Carbon balance and elevated temperature

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Plant material and growing conditions

Five experiments were conducted on two-year-old own-rooted potted Microvines in 2011 (Exp. 1, 2) and in 2013 (Exp. 6, 7 and 8) in growth rooms at Montpellier SupAgro-INRA campus (see general materials and methods, Table 1). Plants were initially grown, up to 30 to 55 unfolded leaves, under control climatic conditions in greenhouse (i.e. day/night temperatures of 25°C/15°C; VPD of 1.28 kPa and daily PAR over a 14-hours photoperiod of 19.2 mol.m-2.d-1). Then, contrasted temperatures treatments were applied in growth rooms of 1m² (Exp. 1, 2, 6 & 7) or 20m² (Exp. 8). Temperature flowering in order to retain dropped flowers and berries due to fruit set failure.
Health, USA). Red line represent the 1 cm scale measured on the image, purple line represent the measured berry diameter. treatments lasted 2-months for Exp. 1 and 1-month for Exp. 2, 6, 7 & 8. Day/night temperatures were set to reach 22°C/12°C and 30°C/20°C, respectively for Exp. 1 and 2. For Exp. 6, 7 and 8, they were set to reach 30/15°C, 30/25°C and 25/15°C, respectively. Other climatic variables were similar for all experiments in 2011 and 2013. Average daily VPD ranged from 0.7 kPa for Exp. 1 to 0.96 kPa for control consisted in removing clusters beyond mid-flowering at T0 for Exp. 6 to 8 or beyond the lag phase at T0 for Exp. 1 & 2, thus corresponding to the ‘All. Inflo’ treatment in general materials and methods Table 1.

Development and growth measurements at the plant level

The date of budburst (modified E.L. stage 4; Coombe, 1995), was assessed from buds observations twice a week on each plant for all Exp. (see general materials and methods, Table 2). Then, the number of unfolded phytomers was counted twice a week from the beginning (T0) to the end (Tf) of the experiments in growth chambers. A phytomer was considered unfolded when the two foliar lobes separated by the main vein were in the same phyllotaxic plan. The phyllochron was calculated for each plant from these measurements, as detailed in chapter 1.
The total plant leaf area and the total flower and berry numbers were determined at T0 and Tf in all Exp. These variables were obtained from the sum of individual leaf area and from flowers or berries numbers at each phytomer position (see below). The last PI lignified was noted at Tf (LLPI). The leaf to fruit ratio (L:F), i.e. the ratio between total plant leaf area (m²) and the total flower and berries fresh weight (kg), and the plant root to shoot dry weights were determined at Tf in Exp. 6 to 8 R:S). For this purpose, total berry fresh weight and total shoot dry weight were calculated from the sum of all berries weight or all above organs weights (leaves, internodes, rachis, flowers and berries) at each phytomer level (see below). The entire root systems were cleaned and oven-dried at 60°C for 7 days (Fig. 1). Total root dry weight was then measured (R0_MS). Dry root systems were then ground with a Retsch Ultra Centrifugal Mill ZM200 (Retsch, Dusseldorf, Germany) and stored in a dry compartment for further carbohydrate assays.

Development and growth measurements at phytomer levels

Spatial development and growth along the proleptic axis were described both at T0 and at Tf in all experiments (Exp. 1 to 2 & 6 to 8; see general materials and methods, Table 2). The presence of any cluster at each P1 and P2 phytomer position (P0 phytomer bearing no inflorescence) was recorded.
Flower buds and berry numbers per cluster were counted from non-destructive observations on attached cluster at T0. Destructive counting of flower and berries were performed on non-attached clusters at Tf. In Exp. 6, nets were used to collect and count the aborted inflorescences, flower buds, flowers and berries over the one-month period in growth chamber (Fig. 2). Nets were positioned on Table 1: PCA variables description. Highlighted variables are related to inflorescence abortion.

Biochemical analyses

Berry glucose and fructose concentrations for Exp. 1 & 2 were assayed by HPLC (Rienth et al., 2014). Leaf, internodes, rachis and roots for Exp. 1 to 8 were assayed for glucose, fructose, sucrose and starch concentrations using enzymatic assays as described by Gibon et al. 2009. Total soluble sugars (TSS) concentration was calculated as the sum of glucose, fructose and sucrose concentrations. Total non- structural carbohydrates (TNC) concentration was calculated as the sum of TSS and starch concentrations. Total soluble sugars and starch contents per organ were calculated from organ fresh weights or dry weight.

