Coupled response of stomatal and mesophyll conductance to light enhances photosynthesis of shade leaves under sunflecks

Light gradients within tree canopies play a major role in the distribution of plant resources that define the photosynthetic capacity of sun and shade leaves. However, the biochemical and diffusional constraints on gas exchange in sun and shade leaves in response to light remain poorly quantified, but critical for predicting canopy carbon and water exchange. To investigate the CO₂ diffusion pathway of sun and shade leaves, leaf gas exchange was coupled with concurrent measurements of carbon isotope discrimination to measure net leaf photosynthesis (A_n), stomatal conductance (g_s) and mesophyll conductance (g_m) in Eucalyptus tereticornis trees grown in climate controlled whole‐tree chambers. Compared to sun leaves, shade leaves had lower A_n, g_m, leaf nitrogen and photosynthetic capacity (A_max) but g_s was similar. When light intensity was temporarily increased for shade leaves to match that of sun leaves, both g_s and g_m increased, and A_n increased to values greater than sun leaves. We show that dynamic physiological responses of shade leaves to altered light environments have implications for up‐scaling leaf level measurements and predicting whole canopy carbon gain. Despite exhibiting reduced photosynthetic capacity, the rapid up‐regulation of g_m with increased light enables shade leaves to respond quickly to sunflecks.     

Seasonal relationships between leaf nitrogen content (photosynthetic capacity) and leaf canopy light exposure in peach (Prunus persica)

Abstract Field gas exchange measurements on intact peach (Prunus persica (L.) Batsch) leaves indicate that leaf nitrogen content (N_L) and leaf weight per unit leaf area (W_a) are highly correlated with CO₂ assimilation rate (A) and mesophyll conductance (g_m). Therefore, N_L and W_a were used to study seasonal relationships between leaf carboxylation capacity and natural light exposure in tree canopies. From mid-season onwards, N_L and W_a were linearly correlated with light exposure expressed as the amount of time during a clear day that a leaf was exposed to a photosynthetic photon flux density (Q) of ≥ 100 μmol m^?2 s^?1. The data support the hypothesis that whole-tree photosynthesis is optimized by partitioning of photosynthetic capacity among leaves in deciduous tree canopies with respect to natural light exposure.     

The rapid A–Ci response: photosynthesis in the phenomic era

Phenotyping for photosynthetic gas exchange parameters is limiting our ability to select plants for enhanced photosynthetic carbon gain and to assess plant function in current and future natural environments. This is due, in part, to the time required to generate estimates of the maximum rate of ribulose‐1,5‐bisphosphate carboxylase oxygenase (Rubisco) carboxylation (V_c,max) and the maximal rate of electron transport (J_max) from the response of photosynthesis (A) to the CO₂ concentration inside leaf air spaces (C_i). To relieve this bottleneck, we developed a method for rapid photosynthetic carbon assimilation CO₂ responses [rapid A–C_i response (RACiR)] utilizing non‐steady‐state measurements of gas exchange. Using high temporal resolution measurements under rapidly changing CO₂ concentrations, we show that RACiR techniques can obtain measures of V_c,max and J_max in ~5 min, and possibly even faster. This is a small fraction of the time required for even the most advanced gas exchange instrumentation. The RACiR technique, owing to its increased throughput, will allow for more rapid screening of crops, mutants and populations of plants in natural environments, bringing gas exchange into the phenomic era.     

Diffusional limitations explain the lower photosynthetic capacity of ferns as compared with angiosperms in a common garden study

Ferns are thought to have lower photosynthetic rates than angiosperms and they lack fine stomatal regulation. However, no study has directly compared photosynthesis in plants of both groups grown under optimal conditions in a common environment. We present a common garden comparison of seven angiosperms and seven ferns paired by habitat preference, with the aims of (1) confirming that ferns do have lower photosynthesis capacity than angiosperms and quantifying these differences; (2) determining the importance of diffusional versus biochemical limitations; and (3) analysing the potential implication of leaf anatomical traits in setting the photosynthesis capacity in both groups. On average, the photosynthetic rate of ferns was about half that of angiosperms, and they exhibited lower stomatal and mesophyll conductance to CO₂ (g_m), maximum velocity of carboxylation and electron transport rate. A quantitative limitation analysis revealed that stomatal and mesophyll conductances were co‐responsible for the lower photosynthesis of ferns as compared with angiosperms. However, g_m alone was the most constraining factor for photosynthesis in ferns. Consistently, leaf anatomy showed important differences between angiosperms and ferns, especially in cell wall thickness and the surface of chloroplasts exposed to intercellular air spaces.     

