Plant Biotech Denmark Meeting
February 2, 2012
Breeding – quality and productivity/
Synthetic - and systems biology
Implications of high-temperature
events and water deficits on
protein profiles in wheat (Triticum
aestivum L. cv. Vinjett) grain
Susanne Jacobsen
Enzyme and Protein Chemistry, Department of Systems Biology,
Technical University of Denmark
Influence of high-temperature events and water deficits
on protein profiles in wheat (Triticum aestivum L. cv.
Vinjett) grain
• Climate change is likely to become an even
more acute problem than previously
anticipated
• Accelerated climatic variability is resulting in
an increase of both the frequency and the
magnitude of extreme events
• More frequent extreme climatic episodes result
in cereals being exposed to several stress events
during the growing season, ultimately affecting
cereal production
Influence of high-temperature events and water deficits
on protein profiles in wheat (Triticum aestivum L. cv.
Vinjett) grain
When?
• The most sensitive growth stages in cereals are spikelet
formation (double ridges to terminal spikelet) and
anthesis
• Extreme heat episodes after anthesis increase
development of the crops, shorten the grain filling
period, and may result in reduction of kernel weight and
of grain quality parameters (starch and protein)
Influence of high-temperature events and water deficits
on protein profiles in wheat (Triticum aestivum L. cv.
Vinjett) grain
• Moderately high temperatures after cold
winther periods increase yields
• An extreme heat event at the ´double-ridge´ stage did
not affect the response of wheat to heat stress at anthesis
• Drought and increased heavy rainfalls
result in yield losses
Influence of high-temperature events and water deficits
on protein profiles in wheat (Triticum aestivum L. cv.
Vinjett) grain
Why?
• 2/3 worldwide human calorie intake is
dependent on cereal production
– Direct intake (e.g. rice)
– Processed products (e.g. bread, pasta, beer...)
– Animal feed (milk, poultry and meat production)
• New biotechnological applications of plants
– Biofuels
– Molecular farming (using plants to produce therapeutic
proteins)
Wheat grain quality receives increasing
attention because of its economic and
nutritional importance
• Studies on responses of wheat grain proteins
to long-term drought or high temperature
• However little is known about the effect of
the combination of water deficits and high
temperature events on grain proteins
• Wheat grain for end use quality: protein fractions
albumins, globulins, gliadins and glutenins
The complexity of the plant response to
abiotic stress
Single and multiple stress factors
Single stress factors
(controlled (!) environmental studies)
versus
Multiple stress factors (nature) for example the
contrasting implications of high temperatures
and water deficits)
- Conclusion: More information is needed on the
interaction of multiple stress events
and the consequences for crop quality
Interaction between multiple drought events
Triticum aestivum L. cv. Vinjett - semi-field experiment subjected to
drought episodes either at terminal spikelet, during grain-filling or
at both stages – effect on total protein and protein fractions
Control:
Watering
throughout
Early drought:
TD
(double ridge)
Late drought:
TA
(anthesis)
TD + TA
(double ridge+
anthesis)
Winning, H. et al., J. Exp. Bot. (2009)
After anthesis
Wheat growth stages
Five-leaf stage
Crop Monitoring and Zadoks Growth Stages for Wheat
High-temperature treatments
temperature in growth chambers was set to increase to 25oC at
the first stage and to 32oC at the second stage
Water suply was withheld
for 7 days at 5-leaf stage
before the first stage
and for 6 days before the
second stage
2007
Influence of high-temperature events and water deficits
on protein profiles in wheat (Triticum aestivum L. cv.
Vinjett) grain
25
A
-1
Albumins[g(mggrain) ]
25
-1
Globulins[g(mggrain) ]
Albumin
Globulin
cA-cB
cA-wdB
cA-htB
htA-htB
20
15
20
15
10
5
5
B
3,0
F
3,0
2,5
2,5
2,0
2,0
1,5
1,5
1,0
1,0
C
-1
Gliadins[g(mggrain) ]
-1
Glutenins[g(mggrain) ]
Glutenin
E
10
30
Gliadin
wdA-cB
wdA-wdB
wdA-htB
htA-cB
htA-wdB
G
30
25
25
20
20
15
15
10
10
D
50
H
50
40
40
30
30
20
20
20
30
40
50
dpa
dpa
20
30
40
50
Influence of high-temperature events and water deficits
on protein profiles in wheat (Triticum aestivum L. cv.
