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Variability in ryegrass responses to elevated CO2 shows that biomass stimulation strongly associates with enhancements in leaf area/tillering capacity and is largely unrelated to leaf-level photosynthetic rate., Whilst a range of strategies have been proposed for enhancing crop productivity, many recent studies have focused primarily on enhancing leaf photosynthesis under current atmospheric CO2 concentrations. Given that the atmospheric CO2 concentration is likely to increase significantly in the foreseeable future, an alternative/complementary strategy might be to exploit any variability in the enhancement of growth/yield and photosynthesis at higher CO2 concentrations. To explore this, we investigated the responses of a diverse range of wild and cultivated ryegrass genotypes, with contrasting geographical origins, to ambient and elevated CO2 concentrations and examined what genetically tractable plant trait(s) might be targeted by plant breeders for future yield enhancements. We found substantial ~7-fold intraspecific variations in biomass productivity among the different genotypes at both CO2 levels, which were related primarily to differences in tillering/leaf area, with only small differences due to leaf photosynthesis. Interestingly, the ranking of genotypes in terms of their response to both CO2 concentrations was similar. However, as expected, estimates of whole-plant photosynthesis were strongly correlated with plant productivity. Our results suggest that greater yield gains under elevated CO2 are likely through the exploitation of genetic differences in tillering and leaf area rather than focusing solely on improving leaf photosynthesis.

Premise of research. Understanding the relationship between field-measured operating stomatal conductance (gop) and theoretical maximum stomatal conductance (gmax), calculated from stomatal density and geometry, provides an important framework that can be used to infer leaf-level gas exchange of historical, herbarium, and fossil plants. To date, however, investigation of the nature of the relationship between gop and theoretical gmax remains limited to a small number of experiments on relatively few taxa and is virtually undefined for plants in natural ecosystems.\ud \ud Methodology. We used the gop measurements of 74 species and 35 families across four biomes from a published contemporary data set of field-measured leaf-level stomatal conductance in woody angiosperms and calculated the theoretical gmax from the same leaves to investigate the relationship between gop and gmax across multiple species and biomes and determine whether such relationships are widely conserved.\ud \ud Pivotal results. We observed significant relationships between gop and gmax, with consistency in the gop ∶ gmax ratio across biomes, growth habits (shrubs and trees), and habitats (open canopy and understory subcanopy). An overall mean gop ∶ gmax ratio of 0.26 ± 0.11 (mean ± SD) was observed. The consistently observed gop ∶ gmax ratio in this study strongly agrees with previous hypotheses that an ideal gop ∶ gmax ratio exists.\ud \ud Conclusions. These results build substantially on previous studies by presenting a new reference for a consistent gop ∶ gmax ratio across many levels and offer great potential to enhance paleoclimate proxies and vegetation-climate models alike.

Background Photosynthesis is a key process in plants that is compromised by the oxygenase activity of Rubisco, which leads to the production of toxic compound phosphoglycolate that is catabolized by photorespiratory pathway. Transformation of plants with photorespiratory bypasses have been shown to reduce photorespiration and enhance plant biomass. Interestingly, engineering of a single gene from such photorespiratory bypasses has also improved photosynthesis and plant productivity. Although single gene transformations may not completely reduce photorespiration, increases in plant biomass accumulation have still been observed indicating an alternative role in regulating different metabolic processes. Therefore, the current study was aimed at evaluating the underlying mechanism (s) associated with the effects of introducing a single cyanobacterial glycolate decarboxylation pathway gene on photosynthesis and plant performance. Methods Transgenic Arabidopsis thaliana plants (GD, HD, OX) expressing independently cyanobacterial decarboxylation pathway genes i.e., glycolate dehydrogenase, hydroxyacid dehydrogenase, and oxalate decarboxylase, respectively, were utilized. Photosynthetic, fluorescence related, and growth parameters were analyzed. Additionally, transcriptomic analysis of GD transgenic plants was also performed. Results The GD plants exhibited a significant increase (16%) in net photosynthesis rate while both HD and OX plants showed a non-significant (11%) increase as compared to wild type plants (WT). The stomatal conductance was significantly higher (24%) in GD and HD plants than the WT plants. The quantum efficiencies of photosystem II, carbon dioxide assimilation and the chlorophyll fluorescence-based photosynthetic electron transport rate were also higher than WT plants. The OX plants displayed significant reductions in the rate of photorespiration relative to gross photosynthesis and increase in the ratio of the photosynthetic electron flow attributable to carboxylation reactions over that attributable to oxygenation reactions. GD, HD and OX plants accumulated significantly higher biomass and seed weight. Soluble sugars were significantly increased in GD and HD plants, while the starch levels were higher in all transgenic plants. The transcriptomic analysis of GD plants revealed 650 up-regulated genes mainly related to photosynthesis, photorespiratory pathway, sucrose metabolism, chlorophyll biosynthesis and glutathione metabolism. Conclusion This study revealed the potential of introduced cyanobacterial pathway genes to enhance photosynthetic and growth-related parameters. The upregulation of genes related to different pathways provided evidence of the underlying mechanisms involved particularly in GD plants. However, transcriptomic profiling of HD and OX plants can further help to identify other potential mechanisms involved in improved plant productivity.

