Photosynthetic heat tolerance of shade and sun leaves of three tropical tree species.

Previous studies of heat tolerance of tropical trees have focused on canopy leaves exposed to full sunlight and high temperatures. However, in lowland tropical forests with leaf area indices of 5-6, the vast majority of leaves experience varying degrees of shade and a reduced heat load compared to sun leaves. Here we tested whether heat tolerance is lower in shade than in sun leaves. For three tropical tree species, Calophyllum inophyllum, Inga spectabilis, and Ormosia macrocalyx, disks of fully developed shade and sun leaves were subjected to 15-min heat treatments, followed by measurement of chlorophyll a fluorescence after 48 h of recovery. In two of the three species, the temperature causing a 50% decrease of the fluorescence ratio Fv/Fm (T50) was significantly lower (by ~ 1.0 °C) in shade than in sun leaves, indicating a moderately decreased heat tolerance of shade leaves. In shade leaves of these two species, the rise in initial fluorescence, F0, also occurred at lower temperatures. In the third species, there was no shade-sun difference in T50. In situ measurements of photosynthetic CO2 assimilation showed that the optimum temperature for photosynthesis tended to be lower in shade leaves, although differences were not significant. At supra-optimal temperatures, photosynthesis was largely constrained by stomatal conductance, and the high-temperature CO2 compensation point, TMax, occurred at considerably lower temperatures than T50. Our study demonstrates that the temperature response of shade leaves of tropical trees differs only marginally from that of sun leaves, both in terms of heat tolerance and photosynthetic performance.

A tribute to Ulrich Heber (1930-2016) for his contribution to photosynthesis research: understanding the interplay between photosynthetic primary reactions, metabolism and the environment.

The dynamic and efficient coordination of primary photosynthetic reactions with leaf energization and metabolism under a wide range of environmental conditions is a fundamental property of plants involving processes at all functional levels. The present historical perspective covers 60 years of research aiming to understand the underlying mechanisms, linking major breakthroughs to current progress. It centers on the contributions of Ulrich Heber who had pioneered novel concepts, fundamental methods, and mechanistic understanding of photosynthesis. An important first step was the development of non-aqueous preparation of chloroplasts allowing the investigation of chloroplast metabolites ex vivo (meaning that the obtained results reflect the in vivo situation). Later on, intact chloroplasts, retaining their functional envelope membranes, were isolated in aqueous media to investigate compartmentation and exchange of metabolites between chloroplasts and external medium. These studies elucidated metabolic interaction between chloroplasts and cytoplasm during photosynthesis. Experiments with isolated intact chloroplasts clarified that oxygenation of ribulose-1.5-bisphosphate generates glycolate in photorespiration. The development of non-invasive optical methods enabled researchers identifying mechanisms that balance electron flow in the photosynthetic electron transport system avoiding its over-reduction. Recording chlorophyll a (Chl a) fluorescence allowed one to monitor, among other parameters, thermal energy dissipation by means of 'nonphotochemical quenching' of the excited state of Chl a. Furthermore, studies both in vivo and in vitro led to basic understanding of the biochemical mechanisms of freezing damage and frost tolerance of plant leaves, to SO2 tolerance of tree leaves and dehydrating lichens and mosses.

Protection by light against heat stress in leaves of tropical crassulacean acid metabolism plants containing high acid levels.

Heat tolerance of plants exhibiting crassulacean acid metabolism (CAM) was determined by exposing leaf sections to a range of temperatures both in the dark and the light, followed by measuring chlorophyll a fluorescence (Fv/Fm and F0) and assessing visible tissue damage. Three CAM species, Clusia rosea Jacq., Clusia pratensis Seem. and Agave angustifolia Haw., were studied. In acidified tissues sampled at the end of the night and exposed to elevated temperatures in the dark, the temperature that caused a 50% decline of Fv/Fm (T50), was remarkably low (40-43°C in leaves of C. rosea). Conversion of chlorophyll to pheophytin indicated irreversible tissue damage caused by malic acid released from the vacuoles. By contrast, when acidified leaves were illuminated during heat treatments, T50 was up to 50-51°C. In de-acidified samples taken at the end of the light period, T50 reached ∼54°C, irrespective of whether temperature treatments were done in the dark or light. Acclimation of A. angustifolia to elevated daytime temperatures resulted in a rise of T50 from ∼54° to ∼57°C. In the field, high tissue temperatures always occur during sun exposure. Measurements of the heat tolerance of CAM plants that use heat treatments of acidified tissue in the dark do not provide relevant information on heat tolerance in an ecological context. However, in the physiological context, such studies may provide important clues on vacuolar properties during the CAM cycle (i.e. on the temperature relationships of malic acid storage and malic acid release).