Photosynthetic limitation of several representative subalpine species in the Catalan Pyrenees in summer

Information on the photosynthetic process and its limitations is essential in order to predict both the capacity of species to adapt to conditions associated with climate change and the likely changes in plant communities. Considering that high‐mountain species are especially sensitive, three species representative of subalpine forests of the Central Catalan Pyrenees: mountain pine (Pinus uncinata Mill.), birch (Betula pendula Roth) and rhododendron (Rhododendron ferrugineum L.) were studied under conditions associated with climate change, such as low precipitation, elevated atmospheric [CO₂] and high solar irradiation incident at Earth's surface, in order to detect any photosynthetic limitations. Short‐term high [CO₂] increased photosynthesis rates (A) and water use efficiency (WUE), especially in birch and mountain pine, whereas stomatal conductance (g_s) was not altered in either species. Birch showed photosynthesis limitation through stomatal closure related to low rainfall, which induced photoinhibition and early foliar senescence. Rhododendron was especially affected by high irradiance, showing early photosynthetic saturation in low light, highest chlorophyll content, lowest gas exchange rates and least photoprotection. Mountain pine had the highest A, photosynthetic capacity (A_max) and light‐saturated rates of net CO₂ assimilation (A_sat), which were maintained under reduced precipitation. Furthermore, maximum quantum yield (F_v/F_m), thermal energy dissipation, PRI and SIPI radiometric index, and ascorbate content indicated improved photoprotection with respect to the other two species. However, maximum velocity of carboxylation of RuBisco (V_cmax) indicated that N availability would be the main photosynthetic limitation in this species.     

Do photosynthetic limitations of evergreen Quercus ilex leaves change with long‐term increased drought severity?

Seasonal drought can severely impact leaf photosynthetic capacity. This is particularly important for Mediterranean forests, where precipitation is expected to decrease as a consequence of climate change. Impacts of increased drought on the photosynthetic capacity of the evergreen Quercus ilex were studied for two years in a mature forest submitted to long‐term throughfall exclusion. Gas exchange and chlorophyll fluorescence were measured on two successive leaf cohorts in a control and a dry plot. Exclusion significantly reduced leaf water potential in the dry treatment. In both treatments, light‐saturated net assimilation rate (A_max), stomatal conductance (g_s), maximum carboxylation rate (V_cmax), maximum rate of electron transport (J_max), mesophyll conductance to CO₂ (g_m) and nitrogen investment in photosynthesis decreased markedly with soil water limitation during summer. The relationships between leaf photosynthetic parameters and leaf water potential remained identical in the two treatments. Leaf and canopy acclimation to progressive, long‐term drought occurred through changes in leaf area index, leaf mass per area and leaf chemical composition, but not through modifications of physiological parameters.     

Maximizing leaf carbon gain in varying saline conditions: An optimization model with dynamic mesophyll conductance

While the adverse effects of elevated salinity levels on leaf gas exchange in many crops are not in dispute, representing such effects on leaf photosynthetic rates (A) continues to draw research attention. Here, an optimization model for stomatal conductance (g_c) that maximizes A while accounting for mesophyll conductance (g_m) was used to interpret new leaf gas exchange measurements collected for five irrigation water salinity levels. A function between chloroplastic CO₂ concentration (c_c) and intercellular CO₂ concentration (c_i) modified by salinity stress to estimate g_m was proposed. Results showed that with increased salinity, the estimated g_m and maximum photosynthetic capacity were both reduced, whereas the marginal water use efficiency λ increased linearly. Adjustments of g_m, λ and photosynthetic capacity were shown to be consistent with a large corpus of drought‐stress experiments. The inferred model parameters were then used to evaluate the combined effects of elevated salinity and atmospheric CO₂ concentration (c_a) on leaf gas exchange. For a given salinity level, increasing c_a increased A linearly, but these increases were accompanied by mild reductions in g_c and transpiration. The c_a level needed to ameliorate A reductions due to increased salinity is also discussed using the aforementioned model calculations.     