Transgenic Arabidopsis thaliana plants expressing cyanobacterial decarboxylation genes GLCD1 (GLYCOLATE DEHYDROGENASE I), HDH (HYDROXYACID DEHYDROGENASE), ODC (OXALATE DECARBOXYLASE) alone, and HDH::ODC simultaneously were successfully developed. Plants independently expressing GLCD1, HDH, ODC, and HDH::ODC were referred to as GD, HD, OX, and HX plants, respectively. The single-copy homozygous GD, HD, OX, and HX plants exhibited appreciable expression of chimeric genes. Phenotypic characterization demonstrated that rosette diameter of GD, HD, OX, and HX was 20, 22, 17, and 16% higher than wild-type (WT) plants. Total numbers of leaves were 32, 35, 37, and 34% more than WT plants after 32 days of sowing. Similarly, all transgenic plants produced more cauline branches than WT plants. All transgenic plants gained more height as compared to WT when recorded after 42 days of growth except HX transgenic plants. Plants vegetative dry biomass was 43% (GD), 35% (HD), 42% (OX), and 36% (HX) higher than WT plants. This is the first report on characterization of cyanobacterial decarboxylation pathway genes, which will pave the way for transformation of complete pathway in plants for better biomass accumulation.

A single species of fossil conifer with well-preserved cuticles from multiple layers of the Dalong Formation in two sections in Duanshan Town, Guizhou Province, Southwest China has been used to reconstruct palaeo-atmospheric pCO2. The age of these layers can be dated back to the late Changhsingian (latest Permian), corresponding to the Clarkina changxingensis and Clarkina yini conodont Zones. Two calibration approaches were employed in this study. Using the stomatal ratio method, the pCO2 curves of the two sections showed matching trends, with levels decreasing in the middle of the profiles, and similar ranges at ca. 340–510 ppm and ca. 300–440 ppm. Similar values (ca. 360–520 ppm), yet with wider error ranges, were obtained when the mechanistic model proposed by Franks et al. in 2014 was applied to one of the sections. Our values are lower than the mean pCO2 values from coeval palaeosols, but partly overlap with them when error ranges are taken into consideration, thus more independent studies are needed to evaluate the discrepancies between the two proxy methods. The low atmospheric CO2 concentrations in this study suggest a cool climate rather than extensive glaciation for the brief period of late Changhsingian, which is supported by the oxygen isotope record of conodont apatite and a major restructuring of Late Permian flora and fauna.

Main conclusion Our study demonstrated that the species respond non-linearly to increases in CO2 concentration when exposed to decadal changes in CO2, representing the year 1987, 2025, 2051, and 2070, respectively. Abstract There are several lines of evidence suggesting that the vast majority of C3 plants respond to elevated atmospheric CO2 by decreasing their stomatal conductance (gs). However, in the majority of CO2 enrichment studies, the response to elevated CO2 are tested between plants grown under ambient (380–420 ppm) and high (538–680 ppm) CO2 concentrations and measured usually at single time points in a diurnal cycle. We investigated gs responses to simulated decadal increments in CO2 predicted over the next 4 decades and tested how measurements of gs may differ when two alternative sampling methods are employed (infrared gas analyzer [IRGA] vs. leaf porometer). We exposed Populus tremula, Popolus tremuloides and Sambucus racemosa to four different CO2 concentrations over 126 days in experimental growth chambers at 350, 420, 490 and 560 ppm CO2; representing the years 1987, 2025, 2051, and 2070, respectively (RCP4.5 scenario). Our study demonstrated that the species respond non-linearly to increases in CO2 concentration when exposed to decadal changes in CO2. Under natural conditions, maximum operational gs is often reached in the late morning to early afternoon, with a mid-day depression around noon. However, we showed that the daily maximum gs can, in some species, shift later into the day when plants are exposed to only small increases (70 ppm) in CO2. A non-linear decreases in gs and a shifting diurnal stomatal behavior under elevated CO2, could affect the long-term daily water and carbon budget of many plants in the future, and therefore alter soil–plant–atmospheric processes.