Vinjett) grain
- At matury highest contents of albumins, globulins
and gliadins after single drought treatment
- In contrast to albumin and gliadin fractions, multiple hightemperature events or water deficits applied at both
terminal spikelet and anthesis resulted in significantly
higher contents of glutenins at maturity than in the single
stress treatment
- Combination of drought and heat stress applied at two stages did
not have a greater effect on albumin, globulin and gliadin
contents during grain filling compared with either of the stresses
applied individually
Interaction of multiple stress events
Proteome analysis - Albumin
cA-cB
htA-htB
cA-htB
cA-wdB
2-DE gels of albumin
fraction from wheat
Identified stress responsive
proteins included proteins
involved in primary metabolism,
storage and stress response such
as late embryogenesis abundant
proteins, peroxiredoxins, αamylase / trypsin inhibitors and
heat shock proteins
Yang et al., Proteomics
(2011) 11: 1684-1695
Interaction of multiple stress events
Proteome analysis - Gliadin
kDa 4 pI 10 kDa 4 pI 10
97
97
2-DE gels of gliadin fraction
from wheat
c4
55
55
37
37
31
21
21
14
cA‐cB 14
kDa
cA‐wdB
d11 kDa
d2
97
97
d4
55
d1 d6 d9
37
31
c3
31
c1
d8
c5
c2
d10
d7
d5
55
f2
f1
f3
f5
f6
f4
f7
37
31
f8
d3
21
14
21
cA‐htB 14
htA‐htB
Identified stress responsive
proteins included
α-gliadins
γ- gliadins
α/β-gliadin
ω -gliadin
Influence of high-temperature events and water deficits on
protein profiles in wheat (Triticum aestivum L. cv.
Vinjett) grain
cA-wdB treatment
Increased
Monomeric α-amylase inhibitor
Late embryogenesis abundant protein H2
Cytosolic 3-phosphoglycerate kinase
Phytochrome B
Embryo-specific protein
Decreased
1-Cys peroxiredoxin
α-amylase/trypsin inhibitor CM3
Glucose and ribitol dehydrogenase
Small Ras-related GTP-binding protein
Calmodulin TaCaM2-2
cA-htB treatment
htA-htB treatment
Increased
1-Cys peroxiredoxin PER1
27K protein, pfam03227,
lysosomal thiol reductase
Serpin 2
α-amylase/trypsin inhibitor CM17
Decreased
Lipoxygenase
cd02947, thioredoxin family
Succinyl-CoA ligase β-chain
Triosephosphate isomerase
Glutamine synthetase
Increased
α-amylase inhibitor CM17
Monomeric α-amylase inhibitor
0.19 dimeric α-amylase inhibitor
Fructose-6-phosphate-2-kinase
Enolase 2
β-amylase
Decreased
27K protein, pfam03227, γ-interferon inducible lysosomal
thiol reductase
α-amylase inhibitor CM1
Multidomain cystatin
14-3-3 protein
Influence of high-temperature events and water deficits
on protein profiles in wheat (Triticum aestivum L. cv.
Vinjett) grain
Conclusion -I
• Combination of drought and heat stress applied at two
stages of grain filling did not have a significant effect on
albumin contents compared with either of the stresses
applied individually
• Globulins and gliadins showed with exception of htAcB increased protein fraction contents from 19 dpa
• Glutenins accumulated during grain filling in response
to different treatments
Influence of high-temperature events and water deficits
on protein profiles in wheat (Triticum aestivum L. cv.
Vinjett) grain
Conclusion -II
• Some stress treatments can significantly modify
contents of individual proteins in grain
• Changes in grain protein contents strongly depended
on the type of stress applied
• Identified proteins are responsive to drought and
high-temperature events
• Proteins play role in anti-desiccation, antioxidation,
defence, carbohydrate metabolism and storage
The complexity of the plant response to
abiotic stress
Influence of high-temperature events and water deficits
on protein profiles in wheat (Triticum aestivum L. cv.
Vinjett) grain
Conclusion and perspectives
• The protein markers identified
are relevant for plant breeding
• Present findings indicating effects of more than one
stress event and type on grain protein composition will
contribute to the identification of cultivars with
increased tolerance to increasing climate variability
Acknowledgements
Aarhus University:
Bernd Wollenweber, technical assistance: Betina Hansen,
Jesper Hjort D. Petersen, Palle Kristiansen Ahm, Ana Andersen,
Mir Aigne
Nanjing Agricultural University:
Dong Jiang, Xia Zhang, Huawei Li, Xiao Wang
Enzyme and Protein Chemistry, Department of Systems Biology,
Technical University of Denmark:
Ljiljana Nesic, Anne Blicher, Birgit Andersen, Rune Halvorsen,
Morten S. Mortensen, Anders Dysted Jørgensen, Fen Yang, Morten Ejby,
Ib Søndergaard, Christine Finnie, Birte Svensson
Funding: Danish Council for Technology and Production Sciences (23-04-0076) to Bernd W.,
Danida (Xia Z., Huawei L.), Chinese Scholarship Council (Dong J., Xiao W.), Cirius (Dong
J.), Directorate for Food, Fisheries and Agri Business grant #3304 FVFP 060678, Centre for
Advanced Food Studies (Fen Y.)
Influence of high-temperature events and water deficits on
protein profiles in wheat (Triticum aestivum L. cv.
Vinjett) grain
Thank you for your attention
Annual Plant Biotech Denmark Meeting
February 2, 2012