This research is part of an effort that the ICT (Institute of Tropical Cultivation) has been doing for several years tending to develop superior genotypes of cocoa (Theobroma cacao L.). That is why this study aims to find tolerant or moderately tolerant cocoa genotypes and accessions to water stress with resistance to pests and diseases and high production and industrial quality. Twenty genotypes of cocoa seedlings were investigated, during the period of 6 months, in a soil with sandy-loam texture under nursery conditions, of controlled irrigation. A split plot design was used, with 40 treatments and 3 repetitions. In addition, daily data of the micro climatic characteristics (T °, HR) were taken, in which different indicators of variable were evaluated such as the stomatal conductance (CE) that is greatly influenced by the T ° and HR. The results obtained indicate that the genotypes that showed moderate tolerance to water stress were UNG - 77, UNG - 53, ICT - 1281 and ICT - 1112; the non-tolerant ones were PAS - 93, CEPEC - 2002, ICT - 2142, ICT - 1092, CP - 2005 - C10, TSH - 1188, CCN - 51, IMC - 67, PH - 17, AYP - 15, ICS - 6, BN - 34, ICT - 1506, PAS - 91, PH - 990 and ICS - 1. Under optimal conditions, the most outstanding genotype was ICS-1, both in plant height, number of leaves, and stomatal conductance, this being proof that this genotype develops excellently and stands out if it has the right conditions and water availability.

Stomatal conductance and photosynthesis

Models of vegetation function are widely used to predict the effects of climate change on carbon, water and nutrient cycles of terrestrial ecosystems, and their feedbacks to climate. Stomatal conductance, the process that governs plant water use and carbon uptake, is fundamental to such models. In this paper, we reconcile two long-standing theories of stomatal conductance. The empirical approach, which is most commonly used in vegetation models, is phenomenological, based on experimental observations of stomatal behaviour in response to environmental conditions. The optimal approach is based on the theoretical argument that stomata should act to minimize the amount of water used per unit carbon gained. We reconcile these two approaches by showing that the theory of optimal stomatal conductance can be used to derive a model of stomatal conductance that is closely analogous to the empirical models. Consequently, we obtain a unified stomatal model which has a similar form to existing empirical models, but which now provides a theoretical interpretation for model parameter values. The key model parameter, g1 ,i s predicted to increase with growth temperature and with the marginal water cost of carbon gain. The new model is fitted to a range of datasets ranging from tropical to boreal trees. The parameter g1 is shown to vary with growth temperature, as predicted, and also with plant functional type. The model is shown to correctly capture responses of stomatal conductance to changing atmospheric CO2, and thus can be used to test for stomatal acclimation to elevated CO2. The reconciliation of the optimal and empirical approaches to modelling stomatal conductance is important for global change biology because it provides a simple theoretical framework for analyzing, and simulating, the coupling between carbon and water cycles under environmental change.

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Reconciling the optimal and empirical approaches to modelling stomatal conductance

Stomatal pores, each surrounded by a pair of guard cells, regulate CO2 uptake and water loss from leaves. Stomatal opening is driven by the accumulation of K+ salts and sugars in guard cells, which is mediated by electrogenic proton pumps in the plasma membrane and/or metabolic activity. Opening responses are achieved by coordination of light signaling, light-energy conversion, membrane ion transport, and metabolic activity in guard cells. In this review, we focus on recent progress in blue- and red-light-dependent stomatal opening. Because the blue-light response of stomata appears to be strongly affected by red light, we discuss underlying mechanisms in the interaction between blue-light signaling and guard cell chloroplasts.

Light Regulation of Stomatal Movement

Publisher Summary This chapter focuses on the metabolite levels in specific cells and the subcellular compartments of plant leaves. Leaf material is difficult to fractionate because a typical plant cell contains several different, mechanically fragile subcellular organelles and is surrounded by a mechanically strong plant cell wall. Rather than attempting to isolate whole organelles or cells, leaves are frozen in liquid N 2 and then broken to small fragments that are enriched in material from a given compartment. These fragments are physically separated under conditions when the metabolic activity or redistribution of metabolites is prevented and subsequently their metabolism is quenched. The exclusion of water, or the use of extremely low temperatures, prevents metabolic activity during the fractionation procedures. The chapter describes the silicone oil centrifugation that allows chloroplasts to be separated from the remainder of the protoplast and to be quenched within 2–3 sec of disrupting the protoplast.

