1. Indian Agricultural Research Institute, New Delhi - 12
Influence of high [CO2] and sulphur levels on root membrane stability and organic acid exudation under low phosphorus in wheat
Milan Kumar Lal* and Renu Pandey
Mineral Nutrition Lab
Division of Plant Physiology
Indian Agricultural Research Institute
New Delhi – , India *
Presented at: National Seminar on Plant Genomics and Biotechnology, OUAT, BBSR

2. Availability of Phosphate rock by the end of this century
Indian Agricultural Research Institute, New Delhi - 12
Availability of Phosphate rock by the end of this century
Mining of Phosphate rock – A non-renewable resource
Syers et al., 2011

3. INTRODUCTION Indian Agricultural Research Institute, New Delhi - 12
Phosphorus is vital to plant growth and is found in every living plant cell, involved in several key plant functions;
Energy transfer
Photosynthesis
Structural component of nucleic acid and membranes
Mostly taken up as the primary orthophosphate ion (H2PO4-) at (pH < 7.0), but also absorbed as secondary orthophosphate (HPO42 -) at (pH > 8.0).
It is incorporated into organic compounds, including nucleic acids (DNA and RNA), phosphoproteins, phospholipids, sugar phosphates, enzymes, and energy-rich phosphate compounds, adenosine triphosphate (ATP) .
Activation of enzymes and enhanced production of phosphatases RNases and organic acids, changes in protein phosphorylation
Organic acid exudates under low P help to mobilize the fixed phosphorus from the soil to plant
Indian Agricultural Research Institute, New Delhi - 12

4. Objectives of Research
Indian Agricultural Research Institute, New Delhi - 12
Objectives of Research
To study the interactive effect of high [CO2] and phosphorus starvation on growth with different sulphur level
To analyze the interactive effect of high [CO2] and phosphorus on membrane injury and root exudation capacity in wheat grown with different sulphur levels
To analyze the expression of candidate genes involved in phosphorus remobilization from membranes in wheat grown under high [CO2] and phosphorus starvation with different sulphur levels
Hypotheses:
Elevated CO2 would provide extra carbon to maintain the flux and thereby improving the organic acid exudation under low P
Since Pi is replaced with S in membrane lipids, the elevated CO2 might also influence membrane permeability and uptake of nutrient.

5. Materials and Methods
Indian Agricultural Research Institute, New Delhi - 12
Crop : Wheat Varieties : a. PDW 233 b. PBW 396 Treatments : P level (sufficient 500 µM and deficient 5 µM) S level (sufficient 2000 µM and deficient 10 µM) Carbon dioxide- Ambient Elevated 700 ppm Replications : Three Design of experiment: Factorial CRD

6. Results- Biomass accumulation
Indian Agricultural Research Institute, New Delhi - 12
Results- Biomass accumulation
0.533
0.422
0.257
0.284
0.062
0.151
0.121
0.075
0.614
0.257
0.296
0.543
0.294
0.265
0.319
0.399
Shoot dry weight averaged over nutrient interaction is increased by 64 and 88% in PDW-233 and PBW-396 respectively
Root dry weight also doubled under eCO2 condition
The total biomass showed 70% more accumulation in PBW 396 under eCO2
The R/S ratio averaged over CO2 X variety increased under low P (41%) and low P/S (12%)

7. PBW 396 PDW 233 Control -P -S -P/-S Elevated CO2 Ambient CO2
Indian Agricultural Research Institute, New Delhi - 12
PBW 396
PDW 233
Control -P -S
-P/-S
Elevated CO2
Ambient CO2

8. Results- Root architecture
Indian Agricultural Research Institute, New Delhi - 12
Results- Root architecture
154.3
325.9
183.1
291.4
17.39
29.92
19.78
32.2
0.16
0.24
0.17
0.28
106.6
102.8
65.7
86.5
The root length increased under eCO2 condition in both varieties and under –P and –P/-S
Root surface area was more than 60% at eCO2 in both varieties averaged over nutrient treatment as compared to aCO2
Low P and low P/S has more root volume and low S perform better under eCO2
No. of lateral increased under eCO2 and also increased in –P/S in PDW 233 under eCO2

9. Root system of PDW 233 seedlings under nutrient treatments
Control -P
-P/-S -S
Ambient CO2
Elevated CO2
Indian Agricultural Research Institute, New Delhi - 12
Root system of PBW 396 seedlings under nutrient treatments

10. Root hairs of PBW 396 seedlings under nutrient treatments
Root hairs of PDW 233 seedlings under nutrient treatments
Indian Agricultural Research Institute, New Delhi - 12
Control -P -S
-P/-S
Ambient CO2
Elevated CO2

11. Membrane Injury of root
Indian Agricultural Research Institute, New Delhi - 12
Membrane Injury of root
53.18
52.78
51.14
54.43
0.10
0.29
0.13
0.12
Membrane injury found to higher in the aCO2 condition compared to eCO2
Overall MII increased by 25 and 27% at low P and low P/S, respectively in comparison to control.
The membrane damage of root was significantly less at low P and low S under eCO2
MGDG content was increased under eCO2 condition in PDW 233 and DGDG content was increased under low P in both eCO2 as well as aCO2
SQDG content decreased under –S and increased under –P and –P/S both in elevated and ambient CO2
The amount of PG under aCO2 was reduced by 58 mol% while the reduction in MGDG was marginal at low P over control