CO2-forced evolution of plant gas exchange capacity and water-use efficiency over the Phanerozoic

The capacity of plants to fix carbon is ultimately constrained by two core plant attributes: photosynthetic biochemistry and the conductance to CO₂ diffusion from the atmosphere to sites of carboxylation in chloroplasts, predominantly stomatal conductance. Analysis of fossilized plant remains shows that stomatal density (number per unit area, D) and size (length by width, S) have fluctuated widely over the Phanerozoic Eon, indicating changes in maximum stomatal conductance. Parallel changes are likely to have taken place in leaf photosynthetic biochemistry, of which maximal rubisco carboxylation rate, V_cmax is a central element. We used measurements of S and D from fossilized plant remains spanning the last 400 Myr (most of the Phanerozoic), together with leaf gas exchange data and modeled Phanerozoic trends in atmospheric CO₂ concentration, [CO₂]_a, to calibrate a [CO₂]_a‐driven model of the long‐term environmental influences on S, D and V_cmax. We show that over the Phanerozoic large changes in [CO₂]_a forced S, D and V_cmax to co‐vary so as to reduce the impact of the change in [CO₂]_a on leaf CO₂ assimilation for minimal energetic cost and reduced nitrogen requirements. Underlying this is a general negative correlation between S and D, and a positive correlation between water‐use efficiency and [CO₂]_a. Furthermore, the calculated steady rise in stomatal conductance over the Phanerozoic is consistent with independent evidence for the evolution of plant hydraulic capacity, implying coordinated and sustained increase in gas exchange capacity and hydraulic capacity parallel long‐term increases in land plant diversity.     

Disentangling the contributions of ontogeny and water stress to photosynthetic limitations in almond trees

Very few studies have attempted to disentangle the respective role of ontogeny and water stress on leaf photosynthetic attributes. The relative significance of both effects on photosynthetic attributes has been investigated in leaves of field‐grown almond trees [Prunus dulcis (Mill.) D. A. Webb] during four growth cycles. Leaf ontogeny resulted in enhanced leaf dry weight per unit area (W_a), greater leaf dry‐to‐fresh weight ratio and lower N content per unit of leaf dry weight (N_w). Concomitantly, area‐based maximum carboxylation rate (V_cmax), maximum electron transport rate (J_max), mesophyll conductance to CO₂ diffusion (gm)′ and light‐saturated net photosynthesis (A_max) declined in both well‐watered and water‐stressed almond leaves. Although g_m and stomatal conductance (g_s) seemed to be co‐ordinated, a much stronger coordination in response to ontogeny and prolonged water stress was observed between g_m and the leaf photosynthetic capacity. Under unrestricted water supply, the leaf age‐related decline of A_max was equally driven by diffusional and biochemical limitations. Under restricted soil water availability, A_max was mainly limited by g_s and, to a lesser extent, by photosynthetic capacity and g_m. When both ontogeny and water stress effects were combined, diffusional limitations was the main determinant of photosynthesis limitation, while stomatal and biochemical limitations contributed similarly.     