The stable carbon isotopes of fossil plants are a reflection of the atmosphere and environment in which they grew. Fossil plant remains have thus stored information about the isotopic composition and concentration of atmospheric carbon dioxide ( p CO 2 ) and possibly p O 2 through time. Studies to date, utilizing extant plants, have linked changes in plant stable carbon isotopes (δ 13 C p ) or carbon isotope discrimination (Δ 13 C) to changes in p CO 2 and/or p O 2 . These studies have relied heavily on angiosperm representatives, a phylogenetic group only present in the fossil record post-Early Cretaceous (∼140 million years ago (mya)), whereas gymnosperms, monilophytes and lycophytes dominated terrestrial ecosystems prior to this time. The aim of this study was to expand our understanding of carbon isotope discrimination in all vascular plant groups of C3 plants including lycophytes, monilophytes, gymnosperms and angiosperms, under elevated CO 2 and sub-ambient O 2 to explore their utility as paleo-atmospheric proxies. To achieve this goal, plants were grown in controlled environment chambers under a range of O 2 :CO 2 ratio treatments. Results reveal a strong phylogenetic dependency on Δ 13 C, where spore-bearing (lycophytes and monilophytes) have significantly higher 13 C discrimination than seed plants (gymnosperms and angiosperms) by ∼5‰. We attribute this strong phylogenetic signal to differences in C i / C a likely mediated by fundamental differences in how spore and seed bearing plants control stomatal aperture. Decreasing O 2 :CO 2 ratio in general resulted in increased carbon isotope discrimination in all plant groups. Notably, while all plant groups respond unidirectionally to elevated atmospheric CO 2 (1900 ppm and ambient O 2 ), they do not respond equally to sub-ambient O 2 (16%). We conclude that (1) Δ 13 C has a strong phylogenetic or ‘reproductive grade’ bias, whereby Δ 13 C of spore reproducing plants is significantly different to seed reproducing taxa. (2) Δ 13 C increases with decreasing O 2 :CO 2 ratios (where significant) but is more likely a result of mechanistically uncoupled responses to elevated p CO 2 and sub-ambient O 2 ; and (3) due to this response we find δ 13 C a cannot be calculated from δ 13 C p unless environmental influences such as O 2 :CO 2 ratio can be independently constrained. Therefore, interpretations of trends in fossil plant δ 13 C p to reconstruct paleoatmospheric CO 2 concentration should include cross calibration based on a nearest living relative, appropriate nearest living equivalent, or utilizing the phylogenetic corrections produced from this study.

Main conclusion Our study demonstrated that the species respond non-linearly to increases in CO2 concentration when exposed to decadal changes in CO2, representing the year 1987, 2025, 2051, and 2070, respectively. Abstract There are several lines of evidence suggesting that the vast majority of C3 plants respond to elevated atmospheric CO2 by decreasing their stomatal conductance (gs). However, in the majority of CO2 enrichment studies, the response to elevated CO2 are tested between plants grown under ambient (380–420 ppm) and high (538–680 ppm) CO2 concentrations and measured usually at single time points in a diurnal cycle. We investigated gs responses to simulated decadal increments in CO2 predicted over the next 4 decades and tested how measurements of gs may differ when two alternative sampling methods are employed (infrared gas analyzer [IRGA] vs. leaf porometer). We exposed Populus tremula, Popolus tremuloides and Sambucus racemosa to four different CO2 concentrations over 126 days in experimental growth chambers at 350, 420, 490 and 560 ppm CO2; representing the years 1987, 2025, 2051, and 2070, respectively (RCP4.5 scenario). Our study demonstrated that the species respond non-linearly to increases in CO2 concentration when exposed to decadal changes in CO2. Under natural conditions, maximum operational gs is often reached in the late morning to early afternoon, with a mid-day depression around noon. However, we showed that the daily maximum gs can, in some species, shift later into the day when plants are exposed to only small increases (70 ppm) in CO2. A non-linear decreases in gs and a shifting diurnal stomatal behavior under elevated CO2, could affect the long-term daily water and carbon budget of many plants in the future, and therefore alter soil–plant–atmospheric processes. Electronic supplementary material The online version of this article (10.1007/s00425-020-03343-z) contains supplementary material, which is available to authorized users.