12. Carboxylate exudation
Indian Agricultural Research Institute, New Delhi - 12
Carboxylate exudation
Sl. No.
Organic acid
CO2 level
PDW 233
PBW 396
Control
Low P
Low S
Low P/S 1.
Oxalic acid AC Nd EC
6.08±0.46
6.71±0.17
4.71±0.28
5.34±0.58
7.86±0.64
9.91±0.36
6.64±0.25
8.47±0.44 2.
Citric Acid
2.43±0.15
2.33±.0.07
2.59±0.10
5.80±0.12
2.15±0.09
1.86±0.14
1.74±0.04
2.34±.0.089
3.57±0.16
2.88±0.12
0.81±0.13
3.10±0.32
4.53±0.12
4.09±0.20
2.67±0.18
7.76±0.18 3.
Pyruvic acid
5.20±0.42
7.95±0.75
3.60±0.18
6.40±0.15
9.07±0.56
3.34±0.33
3.47±0.17
7.37±0.51
6.02±0.39
9.76±0.43
5.77±0.59
7.29±0.72
7.24±0.27
14.89±0.36
11.09±0.67
10.99±0.52 4.
Malic acid
1.31±0.17
1.25±0.21
0.06±0.00
1.14±0.05
1.46±0.20
1.65±0.10
0.51±0.10
1.39±0.31
2.05±0.09
1.82±0.05
0.97±0.02
2.20±0.37
1.07±0.14
3.08±0.10
0.70±0.08
4.88±0.34 5.
Succinic acid
3.42±0.06
2.74±0.35
0.017±0.00
2.15±0.18
2.57±0.05
1.53±0.08
1.73±0.03
4.25±0.79
7.59±0.59
8.51±0.35
2.53±0.41
3.86±0.60
4.07±0.42
13.44±0.54
3.86±0.56
13.21±1.00 6.
Lactic acid
2.21±0.35
4.30±0.67
3.88±0.25
5.66±1.40
3.89±0.51
6.69±0.31
5.21±0.66
8.39±1.48
3.18±0.15
5.58±0.31
4.05±0.27
7.44±0.26
4.72±0.76
8.23±0.30
6.81±0.64
10.25±0.91 7.
Fumaric acid
3.91±0.40
2.38±0.29
2.12±0.09
5.37±0.91
2.66±0.66
5.61±1.39
5.64±0.94
6.98±0.11
4.21±0.20
4.62±0.30
0.63±0.04
4.90±0.50
2.08±0.09
6.00±0.41
2.88±0.66
8.94±0.27 8.
Total Organic acid
18.41±0.69
20.98±2.02
12.29± 0.28
26.55±1.22
21.82±0.731
20.94±1.41
18.32±0.46
30.43±2.68
32.72±1.11
39.91±1.36
19.49±0.63
34.16±2.25
37.77±2.26
59.68±2.04
34.50±2.60
64.52±3.47
In aCO2 no Oxalic acid was detected but in eCO2 there was significant exudation
The low P/S treatment results in 50% increased in citric acid while other low P and low S showed over control
Among varieties, malic acid increased by 87% in PDW-233 and by 2.4-fold in PBW-396 under eCO2
In low P condition have more succinic exudates under eCO2 and similar trend was seen in lactic, fumaric acids.
The total carboxylic acid concentration in root exudates doubled in response to eCO2 over aCO2, with more exudation in PBW 396 as compared to PDW 233

13. Enzymes activity and Nutrient accumulation
Indian Agricultural Research Institute, New Delhi - 12
The PEPC activity under low P resulted in dramatic increase by 141%
For MDH eCO2 in both varieties have more enzyme activity as compared to its control
CS has drastic increase in PBW 396 variety under eCO2 , Increase also seen in –P and –S
In response to eCO2, the activity of PK in root averaged over nutrient treatment and varieties increased by 63% over aCO2
In –S under eCO2 condition the total S uptake was increase this show more S uptake under eCO2
The total P accumulation was enhanced in both varieties under eCO2

14. Conclusion Indian Agricultural Research Institute, New Delhi - 12
The total biomass increases under eCO2 in both varieties as compared to control implying that high CO2 provides more carbon that is accumulated in biomass
The root surface area, length, volume and laterals increased under –P, -S and –P/-S indicating that root architecture is altered under nutrient deprived condition
Membrane injury was less under eCO2 compared to aCO2 indicating extra CO2 provides membrane stability and sulphur provide membrane stability in form of sulpholipid
Carboxylates exudation is higher under eCO2 and also under nutrient deficient condition, eCO2 provides more carbon flux that help in more exudation
Under P stress and eCO2 galactolipid like SQDG, DGDG increased suggesting that the phospholipid was replaced with galactolipids thus preventing membrane damage
The relative expression of TaPT1 gene and other membrane lipid scavenging genes increased drastically under eCO2 indicating higher synthesis of galactolipids involved in membrane lipid recycling.

15. Thank You Acknowledgement:
Chairperson and member of Mineral Nutrition Lab
ICAR for financial support in form of JRF
National Seminar of Plant Genomics and Biotechnology-2015