Leaf photosynthetic capacity is regulated by the interaction of nitrogen and potassium through coordination of CO2 diffusion and carboxylation

Combined application of nitrogen (N) and potassium (K) fertilizer could significantly enhance crop yield. Crop yield and photosynthesis are inseparable. However, the influence of N and K interaction on photosynthesis is still not fully understood. Field and hydroponic experiments were conducted to examine the effects of N and K interaction on leaf photosynthesis characteristics and to explore the mechanisms in the hydroponic experiment. CO₂ conductance and carboxylation characteristic parameters of oilseed leaves were measured under different N and K supplies. Results indicated that detectable increases in leaf area, biomass and net photosynthetic rate (A_n) were observed under optimal N and K supply in field and hydroponic experiments. The ratio of total CO₂ diffusion conductance to the maximum carboxylation rate (g_tot/V_cmax) and A_n presented a linear‐plateau relationship. Under insufficient N, increased K contributed to the CO₂ transmission capacity and improved the proportion of N used for carboxylation, promoting g_tot/V_cmax. However, the low V_cmax associated with N insufficiency limited the A_n. High N supply obviously accelerated V_cmax, yet K deficiency led to a reduction of g_tot, which restricted V_cmax. Synchronous increases in N and K supplementation ensured the appropriate ratio of N to K content in leaves, which simultaneously facilitated g_tot and V_cmax and preserved a g_tot/V_cmax suitable for guaranteeing CO₂ transmission and carboxylation coordination; the overall effect was increased A_n and leaf area. These results highlight the suitable N and K nutrients to coordinate CO₂ diffusion and carboxylation, thereby enhancing photosynthetic capacity and area to obtain high crop yield.     

Mechanisms underlying leaf photosynthetic acclimation to warming and elevated CO2 as inferred from least‐cost optimality theory

The mechanisms responsible for photosynthetic acclimation are not well understood, effectively limiting predictability under future conditions. Least‐cost optimality theory can be used to predict the acclimation of photosynthetic capacity based on the assumption that plants maximize carbon uptake while minimizing the associated costs. Here, we use this theory as a null model in combination with multiple datasets of C₃ plant photosynthetic traits to elucidate the mechanisms underlying photosynthetic acclimation to elevated temperature and carbon dioxide (CO₂). The model‐data comparison showed that leaves decrease the ratio of the maximum rate of electron transport to the maximum rate of Rubisco carboxylation (J_max/V_cmax) under higher temperatures. The comparison also indicated that resources used for Rubisco and electron transport are reduced under both elevated temperature and CO₂. Finally, our analysis suggested that plants underinvest in electron transport relative to carboxylation under elevated CO₂, limiting potential leaf‐level photosynthesis under future CO₂ concentrations. Altogether, our results show that acclimation to temperature and CO₂ is primarily related to resource conservation at the leaf level. Under future, warmer, high CO₂ conditions, plants are therefore likely to use less nutrients for leaf‐level photosynthesis, which may impact whole‐plant to ecosystem functioning.     

The use of low [CO2] to estimate diffusional and non-diffusional limitations of photosynthetic capacity of salt-stressed olive saplings

In this study it has been shown that increased diffusional resistances caused by salt stress may be fully overcome by exposing attached leaves to very low [CO₂] (～ 50 µmol mol^?1), and, thus a non‐destructive‐in vivo method to correctly estimate photosynthetic capacity in stressed plants is reported. Diffusional (i.e. stomatal conductance, g_s, and mesophyll conductance to CO₂, g_m) and biochemical limitations to photosynthesis (A) were measured in two 1‐year‐old Greek olive cultivars (Chalkidikis and Kerkiras) subjected to salt stress by adding 200 mm NaCl to the irrigation water. Two sets of A–C_i curves were measured. A first set of standard A–C_i curves (i.e. without pre‐conditioning plants at low [CO₂]), were generated for salt‐stressed plants. A second set of A–C_i curves were measured, on both control and salt‐stressed plants, after pre‐conditioning leaves at [CO₂] of ～ 50 µmol mol^?1 for about 1.5 h to force stomatal opening. This forced stomata to be wide open, and g_s increased to similar values in control and salt‐stressed plants of both cultivars. After g_s had approached the maximum value, the A–C_i response was again measured. The analysis of the photosynthetic capacity of the salt‐stressed plants based on the standard A–C_i curves, showed low values of the J_max (maximum rate of electron transport) to V_cmax (RuBP‐saturated rate of Rubisco) ratio (1.06), that would implicate a reduced rate of RuBP regeneration, and, thus, a metabolic impairment. However, the analysis of the A–C_i curves made on pre‐conditioned leaves, showed that the estimates of the photosynthetic capacity parameters were much higher than in the standard A–C_i responses. Moreover, these values were similar in magnitude to the average values reported by Wullschleger (Journal of Experimental Botany 44, 907–920, 1993) in a survey of 109 C₃ species. These findings clearly indicates that: (1) salt stress did affect g_s and g_m but not the biochemical capacity to assimilate CO₂ and therefore, in these conditions, the sum of the diffusional resistances set the limit to photosynthesis rates; (2) there was a linear relationship (r² = 0.68) between g_m and g_s, and, thus, changes of g_m can be as fast as those of g_s; (3) the estimates of photosynthetic capacity based on A–C_i curves made without removing diffusional limitations are artificially low and lead to incorrect interpretations of the actual limitations of photosynthesis; and (4) the analysis of the photosynthetic properties in terms of stomatal and non‐stomatal limitations should be replaced by the analysis of diffusional and non‐diffusional limitations of photosynthesis. Finally, the C₃ photosynthesis model parameterization using in vitro‐measured and in vivo‐measured kinetics parameters was compared. Applying the in vivo‐measured Rubisco kinetics parameters resulted in a better parameterization of the photosynthesis model.     