The fossil record and models of atmospheric concentrations of O(2) and CO(2) suggest that past shifts in plant ecological dominance often coincided with dramatic changes in Earth’s atmospheric composition. This study tested the effects of past changes in atmospheric composition on the photosynthetic physiology of a limited range of early-diverging angiosperms (eight), gymnosperms (three), and ferns (two). We performed physiological measurements on all species and used the results to parameterize simulations of their photosynthetic paleophysiology using three independent modeling approaches. Unique physiological attributes were identified for the three evolutionary groups: angiosperm taxa displayed significantly higher mesophyll conductance (g(m)), yet their stomatal conductance (g(s)) was lower than that of ferns. Gymnosperm taxa displayed low g(s) and g(m), but they partially offset their significant diffusional limitations on photosynthesis through their higher maximum Rubisco carboxylation rate. Despite their high total conductance to CO(2), fern taxa lacked an optimized control of g(s), which was reflected in their low intrinsic water use efficiency. Simulations of the photosynthetic physiology of ferns, angiosperms, and gymnosperms through Earth’s history demonstrated that past fluctuations in O(2) and CO(2) concentrations may have resulted in significant shifts in the relative competitiveness of the three evolutionary groups. Although preliminary because of limited species sampling, these findings hint at a potential mechanistic basis for the observed broad temporal correlation between atmospheric change and shifts in plant evolutionary group-level richness observed in the fossil record and are presented as a framework to be tested with paleophotosynthetic proxies and through increased species sampling.

A rapid rise in CO2 has made evergreen trees more waterwise than deciduous trees, especially in cooler parts of the world., Intrinsic water use efficiency (iWUE), defined as the ratio of photosynthesis to stomatal conductance, is a key variable in plant physiology and ecology. Yet, how rising atmospheric CO2 concentration affects iWUE at broad species and ecosystem scales is poorly understood. In a field-based study of 244 woody angiosperm species across eight biomes over the past 25 years of increasing atmospheric CO2 (~45 ppm), we show that iWUE in evergreen species has increased more rapidly than in deciduous species. Specifically, the difference in iWUE gain between evergreen and deciduous taxa diverges along a mean annual temperature gradient from tropical to boreal forests and follows similar observed trends in leaf functional traits such as leaf mass per area. Synthesis of multiple lines of evidence supports our findings. This study provides timely insights into the impact of Anthropocene climate change on forest ecosystems and will aid the development of next-generation trait-based vegetation models.

Intrinsic water use efficiency (iWUE), defined as the ratio of photosynthesis to stomatal conductance, is a key variable in plant physiology and ecology. Yet, how rising atmospheric CO2 concentration affects iWUE at broad species and ecosystem scales is poorly understood. In a field-based study of 244 woody angiosperm species across eight biomes over the past 25 years of increasing atmospheric CO2 (~45 ppm), we show that iWUE in evergreen species has increased more rapidly than in deciduous species. Specifically, the difference in iWUE gain between evergreen and deciduous taxa diverges along a mean annual temperature gradient from tropical to boreal forests and follows similar observed trends in leaf functional traits such as leaf mass per area. Synthesis of multiple lines of evidence supports our findings. This study provides timely insights into the impact of Anthropocene climate change on forest ecosystems and will aid the development of next-generation trait-based vegetation models.

Stomatal conductance (gs) in terrestrial vegetation regulates the uptake of atmospheric carbon dioxide for photosynthesis and water loss through transpiration, closely linking the biosphere and atmosphere and influencing climate. Yet, the range and pattern of gs in plants from natural ecosystems across broad geographic, climatic, and taxonomic ranges remains poorly quantified. Furthermore, attempts to characterize gs on such scales have predominantly relied upon meta-analyses compiling data from many different studies. This approach may be inherently problematic as it combines data collected using unstandardized protocols, sometimes over decadal time spans, and from different habitat groups. Using a standardized protocol, we measured leaf-level gs using porometry in 218 C3 woody angiosperm species in natural ecosystems representing seven bioclimatic zones. The resulting dataset of 4273 gs measurements, which we call STraits (Stomatal Traits), was used to determine patterns in maximum gs (gsmax) across bioclimatic zones and whether there was similarity in the mean gsmax of C3 woody angiosperms across ecosystem types. We also tested for differential gsmax in two broadly defined habitat groups – open-canopy and understory-subcanopy – within and across bioclimatic zones. We found strong convergence in mean gsmax of C3 woody angiosperms in the understory-subcanopy habitats across six bioclimatic zones, but not in open-canopy habitats. Mean gsmax in open-canopy habitats (266 ± 100 mmol m-2 s-1) was significantly higher than in understory-subcanopy habitats (233 ± 86 mmol m-2 s-1). There was also a central tendency in the overall dataset to operate toward a gsmax of ∼250 mmol m-2 s-1. We suggest that the observed convergence in mean gsmax of C3 woody angiosperms in the understory-subcanopy is due to a buffering of gsmax against macroclimate effects which will lead to differential response of C3 woody angiosperm vegetation in these two habitats to future global change. Therefore, it will be important for future studies of gsmax to categorize vegetation according to habitat group.

