5 Results
5.1 Influence of K supply on drought-induced leaf senescence in three different barley lines
5.1.8 Primary metabolite changes under drought stress and varied K supply
HR-C-L HR-S-L HR-C-M HR-S-M HR-C-H HR_S_H
Phenylalanine concentration (nmol g-1 FW)
0 20 40 60 80 100 120 140 160
Control Stress Control Stress
Low K High K
Control Stress Medium K b
b b
b
c
c a a
bc a
ab bc
a bc c
abc a a
HR-C-L HR-S-L HR-C-M HR-S-M HR-C-H HR_S_H Glutamine concentration (nmol g-1 FW)
0 100 200 300 400 500 600
Control Stress Control Stress
Low K High K
Control Stress Medium K a
b
b b b b
c c c bc
a ab
c ab
bc bc a
ab
a )
b )
Figure 14. Effect of drought stress and varied K supply on amino acid concentrations in flag leaves of barley. Concentrations of a) phenylalanine and b) glutamine were measured in flag leaves of LP104, LP110 and LP106 subjected to drought stress at 5 days after flowering. Plants were pre-cultured under sufficient water supply (control) or water limitation (stress) and under low, medium or high K supply. Flag leaves from 12 weeks-old plants were harvested 12 days after imposing drought stress. Bars indicate means ± SD. Different letters denote significant differences according to 3-way ANOVA and Tukey’s test within each K treatment (p<0.05; n=4-6).
5.1.8 Primary metabolite changes under drought stress and varied K supply
_________________________________________________________________ Results supply in the line LP104 compared to other two lines (Figure 15a). In addition, the concentrations of ADP-glucose increased under drought at low K supply in LP104 in comparison with LP110 and LP106 (Figure 15b). Although, the concentration of oxoglutarate decreased under drought in all three investigated lines, however, this reduction was significantly less in the line LP104 compared to other two lines (Figure 15c). An increase in concentration of oxoglutarate was observed under drought stress in both LP110 and LP106 by additional K into the substrate (Figure 15c).
The concentrations of 3-PGA increased in all 3 investigated lines under normal watering regimes. In LP104, the concentration of 3-PGA increased under drought when the K level was high in the substrate (Figure 16c). The concentrations of NADPH, ATP and UDP-glucose were relatively high in the both LP104 and LP110 in comparison with LP106 (Figure 16c, 17a and b). Notably, the concentration of fumarate was also significantly higher under drought in the line LP104 compared to the line LP106 either at low or high K supplies (Figure 16b). Trehalose-6-P level also was increased with high amount of K in the soil in all 3 lines; however, this positive effect was significantly more pronounced in LP106 compared to LP104 and LP110 in spite of different watering regimes (Figure 17c).
In summary, the levels of primary metabolites, in particular those of hexoses were less affected in LP104 and remained at a more constant level under concomitant drought and K deficiency than in the two other lines.
HR-C-L HR-S-L HR-C-M HR-S-M HR-C-H HR_S_H Hexose-6-P concentration (nmol g-1 FW)
0 20 40 60 80 100 120
Control Stress Control Stress
Low K High K
Control Stress Medium K a
a a
b b
b b
b b
b b
ab b
b ab
b a
b
HR-C-L HR-S-L HR-C-M HR-S-M HR-C-H HR_S_H ADP-glucose concentration (nmol g-1 FW)
0.00 0.05 0.10 0.15 0.20 0.25
Control Stress Control Stress
Low K High K
Control Stress Medium K bc
bc bc
a
c ab
a a
a a
a a
bc c c
c ab a
HR-C-L HR-S-L HR-C-M HR-S-M HR-C-H HR_S_H Oxuglutarate concentration (nmol g-1 FW)
0 100 200 300 400 500 600
Control Stress Control Stress
Low K High K
Control Stress Medium K a
b bc bc
a
ab ab
ab
ab bc b
abc
c c
cd d
b
b )
c ) a )
HR-C-L HR-S-L HR-C-M HR-S-M HR-C-H HR_S_H
RelativeHvS40 mRNA level
0 1 2 3 4 5
6 LP104
LP110 LP106
b b b a
b a
b b b a
a a a b
b b b
b
Control Stress Control Stress
Low K High K
Control Stress Medium K HR-C-L HR-S-L HR-C-M HR-S-M HR-C-H HR_S_H Hexose-P concentration (nmol g-1 FW)
0 20 40 60 80 100 120
HR-C-L HR-S-L HR-C-M HR-S-M HR-C-H HR_S_H Oxoglutarate concentration (nmol g-1 FW)
0 100 200 300 400 500 600
Figure 15. Effect of drought stress and varied K supply on primary metabolite concentrations in flag leaves of barley. Concentrations of a) hexose phosphate, b) ADP-Glucose and c) oxoglutarate were measured in flag leaves of LP104, LP110 and LP106 subjected to drought stress at 5 days after flowering. Plants were pre-cultured under sufficient water supply (control) or water limitation (stress) and under low, medium and high K supply. Flag leaves from 12 weeks-old plants were harvested 12 days after imposing drought stress. Bars indicate means ± SD.
Different letters denote significant differences according to 3-way ANOVA and Tukey’s test within each K treatment (p<0.05; n=4-6). Abbreviations are as follows: Hexose-P: hexose phosphate.