Leaf anatomical properties in relation to differences in mesophyll conductance to CO2 and photosynthesis in two related Mediterranean Abies species

Abies alba and Abies pinsapo are closely related species with the same ribulose 1·5‐bisphosphate carboxylase/oxygenase (Rubisco) large subunit (rbcL) but contrasting hydraulic traits and mesophyll structure occurring in the Iberian Peninsula under contrasting conditions. As photosynthesis and hydraulic capacities often co‐scale, we hypothesize that these species differ in mesophyll conductance to CO₂ (g_m). g_m and key anatomical traits were measured in both species. Drought‐adapted population of A. pinsapo has higher photosynthesis than the more mesic population of A. alba, in agreement with its higher hydraulic capacity. However, A. alba exhibits the largest stomatal conductance (g_s), and so water use efficiency (WUE) is much higher in A. pinsapo. The differences in photosynthesis were explained by differences in g_m, indicating a correlation between hydraulic capacity and g_m. We report a case where g_m is the main factor limiting photosynthesis in one species (A. alba) when compared with the other one (A. pinsapo). The results also highlight the discrepancy between g_m estimates based on anatomical measurements and those based on gas exchange methods, probably due to the very large resistance exerted by cell walls and the stroma in both species. Thus, the cell wall and chloroplast properties in relation to CO₂ diffusion constitute a near‐future research priority.     

Relationship between plant hydraulic and biochemical properties derived from a steady-state coupled water and carbon transport model

There is growing evidence that plant stomata have evolved physiological controls to satisfy the demand for CO₂ by photosynthesis while regulating water losses by leaves in a manner that does not cause cavitation in the soil–root–xylem hydraulic system. Whether the hydraulic and biochemical properties of plants evolve independently or whether they are linked at a time scale relevant to plant stand development remains uncertain. To address this question, a steady‐state analytical model was developed in which supply of CO₂ via the stomata and biochemical demand for CO₂ are constrained by the balance between loss of water vapour from the leaf to the atmosphere and supply of water from the soil to the leaf. The model predicts the intercellular CO₂ concentration (C_i) for which the maximum demand for CO₂ is in equilibrium with the maximum hydraulically permissible supply of water through the soil–root–xylem system. The model was then tested at two forest stands in which simultaneous hydraulic, ecophysiological, and long‐term carbon isotope discrimination measurements were available. The model formulation reproduces analytically recent findings on the sensitivity of bulk stomatal conductance (g_s) to vapour pressure deficit (D); namely, g_s = g_ref(1 ? m × lnD), where m is a sensitivity parameter and g_ref is a reference conductance defined at D = 1 kPa. An immediate outcome of the model is an explicit relationship between maximum carboxylation capacity (V_cmax) and soil–plant hydraulic properties. It is shown that this relationship is consistent with measurements reported for conifer and rain forest angiosperm species. The analytical model predicts a decline in V_cmax as the hydraulic capacity of the soil–root–xylem decreases with stand development or age.     