Plants were different at the onset of the experiments in 2011 and 2013

Considering the differences observed between 2011 and 2013 experiments for a large spectrum of the responses to temperature, we suspected that differences in plant vigor between the years could be involved. Total leaf area were different among plants and experiments at T0 (the beginning of the experiment, Fig. 16A, Table 2), although plant leaf area was manually homogenized 2 days before each experiment. The number of phytomers, that depended on the time elapsed between budbreak and the beginning of the experiment, was different between years (58 in 2011 and 32 in 2013, Fig. 16A, Table 2). Moreover, more clusters at older PI were left resulting in more flowers and more berries present at the beginning of the thermal treatment in 2011 as compared to 2013 (500 flowers and berries vs. 50 flowers and berries, Fig. 16B). Furthermore, crop load through cluster removal was differentially managed between 2011 and 2013. In 2011, the last cluster kept was at the lag phase, just phytomers, corresponding to 13 clusters or almost 390 berries. In addition, in 2013, some plants presented hardly any quantifiable flowers or berries and showed aborted inflorescences below PI 6, the rank where floral buds are big enough to be counted, enabling any quantification of fruitfullness. As a result of plant height and vigor differences, the total internode volume was dramatically different, 20 dm3 and 8 dm3 in 2011 and 2013 respectively.

Elevated temperatures fasten and increase leaf and internode expansion, but not berry growth

The consistent increase of leaf expansion rate, and internode elongation rate and maximum size as a function of temperature was unexpected since the impact of temperature on development was taken into account by representing variables as a function of PI. Indeed, it has been regularly reported that leaf area and internode length increase as a function of thermal time follows essentially invariant patterns both in annuals (Turc and Lecoeur, 1997; Granier and Tardieu, 1998) and in perennials, at least those with indeterminate growth (Lebon et al., 2004b; Dambreville et al., 2013). In the case of leaf, this occurred in the absence of effect on the duration of expansion, suggesting a consistent increase of leaf expansion rate (on a thermal time basis) in response to temperature, again in contradiction with several studies. One possibility is that these responses are the signature of the shade avoidance syndrome. Indeed, it has recently been proposed that elevated temperatures lead to similar phenotypes as the so-called shade avoidance syndrome (de Wit et al., 2014) and that this could contribute to reduce leaf temperature and prevent damage to the leaf metabolic apparatus (Franklin, 2008; Crawford et al., 2012). This syndrome leads to increases in specific leaf area and internode length as observed in our experiments. Moreover, architectural changes induced by elevated temperature and shade avoidances share common signaling pathway, both mediated by PIF transcription factors that regulate auxin biosynthesis (de Wit et al., 2014; Franklin et al., 2014).

Table of contents :