Light-stimulated heat tolerance in leaves of two neotropical tree species, Ficus insipida and Calophyllum longifolium.

Previous heat tolerance tests of higher plants have been mostly performed with darkened leaves. However, under natural conditions, high leaf temperatures usually occur during periods of high solar radiation. In this study, we demonstrate small but significant increases in the heat tolerance of illuminated leaves. Leaf disks of mature sun leaves from two neotropical tree species, Ficus insipida Willd. and Calophyllum longifolium Willd., were subjected to 15min of heat treatment in the light (500µmol photons m-2s-1) and in the dark. Tissue temperatures were controlled by floating the disks on the surface of a water bath. PSII activity was determined 24h and 48h after heating using chlorophyll a fluorescence. Permanent tissue damage was assessed visually during long-term storage of leaf sections under dim light. In comparison to heat treatments in the dark, the critical temperature (T50) causing a 50% decline of the fluorescence ratio Fv/Fm was increased by ~1°C (from ~52.5°C to ~53.5°C) in the light. Moreover, illumination reduced the decline of Fv/Fm as temperatures approached T50. Visible tissue damage was reduced following heat treatment in the light. Experiments with attached leaves of seedlings exposed to increasing temperatures in a gas exchange cuvette also showed a positive effect of light on heat tolerance.

Thermal tolerance, net CO2 exchange and growth of a tropical tree species, Ficus insipida, cultivated at elevated daytime and nighttime temperatures.

Global warming and associated increases in the frequency and amplitude of extreme weather events, such as heat waves, may adversely affect tropical rainforest plants via significantly increased tissue temperatures. In this study, the response to two temperature regimes was assessed in seedlings of the neotropical pioneer tree species, Ficus insipida. Plants were cultivated in growth chambers at strongly elevated daytime temperature (39°C), combined with either close to natural (22°C) or elevated (32°C) nighttime temperatures. Under both growth regimes, the critical temperature for irreversible leaf damage, determined by changes in chlorophyll a fluorescence, was approximately 51°C. This is comparable to values found in F. insipida growing under natural ambient conditions and indicates a limited potential for heat tolerance acclimation of this tropical forest tree species. Yet, under high nighttime temperature, growth was strongly enhanced, accompanied by increased rates of net photosynthetic CO2 uptake and diminished temperature dependence of leaf-level dark respiration, consistent with thermal acclimation of these key physiological parameters.

Photosynthesis, photoprotection, and growth of shade-tolerant tropical tree seedlings under full sunlight.

High solar radiation in the tropics is known to cause transient reduction in photosystem II (PSII) efficiency and CO(2) assimilation in sun-exposed leaves, but little is known how these responses affect the actual growth performance of tropical plants. The present study addresses this question. Seedlings of five woody neotropical forest species were cultivated under full sunlight and shaded conditions. In full sunlight, strong photoinhibition of PSII at midday was documented for the late-successional tree species Ormosia macrocalyx and Tetragastris panamensis and the understory/forest gap species, Piper reticulatum. In leaves of O. macrocalyx, PSII inhibition was accompanied by substantial midday depression of net CO(2) assimilation. Leaves of all species had increased pools of violaxanthin-cycle pigments. Other features of photoacclimation, such as increased Chl a/b ratio and contents of lutein, ß-carotene and tocopherol varied. High light caused strong increase of tocopherol in leaves of T. panamensis and another late-successional species, Virola surinamensis. O. macrocalyx had low contents of tocopherol and UV-absorbing substances. Under full sunlight, biomass accumulation was not reduced in seedlings of T. panamensis, P. reticulatum, and V. surinamensis, but O. macrocalyx exhibited substantial growth inhibition. In the highly shade-tolerant understory species Psychotria marginata, full sunlight caused strongly reduced growth of most individuals. However, some plants showed relatively high growth rates under full sun approaching those of seedlings at 40 % ambient irradiance. It is concluded that shade-tolerant tropical tree seedlings can achieve efficient photoacclimation and high growth rates in full sunlight.

Sun-shade patterns of leaf carotenoid composition in 86 species of neotropical forest plants.