Metabolite levels in specific cells and subcellular compartments of plant leaves

Stomata can be regarded as hydraulically driven valves in the leaf surface, which open to allow CO2 uptake and close to prevent excessive loss of water. Movement of these 'Watergates' is regulated by environmental conditions, such as light, CO2 and humidity. Guard cells can sense environmental conditions and function as motor cells within the stomatal complex. Stomatal movement results from the transport of K+ salts across the guard cell membranes. In this review, we discuss the biophysical principles and mechanisms of stomatal movement and relate these to ion transport at the plasma membrane and vacuolar membrane. Studies with isolated guard cells, combined with recordings on single guard cells in intact plants, revealed that light stimulates stomatal opening via blue light-specific and photosynthetic-active radiation-dependent pathways. In addition, guard cells sense changes in air humidity and the water status of distant tissues via the stress hormone abscisic acid (ABA). Guard cells thus provide an excellent system to study cross-talk, as multiple signaling pathways induce both short- and long-term responses in these sensory cells.

In the light of stomatal opening: new insights into ‘the Watergate’

Using quantitative histochemical techniques, the carbohydrate levels of guard cells from open and closed stomatal apparatus of Vicia faba L. were compared. To minimize experimental error, all comparisons were between leaflets of the same pair. Stomata on one leaflet were caused to open by light and reduced CO(2). The other leaflet, which was in darkness, had closed stomata. In one experiment, data were also collected on palisade parenchyma, spongy parenchyma, and epidermal cells.Guard cell starch concentration was higher in the leaflets with closed stomata than in open stomata by 72 +/- 16 millimoles per kilogram dry weight (anhydroglucosyl equivalents) (N = 117, P < 0.02). Variation in guard cell starch concentration from one part of a leaflet to another was small. The data are consistent with the hypothesis that starch degradation provides the carbon skeletons for anion synthesis in guard cells during stomatal opening.Sucrose concentration was higher in guard cells when stomata were open than when they were closed in all three experiments (average difference = 45 +/- 7 millimoles per kilogram dry weight [N = 59, P < 0.01]). The variability of sucrose concentration within test leaflets prevented an unequivocal interpretation of these results. When all data are considered, it appears that soluble sugars increase in guard cells when stomata of Vicia faba open.

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Guard cell starch concentration quantitatively related to stomatal aperture.

A new procedure is reported for high-yield isolation of guard cell protoplasts from Vicia faba L. Delayed light emission and P700 content plus absorption and fluorescence emission spectra of these protoplast extracts are reported. It is concluded that both photosystems are present. The presence of photosystem II and the absence of the reductive-step enzyme of the Calvin-Benson Cycle (Outlaw WH Jr, J Manchester, CH DiCamelli, DD Randall, B Rapp, GM Veith 1979 Proc Natl Acad Sci USA 76: 6371-6375) in a cell has no precedent in the literature. It is speculated that noncyclic photosynthetic electron flow is an environmental sensor which causes stomata to remain open in light.

/pdf/presence-of-both-photosystems-in-guard-cells-of-vicia-faba-l-13s29bswp7.pdf

Presence of Both Photosystems in Guard Cells of Vicia faba L: IMPLICATIONS FOR ENVIRONMENTAL SIGNAL PROCESSING