General introduction
I. Grapevine and climate change
I.1. Grapevine economic trends
I.2. Climate change
I.3. First observations and expectations of climate change impact on Grapevine
II. Yield and elevated temperature
II.1. Yield definition in grapevine
(i) Grapevine reproductive development description
(ii) Yield components
(iii) Quality aside
II.2. Elevated temperature effect on crop yield
II.3. Elevated temperature effect on grapevine yield
(i) During the first year
(ii) During the second year: budburst
(iii) During the second year: flowering and fruit set
(iv) During the second year: first phase of berry growth
(v) During the second year: berry maturing
III. Carbon balance and elevated temperature
III.1. Elevated temperature effect on carbon acquisition
(i) Temperature effect on Photosynthesis
(ii) Temperature acclimation
III.2. Elevated temperature effect on carbon use
(i) Temperature effect on Respiration
(ii) Temperature effect on Photo-Respiration
(iii) Temperature effect on organ production
(iv) Temperature effect on organ expansion
(v) Temperature effect on biomass accumulation
(vi) Temperature effect on reserves dynamics
III.3. Elevated temperature effect on carbon balance
(i) Carbon balance definition
(ii) Existing models?
(iii) Temperature effect on Carbon Balance
IV. Working hypothesis: temperature effects on fruit set are related to carbon balance and dynamics?
V. The Microvine: a new model to address scientific issues on the grapevine response to temperature?
V.1. Wild type grapevine limitations
V.2. Creation and genetic description of Microvine
V.3. Phenotype description
V.4. Gibberellic acids in grapevine
(i) Synthesis and signalization pathway
(ii) Implication in development
Summary
(iii) Implication in response to biotic and abiotic stresses
(iv) What about Microvine?
V.5. Other dwarf model plants
VI. Working plan
General material and method
I. Plant material
II. Climatic conditions
III. Plant measurements global chart
IV. Statistical analysis
References
Chapter 1: Microvine, a new model for exploring grapevine response to climate warming
Abstract
Keywords
I. Introduction
I.1. Materials and methods
I.2. Plant material and growing conditions
I.3. Temporal reproductive and vegetative variables measurements
I.4. Spatial reproductive and vegetative variables measurements
I.5. Biochemical analyses
I.6. Calculations of leaf area, internode and berry volume, phyllochron and spatio-temporal conversion
I.7. Statistical analysis
II. Results
II.1. Microvine spatial patterns mimic grapevine dynamic patterns for vegetative and reproductive developments and berry metabolite accumulation
II.2. The temporal changes in berry and leaf sizes were conserved among phytomers
II.3. The spatial changes in berry, leaf and internode sizes were stable over time 79
II.4. The temporal leaf and berry developments patterns were accurately inferred from spatial profiles along the axis
II.5. Elevated temperature uncoupled biomass from volumetric growth, cut down starch storage in internodes and delayed ripening
III. Discussion
Summary
III.1. Microvine, a model that resembles grapevine
III.2. Microvine, a grapevine model with continuous and regular vegetative and
reproductive development
III.3. Microvine, a grapevine model with a few limitations
III.4. Microvine, a model for addressing grapevine responses to abiotic stresses
IV. Conclusion
V. Acknowledgements
VI. References
Chapter 2: Microvine developmental and metabolic responses to elevated temperature
I. Introduction
II. Materials and Method
II.1. Plant material and growing conditions
II.2. Development and growth measurements at the plant level
II.2. Development and growth measurements at phytomer levels
II.3. Biochemical analyses
II.4. Calculations of inflorescence abortion rates
II.5. Statistical analysis
III. Results
III.1. The rate of vegetative development was neither affected by years nor by temperature, in contrast with reproductive development
III.2. Spatial profiles of organ size, biomass and TNC concentration were altered by temperature
III.3. Fruitfulness was lower under elevated temperatures, due to inflorescences abortions
III.4. Plants were different at the onset of the experiments in 2011 and 2013
III.5. Day vs. Night temperature
IV. Discussion
IV.1. Vegetative development is stable under elevated temperatures, but reproductive development is delayed
IV.2. Elevated temperatures fasten and increase leaf and internode expansion, but not berry growth
IV.3. Inflorescence abortion present 2 different profiles from the two years of experiments
IV.4. 2011, vigorous plants, late abortion: possible explanations
IV.5. 2013, low vigor plants, early abortion: possible explanations
IV.6. Different insights from 2011 or 2013 experiments
V. Conclusion
VI. Acknowledgments
VII. References
Chapter 3: Réponses du bilan carboné de la Microvigne à la température
I. Introduction
II. Matériel & Méthode
II.1. Matériel végétal
II.2. Conditions climatiques
II.3. Mesures d’échanges gazeux
(i) Mesures d’échanges gazeux des feuilles
(ii) Mesures d’échanges gazeux des organes reproducteurs
(iii) Mesures d’échanges gazeux des plantes entières
II.4. Calculs des surfaces foliaires et des volumes d’inflorescences
II.5. Modélisation de la réponse des échanges gazeux à la température aux échelles organe et plante entière
(i) Courbes enveloppes de Amax et Rmax à l’échelle organe en fonction de la température
(ii) Analyse de covariance des résidus relatifs de An et R par rapport à Amax et Rmax à l’échelle organe
(iii) (iii)Test de la qualité des estimations de An et R aux échelles organe et plante entière
II.6. Modélisation du bilan du carbone à l’échelle plante entière
(i) Formalismes de simulation des BC aérien ou total (BCa ou BCtot)
(ii) Quantité de sucres non structuraux accumulés (Exp. 1 à 2 & 6 à 8)
(iii) Simulation du BCa pour une plante fictive soumise à différents
traitements thermiques
II.7. Analyses statistiques
III. Résultats
III.1. Réponses des échanges gazeux à la température et à la charge en fruit à l’échelle de l’organe
III.2. Réponses des échanges gazeux à la température et à la charge en fruit à l’échelle de la plante entière
III.3. Simulation et évaluation des échanges gazeux aux échelles organe et plante entière pour les expérimentations
III.4. Simulations et évaluation des bilans carbonés aérien et total pour les expérimentations
III.5. Simulation du bilan carboné aérien pour des plantes fictives soumises à un gradient de température et différents niveaux de charge en fruit
IV. Discussion
IV.1. L’élévation de la température à court terme réduit le BCa de la Microvigne via la diminution du gain de carbone net des organes aériens (DCAn-R) 203
IV.2. A long terme, l’acclimatation aux températures élevées atténue les diminutions du BCa, malgré l’augmentation de BMa, en (DCAn-R)
IV.3. Les diminutions de la charge en fruit favorisent le gain de carbone net des organes aériens (DCAn-R) et augmentent le BCa
Summary
IV.4. Des différences de BCa et d’accumulations de sucres TNC marquées entre les années : quels rôles des états initiaux de développement aérien et racinaires? 211
V. Conclusion
VI. Références

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