Summary Understanding the drivers of geological‐scale patterns in plant macroevolution is limited by a hesitancy to use measurable traits of fossils to infer palaeoecophysiological function.Here, scaling relationships between morphological traits including maximum theoretical stomatal conductance (g max) and leaf vein density (D v) and physiological measurements including operational stomatal conductance (g op), saturated (A sat ) and maximum (A max) assimilation rates were investigated for 18 extant taxa in order to improve understanding of angiosperm diversification in the Cretaceous.Our study demonstrated significant relationships between g op, g max and D v that together can be used to estimate gas exchange and the photosynthetic capacities of fossils. We showed that acquisition of high g max in angiosperms conferred a competitive advantage over gymnosperms by increasing the dynamic range (plasticity) of their gas exchange and expanding their ecophysiological niche space. We suggest that species with a high g max (> 1400 mmol m−2 s−1) would have been capable of maintaining a high A max as the atmospheric CO 2 declined through the Cretaceous, whereas gymnosperms with a low g max would experience severe photosynthetic penalty.Expansion of the ecophysiological niche space in angiosperms, afforded by coordinated evolution of high g max , D v and increased plasticity in g op , adds further functional insights into the mechanisms driving angiosperm speciation.

Background and Aims: Studies have indicated that plant stomatal conductance (gs) decreases in response to elevated atmospheric CO2, a phenomenon of significance for the global hydrological cycle. However, gs increases across certain CO2 ranges have been predicted by optimisation models. The aim of this work was to demonstrate that under certain environmental condition, gs can increase in response to elevated CO2. Methods: When using (i) an extensive, up-to-date, synthesis of gs responses in FACE experiments, (ii) in situ measurements across four biomes showing dynamic gs responses to a CO2 rise of ~50ppm (characterising the change in this greenhouse gas over the past three decades) and (iii) a photosynthesis-stomatal conductance model, it is demonstrated that gs can in some cases increase in response to increasing atmospheric CO2. Key Results: Field observations are corroborated by an extensive synthesis of gs responses in FACE experiments showing that 11.8% of gs responses under experimentally elevated CO2 are positive. They are further supported by a strong data-model fit (r2=0.607) using a stomatal optimization model applied to the field gs dataset. A parameter space identified in the Farquhar-Ball-Berry photosynthesis-stomatal conductance model confirms field observations of increasing gs under elevated CO2 in hot dry conditions. It was shown that contrary to the general assumption, positive gs responses to elevated CO2, although relatively rare, are a feature of woody taxa adapted to warm, low-humidity conditions, and that this response is also demonstrated in global simulations using the Community Land Model (CLM4). Conclusions: The results contradict the over-simplistic notion that global vegetation always responds with decreasing gs to elevated CO2, a finding that has important implications for predicting future vegetation feedbacks on the hydrological cycle at the regional level. Irish Research Council Science Foundation Ireland

Background and aims Fluctuations in [CO 2 ] have been widely studied as a potential driver of plant evolution; however, the role of a fluctuating [O 2 ]:[CO 2 ] ratio is often overlooked. The present study aimed to investigate the inherent physiological plasticity of early diverging, extant species following acclimation to an atmosphere similar to that across the Triassic-Jurassic mass extinction interval (TJB, approx. 200 Mya), a time of major ecological change. Methods Mature plants from two angiosperm ( Drimys winteri and Chloranthus oldhamii ), two monilophyte ( Osmunda claytoniana and Cyathea australis ) and one gymnosperm ( Ginkgo biloba ) species were grown for 2 months in replicated walk-in Conviron BDW40 chambers running at TJB treatment conditions of 16 % [O 2 ]-1900 ppm [CO 2 ] and ambient conditions of 21 % [O 2 ]-400 ppm [CO 2 ], and their physiological plasticity was assessed using gas exchange and chlorophyll fluorescence methods. Key results TJB acclimation caused significant reductions in the maximum rate of carboxylation ( V Cmax ) and the maximum electron flow supporting ribulose-1,5-bisphosphate regeneration ( J max ) in all species, yet this downregulation had little effect on their light-saturated photosynthetic rate ( A sat ). Ginkgo was found to photorespire heavily under ambient conditions, while growth in low [O 2 ]:[CO 2 ] resulted in increased heat dissipation per reaction centre ( DI o / RC ), severe photodamage, as revealed by the species' decreased maximum efficiency of primary photochemistry ( F v / F m ) and decreased in situ photosynthetic electron flow ( Jsitu ). Conclusions It is argued that the observed photodamage reflects the inability of Ginkgo to divert excess photosynthetic electron flow to sinks other than the downregulated C 3 and the diminished C 2 cycles under low [O 2 ]:[CO 2 ]. This finding, coupled with the remarkable physiological plasticity of the ferns, provides insights into the underlying mechanism of Ginkgoales' near extinction and ferns' proliferation as atmospheric [CO 2 ] increased to maximum levels across the TJB.