In situ estimation of net CO2 assimilation, photosynthetic electron flow and photorespiration in Turkey oak (Q. cerris L.) leaves: diurnal cycles under different levels of water supply

Diurnal time courses of net CO₂ assimilation rates, stomatal conductance and light-driven electron fluxes were measured in situ on attached leaves of 30-year-old Turkey oak trees (Quercus cerris L.) under natural summer conditions in central Italy. Combined measurements of gas exchange and chlorophyll a fluorescence under low O₂ concentrations allowed the demonstration of a linear relationship between the photochemical efficiency of PSII (fluorescence measurements) and the apparent quantum yield of gross photosynthesis (gas exchange). This relationship was used under normal O₂ to compute total light-driven electron fluxes, and to partition them into fractions used for RuBP carboxylation or RuBP oxygenation. This procedure also yielded an indirect estimate of the rate of photorespiration in vivo. The time courses of light-driven electron flow, net CO₂ assimilation and photorespiration paralleled that of photosynthetic photon flux density, with important afternoon deviations as soon as a severe drought stress occurred, whereas photochemical efficiency and maximal fluorescence underwent large but reversible diurnal decreases. The latter observation indicated the occurrence of a large non-photochemical energy dissipation at PSII. We estimated that less than 60% of the total photosynthetic electron flow was used for carbon assimilation at midday, while about 40% was devoted to photorespiration. The rate of carbon loss by photorespiration (R₁) reached mean levels of 56% of net assimilation rates. The potential application of this technique to analysis of the relative contributions of thermal de-excitation at PSII and photorespiratory carbon recycling in the protection of photosynthesis against stress effects is discussed.     

Hemp (Cannabis sativa L.) leaf photosynthesis in relation to nitrogen content and temperature: implications for hemp as a bio‐economically sustainable crop

Hemp (Cannabis sativa L.) may be a suitable crop for the bio‐economy as it requires low inputs while producing a high and valuable biomass yield. With the aim of understanding the physiological basis of hemp's high resource‐use efficiency and yield potential, photosynthesis was analysed on leaves exposed to a range of nitrogen and temperature levels. Light‐saturated net photosynthesis rate (A_max) increased with an increase in leaf nitrogen up to 31.2 ± 1.9 μmol m^?2 s^?1 at 25 °C. The A_max initially increased with an increase in leaf temperature (T_L), levelled off at 25–35 °C and decreased when T_L became higher than 35 °C. Based on a C₃ leaf photosynthesis model, we estimated mesophyll conductance (g_m), efficiency of converting incident irradiance into linear electron transport under limiting light (κ_2LL), linear electron transport capacity (J_max), Rubisco carboxylation capacity (V_cmax), triose phosphate utilization capacity (T_p) and day respiration (R_d), using data obtained from gas exchange and chlorophyll fluorescence measurements at different leaf positions and various levels of incident irradiance, CO₂ and O₂. The effects of leaf nitrogen and temperature on photosynthesis parameters were consistent at different leaf positions and among different growth environments except for κ_2LL, which was higher for plants grown in the glasshouse than for those grown outdoors. Model analysis showed that compared with cotton and kenaf, hemp has higher photosynthetic capacity when leaf nitrogen is <2.0 g N m^?2. The high photosynthetic capacity measured in this study, especially at low nitrogen level, provides additional evidence that hemp can be grown as a sustainable bioenergy crop over a wide range of climatic and agronomic conditions.     

