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Poplar cambium and leaf tissues present specific changes in 2D-page patterns in response to drought of heat stress
Thomas Durand, Domenico Morabito, Sébastien Planchon, Kjell Sergeant, Sabine Carpin, Philippe Label, Jean-François Hausman, Jenny Renault
To cite this version:
Thomas Durand, Domenico Morabito, Sébastien Planchon, Kjell Sergeant, Sabine Carpin, et al..
Poplar cambium and leaf tissues present specific changes in 2D-page patterns in response to drought
of heat stress. 2. Poplar Symposium, Mar 2009, Göttingen, Germany. 1 p., 2009. �hal-02824773�
T h is w o rk i s s u p p o rt e d b y a g ra n t fr o m t h e L u xe m b o u rg F o n d s N a ti o n a l d e l a R e c h e rc h e (A F R 0 5 / 0 9 4 ).
a : Laboratoire de Biologie des Ligneux et des Grandes Cultures. UFR – Faculté des Sciences.
Université d’Orléans. UPRES EA-1207. rue de Chartres. B.P. 6759. 45067 Orléans CEDEX 2.
France.
b : CRP-Gabriel Lippmann. 41 rue du Brill. L-4422 Belvaux. GD Luxembourg.
c : Institut National de la Recherche Agronomique. 2163 avenue de la Pomme de Pin. B.P.
20619 Ardon. 45166 Olivet CEDEX. France.
d : INRA USC2030 'Arbres et Réponses aux Contraintes Hydriques et Environnementales' (ARCHE) F-45067 Orléans cedex.
POPLAR CAMBIUM AND LEAF TISSUES PRESENT SPECIFIC CHANGES IN 2D-PAGE PATTERNS IN RESPONSE TO DROUGHT OR HEAT STRESS
Poplar Symposium
Göttingen, march 11-13 2009
Durand, Thomas C. abcd ; Morabito, Domenico ad ; Planchon, Sébastien b ; Sergeant, Kjell b ; Carpin, Sabine ad ; Label, Philippe c ; Hausman, Jean-François b ; Renaut, Jenny b
Introduction.
Plant stress tolerance is determined by the ability to preserve tissue- and cell- functions. The mechanisms involved in the metabolic defence partly depend on the physiological status, but also on stress sensing and signalling pathways. Accordingly, stress tolerance arises through molecular responses, depending on cell and tissue specific structures.
Thus specific proteomic changes can be expected in leaves and in cambium of Populus tremula x P. alba genotype 717-1B4 exposed to stress.
tdurand@lippmann.lu
Conclusion.
Each kind of abiotic stress endured by the plants triggers partly specific changes in the studied proteomes (fig 2&3). In leaf, 20 out of the 38 identified protein functions present a stress-specificity. They are 44 out of 72 in the cambial tissue. In spite of that relative stress specificity, the same metabolisms are involved in the responses.
When comparing proteome changes between leaf and cambial zone, only 6 protein functions, among the 104 identified, were common between tissues in the responses to the different stresses applied.
The tissue specificity of the response exceeds the stress specificity.
Proteomic analysis
Protein extracts were separated by 2D-DIGE. From 4 biological replicates of each sample, a relative quantification was performed, using DeCyder software.
Leaf tissue analysis showed 169 proteins spots which abundance was significantly (p<0.05) altered under the constraints presented in Figure 1, with an absolute value of the average ratio of at least 1.3.
MALDI TOF-TOF analysis, still in progress, already allowed the confident identification of 102 proteins.
Cambial zone analysis showed 197 protein spots with differential abundance, among which 155 were confidently identified.
Stress characterisation
Young pot-grown poplar plants were submitted to drought or heat stress (with or without a previous temperature gradient) while physiological parameters were monitored (fig. 1). A 7 days return to control conditions period was performed, allowing plants to recover. Tissues were sampled at key steps of the experimental set-up, indicated by ( ) in the figure 1.
Drought decreased H
2O stomatal conductance, CO
2assimilation, predawn leaf water potential and leaf relative water content.
Heat treatments triggered a shorter stress episode, which affected leaf water potential and CO
2assimilation.
The plants experienced characterised and reversible stresses.
Fig 1: Experimental set of drought and heat stress applied to Populus tremula x P. alba717 - 1B4 : sampling of tissue
42°C
22°C 22°C
D12 D19
D1
Drought stress : 12 days of withholding water
Heat Gradient
Heat Shock
25°C 30°C
42°C
22°C 22°C
D19 D12
D1
7 days of recovery
D1 D8 D12 D19
Re-watering
Fig 3.Bidimensional electophoretic separation of proteins isolated from poplar stem cambial tissue.
Underlined spots, differential in abundance between control and stressed plants, were confidently identified.
Proteome changes in Leaf and Cambial zone tissues
Fig 2.Bidimensional electophoretic separation of proteins from poplar leaves.
The underlined spots were quantitatively different between control and stressed plants.
The abundance of only 6 proteins (present in 43 protein spots in total) were altered in both leaf and cambial proteomes, for instance glyceraldehyde-3-phosphate dehydrogenase ( ) fructose-bisphosphate aldolase ( ), and thioredoxin peroxidase ( ). However, the changes observed in these protein abundances in response to the applied constraints were always dissimilar in the two tissues.
The sole exception was one Quinone oxidoreductase ( ) (cf fig 2 & 3).
In Cambial zone tissue:
➢Heat Gradienthad a particularly strong impact on glycolysis, and among stress-responsive proteins, it affected 9 isoforms of Osmotin( ).
➢Heat Shock impacted almost specificalySAM Cycle( ) & One- carbon Metabolism(especially serine hydroxymethyltransferase ),
➢Water deficitinduced an important redox response (GPX, APX, PRX2 ) and a massive decrease of storage proteins ( ).
These reponses are particularly important on day 12, corresponding to the strongest intensity of the constraint.
Specific features appeared between treatments.
In leaf tissue:
➢Heat Gradienthad an important inhibitory effect on the Calvin cycle aldolases ( & ).
➢Heat Shock induced an increase of 5 Oxygen- evolving enhancer protein isoforms ( ).
➢Water deficit particularly affected RuBisCO ( ), with an increased abundance of 5 isoforms at the recovery stage.
Fig 4.Patterns of the proteomic changes observed in various metabolisms in leaf and cambial zone tissues
Electrofocusing SDS-PAGE
Protein separation
Image capture Bioinformatics (DeCyder)
Spot picking
& digestion
MALDI-TOF-TOF identification
pI=3 Non-linear pI gradient 10
Increasing molecular weight
pI=3 Non-linear pI gradient 10
Increasing molecular weight
Carbon metabolism Starch and sucrose metabolism Glycolysis
Respiration One-carbon Pathway SAM cycle
Nitrogen & Protein Metabolism Protein folding
Redox status and Stress response Transcription & Translation Factors Signal transduction Cell Structure Secondary metabolism Miscellaneous Nbof differential isoforms Underexpressedoverexpressed Cambial zone
Heat Gradient Heat Shock Drought
15 10 5 0 5 10 15 20 Carbon metabolism
Photosynthesis & electron transport Respiration
Nitrogen & amino acid metabolism Glycolysis
Redox status & Stress response Miscellaneous
Heat
Gradient Heat Shock Drought Leaf tissue
Nbof differential isoforms Underexpressedoverexpressed 15 10 5 0 5 10 15 20
Constraints Constraints