In order to be successful in a given environment a plant should invest in a vein and stomatal network that ensures balance between both water supply and demand. Vein density (Dv) and stomatal density (SD) have been shown to be strongly positively correlated in response to a range of environmental variables in more recently evolved plant species, but the extent of this relationship has not been confirmed in earlier diverging plant lineages. In order to examine the effect of a changing atmosphere on the relationship between Dv and SD, five early-diverging plant species representing two different reproductive plant grades were grown for seven months in a palaeo-treatment comprising an O2:CO2 ratio that has occurred multiple times throughout plant evolutionary history. Results show a range of species-specific Dv and SD responses to the palaeo-treatment, however we show that the strong relationship between Dv and SD under modern ambient atmospheric composition is maintained following exposure to the palaeo-treatment. This suggests strong co-ordination between vein and stomatal traits even under relatively extreme environmental change. This co-ordination supports existing plant function proxies that use the distance between vein endings and stomata (Dm) to infer plant palaeo-physiology such as assimilation rate, and as a result, lends confidence to future application of palaeo-CO2 proxy models that require robust estimates of palaeo-assimilation rate as key initialisation parameters. European Research Council

The fine structure of the green stem of Dianthus caryophyllus , the leaf petiole and the flower pedicel of Zantedeschia aethiopica were studied using light and scanning electron microscopy. It was revealed that these non-foliar plant parts of both species possess epidermis with numerous stomata. Stomatal density of D. caryophyllus stem was found to be relatively high (79 vs 100 per mm 2 found on leaf surface). Z. aethiopica petioles and pedicels also possess numerous stomata (17 per mm 2 ), yet stomatal density was found to be about half of that of leaves. Anatomical differences observed between petioles and pedicels were only minor. Stems of D. caryophyllus as well as petioles and pedicels of Z. aethiopica have a chlorenchyma-type tissue whose fine structure is quite similar to the leaf palisade chlorenchyma. Yet, the palisade of Z. aethiopica petioles and pedicels shows a peculiar arrangement: palisade cells are arranged with their long axis parallel to the longitudinal organ axis. Palisade tissue found in the aforementioned non-foliar plant organs in both species shows strong red chlorophyll auto-fluorescence under epi-fluorescence optics, consists of cells with abundant chloroplasts, possesses high percentage of intercellular spaces (13 and 20%, respectively) and its cells expose considerable part of their surface to the intercellular air. The fine structure of this stem palisade tissue along with the abundance of functional stomata found on the epidermis may support efficient photosynthesis.

Recent reports have indicated a considerably inactivated PSII in twig cortices, in spite of the low light transmittance of overlying periderms. Corresponding information for more deeply located and less illuminated tissues like xylem rays and pith are lacking. In this investigation we aimed to characterize the efficiency of PSII and its light sensitivity along twig depth, in conjunction with the prevailing light quantity and quality. To that aim, optical methods (spectral reflectance and transmittance, chlorophyll fluorescence imaging, low temperature fluorescence spectra) and photoinhibitory treatments were applied in cut twig sections of four tree species, while corresponding leaves served as controls. Compared to leaves, twig tissues displayed lower chlorophyll (Chl) levels and dark-adapted PSII efficiency, with strong decreasing gradients towards the twig center. The low PSII efficiencies in the inner stem were not an artifact due to an actinic effect of measuring beam or to an enhanced contribution of PSI fluorescence. In fact, the PSII/PSI ratios in cortices were higher and those in the xylem rays similar to that of leaves. Inner twig tissues were quite resistant to photoinhibitory treatments, tolerating irradiation levels several-fold higher than those encountered in their microenvironment. Moreover, the extent of high light tolerance was similar in naturally exposed and shaded twig sides. The results indicate an increasing, inherent and light-independent inactivation of PSII along twig depth. The findings are discussed on the basis of a recently proposed model for photosynthetic electron flow in twigs, taking into account the specific atmospheric and light microenvironment as well as the possible metabolic needs of such bulky organs.