Diffusion limitations and metabolic factors associated with inhibition and recovery of photosynthesis from drought stress in a C3 perennial grass species

Stomatal closure and metabolic impairment under drought stress limits photosynthesis. The objective of this study was to determine major stomatal and metabolic factors involved in photosynthetic responses to drought and recovery upon re‐watering in a C₃ perennial grass species, Kentucky bluegrass (Poa pratensis L.). Two genotypes differing in drought resistance, ‘Midnight’ (tolerant) and ‘Brilliant’ (sensitive), were subjected to drought stress for 15 days and then re‐watered for 10 days in growth chambers. Single‐leaf net photosynthetic rate (A), stomatal conductance (g_s) and transpiration rate (Tr) decreased during drought, with a less rapid decline in ‘Midnight’ than in ‘Brilliant’. Photochemical efficiency, Rubisco activity and activation state declined during drought, but were significantly higher in ‘Midnight’ than in ‘Brilliant’. The relationship between A and internal leaf CO₂ concentration (A/Ci curve) during drought and re‐watering was analyzed to estimate the relative influence of stomatal and non‐stomatal components on photosynthesis. Stomatal limitation (Ls %), non‐stomatal limitation (Lns %), CO₂ compensation point (CP) and dark respiration (Rd) increased with stress duration in both genotypes, but to a lesser extent in ‘Midnight’. Maximum CO₂ assimilation rate (A_max), carboxylation efficiency (CE) and mesophyll conductance (g_m) declined, but ‘Midnight’ had significantly higher levels of A_max, CE and g_m than ‘Brilliant’. Maximum carboxylation rate of Rubisco (V_cmax) and ribulose‐1,5‐bisphospate (RuBP) regeneration capacity mediated by maximum electron transport rate (J_max) decreased from moderate to severe drought stress in both genotypes, but to a greater extent in ‘Brilliant’ than in ‘Midnight’. After re‐watering, RWC restored to about 90% of the control levels in both genotypes, whereas A, g_s, Tr and Fv/Fm was only partially recovered, with a higher recovery level in ‘Midnight’ than in ‘Brilliant’. Rubisco activity and activation state restored to the control level after re‐watering, with more rapid increase in ‘Midnight’ than in ‘Brilliant’. The values of Ls, Lns, CP and Rd declined, and A_max, CE, V_cmax, J_max and g_m increased after re‐watering, with more rapid change in all parameters in ‘Midnight’ than in ‘Brilliant’. These results indicated that the maintenance of higher A and A_max under drought stress in drought‐tolerant Kentucky bluegrass could be attributed to higher Rubico activation state, higher CE and less stomatal limitation. The ability to resume metabolic activity (A_max, CE, Fv/Fm and Rubisco) was observed in the drought‐tolerant genotype and is the most likely cause for the increased recuperative ability of photosynthesis. Incomplete recovery of photosynthesis upon re‐watering could be attributable to lasting stomatal limitations caused by severe drought damage in both genotypes. Promoting rapid stomatal recovery from drought stress may be critical for plants to resume full photosynthetic capacity in C₃ perennial grass species.     

Low conductances for CO2 diffusion from stomata to the sites of carboxylation in leaves of woody species

Concurrent measurements of leaf gas exchange and on-line ¹³C discrimination were used to evaluate the CO₂ conductance to diffusion from the stomatal cavity to the sites of carboxylation within the chloroplast (internal conductance; g_i). When photon irradiance was varied it appeared that g_i and/or the discrimination accompanying carboxylation also varied. Despite this problem, g_i, was estimated for leaves of peach (Prunus persica), grapefruit (Citrus paradisi), lemon (C. limon) and macadamia (Macadamia integrifolia) at saturating photon irradiance. Estimates for leaves of C. paradisi, C. limon and M. integrifolia were considerably lower than those previously reported for well-nourished herbaceous plants and ranged from 1.1 to2.2μmol CO₂ m^?2 s^?1 Pa^?1, whilst P. persica had a mean value of 3.5 μmol CO₂ m^?2 s^?1 Pa^?1. At an ambient CO₂ partial pressure of 33Pa, estimates of chloroplastic partial pressure of CO₂ (C_c) using measurements of CO₂ assimilation rate (A) and calculated values of g_i, and of partial pressure of CO₂ in the stomatal cavity (C_st) were as low as 11.2 Pa for C. limon and as high as 17.8Pa for peach. In vivo maximum rubisco activities (V_max) were also determined from estimates of C_c. This calculation showed that for a given leaf nitrogen concentration (area basis) C. paradisi and C. limon leaves had a lower V_max than P. persica, with C. paradisi and C. limon estimated to have only 10% of leaf nitrogen present as rubisco. Therefore, low CO₂ assimilation rates despite high leaf nitrogen concentrations in leaves of the evergreen species examined were explained not only by a low C_c but also by a relatively low proportion of leaf nitrogen being used for photosynthesis. We also show that simple one-dimensional equations describing the relationship between leaf internal conductance from stomatal cavities to the sites of carboxylation and carbon isotope discrimination (Δ) can lead to errors in the estimate of g_i. Potential effects of heterogeneity in stomatal aperture on carbon isotope discrimination may be particularly important and may lead to a dependence of g_i upon CO₂ assimilation rate. It is shown that for any concurrent measurement of A and Δ, the estimate of C_c is an overestimate of the correct photosynthetic capacity-weighted value, but this error is probably less than 1.0 Pa.     