Leaflets of Vicia faba L. with either open or closed stomata were quick-frozen and freeze-dried. Individual guard cell pairs and pure samples of palisade parenchyma, spongy parenchyma, and epidermis lacking guard cells were dissected from the leaflets, weighed, and assayed for organic acids or K+.K+ was measured by a new enzymatic method. In guard cells of open stomata, as compared to closed stomata, K+ was 2- to 4-fold higher, malic acid 6-fold higher, and citric acid 3-fold higher. Both aspartic and glutamic acids were also higher, but the amounts present were low compared to malic and citric acids. Isocitric acid was significantly higher in one experiment, but not in another. Glyceric acid was not increased. Succinic acid was too low to detect by the method used; but in guard cells of open stomata the concentration must have been less than 2% of that of malic acid. Malic acid was higher in the palisade parenchyma from the leaflet with open stomata. The ion balance shows that malic and citric acids provide much of the counter ion for the K+ taken up during stomatal opening

https://www.pnas.org/content/pnas/74/10/4434.full.pdf

Organic acid and potassium accumulation in guard cells during stomatal opening.

Four cell types from Vicia faba Linnaeus “Long Pod” leaflets were assayed for three enzymes unique to the photosynthetic carbon reduction pathway. The enzymes were ribulosebisphosphate carboxylase [3-phospho-D-glycerate carboxy-lyase (dimerizing), EC 4.1.1.39], phosphoribulokinase (ATP:D-ribulose-5-phosphate 1-phosphotransferase, EC 2.7.1.19), and glyceraldehyde-phosphate dehydrogenase (NADP+) (phosphorylating) [D-glyceraldehyde-3-phosphate:NADP+ oxidoreductase (phosphorylating), EC 1.2.1.13]. On a dry weight basis, these enzyme activities were about twice as high in palisade as in spongy parenchyma. Two of the enzymes were not detected in epidermal cells and the other was present in only a trace amount. In guard cells, these enzyme activities were absent or present at les than 1% of the amount in palisade cells. Immunoelectrophoresis showed that ribulosebisphosphate carboxylase was absent in extracts of guard cell protoplasts. Microscopy confirmed the abundance of typical guard cell chloroplasts. These results demonstrate the absence of the photosynthetic carbon reduction pathway in guard cell chloroplasts. This is the only chloroplast type known to be deficient in this pathway in plants whose primary CO2 acceptor is ribulose bisphosphate. Possible reasons for the absence of this pathway in guard cells are discussed.

http://www.pnas.org/content/76/12/6371.full.pdf

Photosynthetic carbon reduction pathway is absent in chloroplasts of Vicia faba guard cells

Specific activities of NADP-malic enzyme, NAD-malic enzyme, phosphoenolpyruvate carboxykinase and pyruvate, orthophosphate dikinase in various cells of Vicia faba L leaflets were determined Expressed on dry weight, chlorophyll or protein basis, the averages for NADP- and NAD-malic enzyme specific activities were higher in guard cells than in photosynthetic parenchyma cells Malic enzyme-specific activities were also high in epidermal cells Phosphoenolypyruvate carboxykinase activity was not detected in Vicia leaf extracts or guard cells; the assay techniques were validated by mixed Vicia-Brachiaria leaf extraction and assays on nanogram samples of Brachiaria bundlesheath cells It was inferred from these data that guard cell malate depletion is by decarboxylation to pyruvate in the epidermal layer, but how the various epidermal cells interact remains obscure Pyruvate, orthophosphate dikinase activity could not be demonstrated unequivocally in Vicia leaf extracts, Vicia guard cell protoplast extracts, or in Vicia guard cells The assay techniques were validated by mixed Vicia-Kochia leaf extraction and assays on nanogram samples of Kochia mesophyll cells How (or if) pyruvate is phosphorylated by epidermal tissue for entry into gluconeogenesis is unknown

William H. Outlaw

Papers

Guard cell starch concentration quantitatively related to stomatal aperture.

Presence of Both Photosystems in Guard Cells of Vicia faba L: IMPLICATIONS FOR ENVIRONMENTAL SIGNAL PROCESSING

Organic acid and potassium accumulation in guard cells during stomatal opening.

Photosynthetic carbon reduction pathway is absent in chloroplasts of Vicia faba guard cells

High Levels of Malic Enzyme Activities in Vicia faba L. Epidermal Tissue.