The leaves of some plants display an optical patchiness on their upper side, displaying light- and dark-green areas with high and low reflectance, respectively. In this investigation, we studied the fine structure of the corresponding sectors and we asked whether the lost reflected light entails a photosynthetic cost to these leaves. Four species, i.e. Arum italicum , Ranunculus ficaria , Cyclamen hederifolium and Cyclamen persicum were investigated. Scanning electron microscope examination revealed that epidermal cells of light-green sectors of all species are more bulgy than corresponding cells of neighboring dark-green leaf sectors. The comparative anatomical study revealed that (i) epidermis thickness of the light-green areas and the number of mesophyll cell layers does not differ from those of the adjacent dark-green leaf sectors and (ii) palisade cells of light-green sectors are slightly larger and more loosely arranged, allowing a much higher percentage of intercellular air spaces. The latter histological feature seems to provide the structural basis for the different optical properties between the two leaf sectors. Contrary to expectations, net photosynthetic rates (expressed on a leaf area basis) were similar in the light-green and the dark-green areas of the two cyclamen species. Yet, in C. persicum net photosynthesis was higher in the light-green areas, if expressed on a dry mass basis. The small size of the light-green spots in the rest of the test plants precluded CO 2 assimilation measurements, yet maximum linear photosynthetic electron transport rates displayed no differences between the two sectors in all plants. Thus, the assumption of a photosynthetic cost in the light-green areas was not confirmed. On the contrary, a higher construction cost was evident in the dark-green areas of three species, displaying a significantly higher specific leaf mass, without any photosynthetic benefit. The results on net photosynthesis were compatible with leaf optical properties and pigment levels. Thus, in spite of the considerably higher reflectance of the light-green areas and their lower (yet normal for a green leaf) chlorophyll levels, corresponding differences in absorptance were slight. In addition, dry mass-based pigment contents in dark-green areas were higher, while chlorophyll a / b (in two species) and carotenoid/chlorophyll ratios (in three species) were lower, pointing to a shade adaptation in these sectors. We conclude that in variegated leaves of this kind, dark-green areas are more costly to build and probably less photosynthetically active. We argue that the high pigment contents of dark-green areas establish steep light gradients in the corresponding mesophyll, rendering deeper chloroplast layers more shade adapted.

Background: Plant growth chambers provide a controlled environment to analyse the effects of environmental parameters (light, temperature, atmospheric gas composition etc.) on plant function. However, it has been shown that a ‘chamber effect’ may exist whereby results observed are not due to an experimental treatment but to inconspicuous differences in supposedly identical chambers. In this study, Vicia faba L. 'Aquadulce Claudia' (broad bean) plants were grown in eight walk-in chambers to establish if a chamber effect existed, and if so, what plant traits are best for detecting such an effect. A range of techniques were used to measure differences between chamber plants, including chlorophyll fluorescence measurements, gas exchange analysis, biomass, reproductive yield, anatomical traits and leaf stable carbon isotopes. Results and discussion: Four of the eight chambers exhibited a chamber effect. In particular, we identified two types of chamber effect which we term 'resolvable' or 'unresolved'; a resolvable chamber effect is caused by malfunctioning components of a chamber and an unresolved chamber effect is caused by unknown factors that can only be mitigated by appropriate experimental design and sufficient replication. Not all measured plant traits were able to detect a chamber effect and no single trait was capable of detecting all chamber effects. Fresh weight and flower count detected a chamber effect in three chambers, stable carbon isotopes (δ13C) and net rate CO2 assimilation (An) identified a chamber effect in two chambers, stomatal conductance (gs) and total performance index detected an effect only in one chamber. Conclusion: (1) Chamber effects can be adequately detected by fresh weight measurements and flower counts on Vicia faba plants. These methods were the most effective in terms of detection and most efficient in terms of time. (2) δ13C, gs and An measurements help distinguish between resolvable and unresolved chamber effects. (3) Unresolved chamber effects require experimental unit replication while resolvable chamber effects require investigation, repair and retesting in advance of initiating further experiments. European Research Council Science Foundation Ireland

Fast and slow chlorophyll fluorescence induction curves at high and low actinic visible light, post-illumination changes in fluorescence yield and reflectance changes at 820 nm induced by far-red light were used to characterize the state of PSII and PSI and their electron transport capabilities in chlorophyllous twig cortices of Eleagnus angustifolius L., while corresponding leaves served as controls. Twigs displayed low dark-adapted PSII photochemical efficiencies and particularly low linear electron transport rates when illuminated. In addition, their PSII population was characterized by a high proportion of inactive, non-Q(B)-reducing centers and an incomplete quenching of fluorescence during the slow induction phase. It is suggested that PSII in twigs is an inefficient electron donor to PSI and/or the reductive pentose phosphate cycle. Yet, in spite of this apparent PSII deficiency, pools of intermediate electron carriers and potential PSI activity were more than sufficient to support the observed linear electron transport rates. Moreover, the rate of PSI reduction upon far-red/dark transitions and the magnitude of fluorescence yield increase upon white light/dark transitions were compatible with an efficient electron flow to PSI from stromal donors in the absence of PSII activity. We conclude that corticular chlorenchyma may be actively engaged in cyclic at the expense of a linear electron flow and discuss the possible physiological significance of this finding in conjunction with the particular microenvironmental conditions encountered within twigs.