Photosynthesis limitations in three fern species

Maximum photosynthesis rates in ferns are generally lower than those of seed plants, but little is known about the limiting factors, which are crucial to understand the evolution of photosynthesis in land plants. To address this issue, a gas exchange/chlorophyll fluorescence analysis was performed in three fern species spanning high phylogenetic range within Polypodiopsida (Osmunda regalis, Blechnum gibbum and Nephrolepis exaltata) to determine their maximum net photosynthesis (A_N), stomatal (g_s) and mesophyll (g_m) conductances to CO₂, and the maximum velocity of carboxylation (V_c,max). The in vitro Rubisco specificity factor (S_C/O) was also determined. All three species had values for S_C/O similar to those typical of seed plants, but values of A_N, g_s, g_m and V_c,max were within the lowest range of those observed in seed plants. In addition, g_s was unresponsive to light and CO₂, as already described in other fern species. On the contrary, g_m varied with changes CO₂. A quantitative photosynthesis limitation analysis suggested that early land plants (ferns) presented not only stomatal limitations—which were less adjustable to the environment—but also restricted g_m and V_c,max, resulting in limited maximum photosynthesis rates.     

The apoplastic antioxidant system and altered cell wall dynamics influence mesophyll conductance and the rate of photosynthesis

Mesophyll conductance (g_m), the diffusion of CO₂ from substomatal cavities to the carboxylation sites in the chloroplasts, is a highly complex trait driving photosynthesis (net CO₂ assimilation, A_N). However, little is known concerning the mechanisms by which it is dynamically regulated. The apoplast is considered as a ‘key information bridge’ between the environment and cells. Interestingly, most of the environmental constraints affecting g_m also cause apoplastic responses, cell wall (CW) alterations and metabolic rearrangements. Since CW thickness is a key determinant of g_m, we hypothesize that other changes in this cellular compartiment should also influence g_m. We study the relationship between the antioxidant apoplastic system and CW metabolism and the g_m responses in tobacco plants (Nicotiana sylvestris L.) under two abiotic stresses (drought and salinity), combining in vivo gas‐exchange measurements with analyses of antioxidant activities, CW composition and primary metabolism. Stress treatments imposed substantial reductions in A_N (58–54%) and g_m (59%), accompanied by a strong antioxidant enzymatic response at the apoplastic and symplastic levels. Interestingly, apoplastic but not symplastic peroxidases were positively related to g_m. Leaf anatomy remained mostly stable; however, the stress treatments significantly affected the CW composition, specifically pectins, which showed significant relationships with A_N and g_m. The treatments additionally promoted a differential primary metabolic response, and specific CW‐related metabolites including galactose, glucosamine and hydroxycinnamate showed exclusive relationships with g_m independent of the stress. These results suggest that g_m responses can be attributed to specific changes in the apoplastic antioxidant system and CW metabolism, opening up more possibilities for improving photosynthesis using breeding/biotechnological strategies.