu. Gels were stained with Coomassie G-250 (scanned with GS-800 calibrated densitometer) and with IAF labeling (scanned with Typhoon 9400 scanner).
Total optical densities for each lane obtained from IAF staining were normalized with those from Coomassie G-250 staining of the same gel. (C, D) Levels of proteins containing thiol groups. Values shown are (C) means of 4 biological replicates (±SD) numbered from 1 to 4, and (D) means of 4 technical replicates (±SD). Each measurement was performed in an extract obtained from several cotyledons. Analyses were performed using ANOVA, student’s T test; **p<0.01, ***p<0.001.
Fig 6.
Profiles of the expression of proteins containing thiol groups in (A, B) seedlings and (C, D) cotyledons of bean seeds germinated for 9 days in the presence of distilled water (CTR) or 200 μM Cu.
Proteins (800 μg) were labeled with IAF and separated by 2-D SDS-PAGE. Figures show spots of interest in representative gels from (A, C) colloidal Coomassie Brilliant G-250 staining (scanned with GS-800 calibrated densitometer) and (B, D) IAF labeling (scanned with Typhoon 9400 scanner; 800 PMT). Numbers correspond to spots of p<0.05 and Fold induction >1.5 (spots identified).
Fig 7.
Representative images of 1DE gels of proteins (150 μg) in seedlings of bean seeds germinated for 9 days in (A) the presence of distilled water (CTR) or (B) 200 μM Cu.
Gels were stained with Coomassie G-250 (scanned with GS-800 calibrated densitometer) and with FTSC labeling (scanned with Typhoon 9400 scanner). Total optical densities for each lane obtained from FTSC staining were normalized with those from Coomassie G-250 staining of the same gel. (C, D) Levels of proteins containing carbonyl groups. Values shown are means of (C) 4 biological replicates (±SD) numbered from 1 to 4, and means of (D) 4 technical replicates (±SD). Each measurement was performed in an extract obtained from several cotyledons. Analyses were performed using ANOVA, student’s T test; *p<0.05, ***p<0.001.
Fig 8.
Representative images of 1DE gels of proteins (100 μg) in cotyledons of bean seeds germinated for 3 days in the presence of (A) distilled water (CTR) or (B) 200 μM Cu.
Gels were stained with Coomassie G-250 (scanned with GS-800 calibrated densitometer) and with FTSC labeling (scanned with Typhoon 9400 scanner). Total optical densities for each lane obtained from FTSC staining were normalized with those from Coomassie G-250 staining of the same gel. (C, D) Levels of proteins containing carbonyl groups. Values shown are (C) means of 4 biological replicates (±SD) numbered from 1 to 4, and (D) means of 4 technical replicates (±SD). Each measurement was performed in an extract obtained from several cotyledons. Analyses were performed using ANOVA, student’s T test; ***p<0.001.
Fig 9.
Profiles of the expression of proteins containing carbonyl groups in (A, B) seedlings and (C, D) cotyledons of bean seeds germinated for 9 days in the presence of distilled water (CTR) or 200 μM Cu.
Proteins (1200 μg) were labeled with FTSC and separated by 2-D SDS-PAGE. Figures show spots of interest in representative gels from (A) colloidal Coomassie Brilliant G-250 staining (scanned with GS-800 calibrated densitometer) and (B) FTSC labeling (scanned with Typhoon 9400 scanner; 600 PMT). Numbers correspond to spots of p<0.05 and Fold induction >1.5 (spots identified).
Table 5.
Proteins containing thiol groups (IAF labeling) in the seedlings (3 days-old) (a) and the cotyledons (9 days-old) (b) of germinated bean seeds in the presence of H2O (CTR) or 200 μM Cu.
Table 6.
Proteins containing carbonyl groups (FTSC labeling) in the seedlings (3 days-old) (a) and the cotyledons (9 days-old) (b) of germinated bean seeds in the presence of H2O (CTR) or 200 μM Cu.