_________________________________________________________________ Results
HR-C-L HR-S-L HR-C-M HR-S-M HR-C-H HR_S_H 3PGA concentration (nmol g-1 FW)
0 100 200 300 400 500 600 700
Control Stress Control Stress
Low K High K
Control Stress Medium K a
b
c ab b
c a
bbc b
bc c
a
b b
b
c c
HR-C-L HR-S-L HR-C-M HR-S-M HR-C-H HR_S_H Fumarate concentration (nmol g-1 FW)
0 1000 2000 3000 4000 5000 6000
Control Stress Control Stress
Low K High K
Control Stress Medium K a
c bc ab
bc c
ab ab ab
a
b ab a
bc b bc
bc c
HR-C-L HR-S-L HR-C-M HR-S-M HR-C-H HR_S_H
NADPH concentration (nmol g-1 FW)
0 1 2 3 4 5 6
Control Stress Control Stress
Low K High K
Control Stress Medium K ab
b a
a
a
abc
b b
c abc
ab
bc a
b b
c bc
b
b )
c ) a )
HR-C-L HR-S-L HR-C-M HR-S-M HR-C-H HR_S_H
RelativeHvS40 mRNA level
0 1 2 3 4 5
6 LP104
LP110 LP106
b b b a
b a
b b b a
a a a b
b b b
b
Control Stress Control Stress
Low K High K
Control Stress Medium K
Figure 16. Effect of the drought stress and varied K supply on primary metabolite concentrations in flag leaves of barley. Concentrations of a) 3PGA, b) fumarate and c) NADPH were measured in flag leaves of LP104, LP110 and LP106 subjected to drought stress at 5 days after flowering. Plants were pre-cultured under sufficient water supply (control) or water limitation (stress) and under low, medium and high K supply. Flag leaves from 12 weeks-old plants that were harvested 12 days after imposing drought stress. Bars indicate means ± SD. Different letters denote significant differences according to 3-way ANOVA and Tukey’s test within each K treatment (p<0.05; n=4-6). Abbreviations are as follows: 3PGA: 3-phosphoglycerate, NADPH: Nicotinamide adenine dinucleotide phosphate.
HR-C-L HR-S-L HR-C-M HR-S-M HR-C-H HR_S_H ATP concentration (nmol g-1 FW)
0 5 10 15 20 25 30
Control Stress Control Stress
Low K High K
Control Stress Medium K ab
ab bc
bc a
c a
ab
cd bc
cd d
c ab ab a a
c
HR-C-L HR-S-L HR-C-M HR-S-M HR-C-H HR_S_H UDP-glucose concentration (nmol g-1 FW)
0 10 20 30 40 50 60
Control Stress Control Stress
Low K High K
Control Stress Medium K ab ab
a
ab
a ab a
ab ab ab
a
abc
bc bc c c b
b
HR-C-L HR-S-L HR-C-M HR-S-M HR-C-H HR_S_H Trehalose-6-P concentration (nmol g-1 FW)
0.0 0.5 1.0 1.5 2.0 2.5
Control Stress Control Stress
Low K High K
Control Stress Medium K abc
a ab
c c bc
c
c ab
bc c
a
c c
b b
c a
b )
c ) a )
HR-C-L HR-S-L HR-C-M HR-S-M HR-C-H HR_S_H
RelativeHvS40 mRNA level
0 1 2 3 4 5
6 LP104
LP110 LP106
b b b a
b a
b b b a
a a a b
b b b
b
Control Stress Control Stress
Low K High K
Control Stress Medium K
Figure 17. Effect of drought stress and varied K supply on primary metabolite concentrations in flag leaves of barley. Concentrations of a) ATP, b) UDP-glucose and c) trehalose-6-P were measured in flag leaves of LP104, LP110 and LP106 subjected to drought stress at 5 days after flowering. Plants were pre-cultured under sufficient water supply (control) or water limitation (stress) and under low, medium and high K supply. Flag leaves from 12 weeks-old plants that were harvested 12 days after imposing drought stress. Bars indicate means ± SD.
Different letters denote significant differences according to 3-way ANOVA and Tukey’s test within each K treatment (p<0.05; n=4-6). Abbreviations are as follows: Trehalose-6-p: trehalose-6-phosphate, ATP: Adenosine triphosphate.
_________________________________________________________________ Results
5.2 Influence of K supply on the hormonal regulation of two barley lines with different behaviour under drought stress
Based on the higher carbohydrate and starch levels in LP104, the question arose whether the different behaviour of the investigated barley under drought stress may be related to the stress hormone ABA. Previously, a cross talk has been postulated between starch metabolism in barley leaves and the ABA signaling pathway (Fujii et al., 2009; Seiler et al., 2011). To investigate ABA-related responses in the two most contrasting lines LP104 and LP106, plants were grown in a pot experiment in the greenhouse for a period of 12 weeks and fertilized with an adequate (high K treatment, 4g/kg dry soil of K2SO4) or deficient (low K treatment, without adding K2SO4) amount of K. Five days after flowering (DAF) drought stress was imposed by reducing the soil moisture content to 10%
(corresponding to 25% field capacity). After 12 days of drought stress flag leaves were harvested for the assessment of physiological and biochemical parameters induced by drought stress.
5.2.1 Chlorophyll concentrations in flag leaves under drought stress and