Light and scanning electron microscopy were used to study leaf and stem fine structure of the drought deciduous green-stemmed Mediterranean shrub Calicotome villosa (Poiret) Link. (Leguminosae). Each leaf consists of three small obovate leaflets with abundant but small (16 μm length) anomocytic stomata on both surfaces. Adaxial surface exhibits more than double stomatal density (440±8 mm −2 ) than the abaxial one (185±4 mm −2 ). T-shaped trichomes (36±3 mm −2 ) are present only on the abaxial leaf surface. Leaves are unifacial, furnished with palisade parenchyma on both sides. The stem is characterized by raised ridges and grooves. Beneath the one-cell-layered epidermis sclerenchyma is found on ridges, whereas stomata and palisade chlorenchyma are found in grooves. Hairs are abundant, especially in grooves. Stem and leaf palisade chlorenchymas are structurally similar. According to these data, photosynthesis could be efficiently supported by the stem.

A combination of gas exchange and various chlorophyll fluorescence measurements under varying O(2) and CO(2) partial pressures were used to characterize photosynthesis in green, stomata-bearing petioles of Zantedeschia aethiopica (calla lily) while corresponding leaves served as controls. Compared to leaves, petioles displayed considerably lower CO(2) assimilation rates, limited by both stomatal and mesophyll components. Further analysis of mesophyll limitations indicated lower carboxylating efficiencies and insufficient RuBP regeneration but almost similar rates of linear electron transport. Accordingly, higher oxygenation/carboxylation ratios were assumed for petioles and confirmed by experiments under non-photorespiratory conditions. Higher photorespiration rates in petioles were accompanied by higher cyclic electron flow around PSI, the latter being possibly linked to limitations in electron transport from intermediate electron carriers to end acceptors and low contents of PSI. Based on chlorophyll fluorescence methods, similar conclusions can be drawn for green pedicels, although gas exchange in these organs could not be applied due to their bulky size. Since our test plants were not subjected to stress we argue that higher photorespiration and cyclic electron flow rates are innate attributes of photosynthesis in stalks of calla lily. Active nitrogen metabolism may be inferred, while increased cyclic electron flow may provide the additional ATP required for the enhanced photorespiratory activity in petiole and pedicel chloroplasts and/or the decarboxylation of malate ascending from roots.

Some photosynthetic attributes of leaves and stems were seasonally followed in the small-leaved, summer-deciduous, green-stemmed Mediterranean shrub Calicotome villosa. Both leaves and stems displayed similar photon energy-saturated photosystem 2 (PS2) efficiencies with a minimum during winter. A second minimum in stems during the leafless summer period could be ascribed to sustained photoinhibition. Yet, stems were slightly inferior in photon capture, resulting partly from lower chlorophyll (Chl) contents and partly from higher reflectance due to pubescence. As a result, photon energy-saturated linear electron transport rates were slightly higher in leaves. However, when the total leaf and stem areas were taken into account, this superiority was abolished during autumn and winter and more than overturned during spring. Given that during summer the stems were the only photosynthetic organs, the yearly photosynthetic contribution of stems was much higher. Chl contents in stems displayed a transient and considerable summer drop, accompanied by an increase in the carotenoid to Chl ratio, indicating a photo-protective adaptation to summer drought through a decrease of photo-selective capacity, typical for leaves of many Mediterranean plants.

Atmospheric carbon dioxide concentration has shifted dynamically over the Phanerozoic according to mass balance models and the majority of proxy estimates. A new paleo-CO 2 proxy method underpinned by mechanistic understanding of plant stomatal, isotopic and photosynthetic responses to CO 2 has provocatively claimed that maximum paleoatmospheric CO 2 was capped at 1000 ppm for the majority of the past 300 million years. Here we evaluate the robustness of the new paleo-proxy CO 2 model by testing its sensitivity to initial parameterization and to scaling factors employed to estimate paleophysiological function from anatomical and morphological traits. A series of sensitivity analyses find that the model is robust to modification in some of the constants employed, such as CO 2 compensation point and mesophyll conductance, resulting in variability in paleo-CO 2 estimates which are already accounted for in the error propagation of the model. We demonstrate high sensitivity in the model to key input parameters such as initial fossil plant assimilation rate, termed A 0 and scaling factors used to estimate stomatal conductance from measurements of fossil stomata. Incorrect parameterization of A 0 has resulted in under estimation of p CO 2 by as much as 600 ppm. Despite these uncertainties, our analysis highlights that the new mechanistic paleo-CO 2 proxy of Franks et al. (2014) has significant potential to derive robust and more accurate CO 2 estimates from fossil plant stomata, as long as parameterization of A 0 is strongly justified with species appropriate morphological and anatomical data. We highlight methods that can be used to improve current estimates of fossil plant assimilation rates, reduce uncertainty associated with implementation of the Franks et al. (2014) model and importantly add to understanding of patterns of plant productivity over the Phanerozoic, for which there currently is no consensus.