A survey of photosynthetic pigments, including 86 species from 64 families, was conducted for leaves of neotropical vascular plants to study sun-shade patterns in carotenoid biosynthesis and occurrence of α-carotene (α-Car) and lutein epoxide (Lx). Under low light, leaves invested less in structural components and more in light harvesting, as manifested by low leaf dry mass per area (LMA) and enhanced mass-based accumulation of chlorophyll (Chl) and carotenoids, especially lutein and neoxanthin. Under high irradiance, LMA was greater and ß-carotene (ß-Car) and violaxanthin-cycle pool increased on a leaf area or Chl basis. The majority of plants contained α-Car in leaves, but the α- to ß-Car ratio was always low in the sun, suggesting preference for ß-Car in strong light. Shade and sun leaves had similar ß,ε-carotenoid contents per unit Chl, whereas sun leaves had more ß,ß-carotenoids than shade leaves. Accumulation of Lx in leaves was found to be widely distributed among taxa: >5 mmol mol Chl-1 in 20% of all species examined and >10 mmol mol Chl-1 in 10% of woody species. In Virola elongata (Benth.) Warb, having substantial Lx in both leaf types, the Lx cycle was operating on a daily basis although Lx restoration in the dark was delayed compared with violaxanthin restoration.

Lutein epoxide cycle, light harvesting and photoprotection in species of the tropical tree genus Inga.

Dynamics and possible function of the lutein epoxide (Lx) cycle, that is, the reversible conversion of Lx to lutein (L) in the light-harvesting antennae, were investigated in leaves of tropical tree species. Photosynthetic pigments were quantified in nine Inga species and species from three other genera. In Inga, Lx levels were high in shade leaves (mostly above 20 mmol mol(-1) chlorophyll) and low in sun leaves. In Virola surinamensis, both sun and shade leaves exhibited very high Lx contents (about 60 mmol mol(-1) chlorophyll). In Inga marginata grown under high irradiance, Lx slowly accumulated within several days upon transfer to deep shade. When shade leaves of I. marginata were briefly exposed to the sunlight, both violaxanthin and Lx were quickly de-epoxidized. Subsequently, overnight recovery occurred only for violaxanthin, not for Lx. In such leaves, containing reduced levels of Lx and increased levels of L, chlorophyll fluorescence induction showed significantly slower reduction of the photosystem II electron acceptor, Q(A), and faster formation as well as a higher level of non-photochemical quenching. The results indicate that slow Lx accumulation in Inga leaves may improve light harvesting under limiting light, while quick de-epoxidation of Lx to L in response to excess light may enhance photoprotection.

Photoprotection, photosynthesis and growth of tropical tree seedlings under near-ambient and strongly reduced solar ultraviolet-B radiation.

Seedlings of two late-successional tropical rainforest tree species, Tetragastris panamensis (Engler) O. Kuntze and Calophyllum longifolium (Willd.), were field grown for 3-4 months at an open site near Panama City (9 degrees N), Panama, under plastic films that either transmitted or excluded most solar UV-B radiation. Experiments were designed to test whether leaves developing under bright sunlight with strongly reduced UV-B are capable of acclimating to near-ambient UV-B conditions. Leaves of T. panamensis that developed under near-ambient UV-B contained higher amounts of UV-absorbing substances than leaves of seedlings grown under reduced UV-B. Photosynthetic pigment composition, content of alpha-tocopherol, CO(2) assimilation, potential photosystem II (PSII) efficiency (evaluated by F(v)/F(m) ratios) and growth of T. panamensis and C. longifolium did not differ between seedlings developed under near-ambient and reduced solar UV-B. When seedlings were transferred from the reduced UV-B treatment to the near-ambient UV-B treatment, a pronounced inhibition of photosynthetic capacity was observed initially in both species. UV-B-mediated inhibition of photosynthetic capacity nearly fully recovered within 1 week of the transfer in C. longifolium, whereas in T. panamensis an about 35% reduced capacity of CO(2) uptake was maintained. A marked increase in UV-absorbing substances was observed in foliage of transferred T. panamensis seedlings. Both species exhibited enhanced mid-day photoinhibition of PSII immediately after being transferred from the reduced UV-B to the near-ambient UV-B treatment. This effect was fully reversible within 1d in T. panamensis and within a few days in C. longifolium. The data show that leaves of these tropical tree seedlings, when developing in full-spectrum sunlight, are effectively protected against high solar UV-B radiation. In contrast, leaves developing under conditions of low UV-B lacked sufficient UV protection. They experienced a decline in photosynthetic competence when suddenly exposed to near-ambient UV-B levels, but exhibited pronounced acclimative responses.

Do mature shade leaves of tropical tree seedlings acclimate to high sunlight and UV radiation?

Seedlings of neotropical forest trees grown in low light were exposed to 0.5-9 h d-1 direct sunlight, for up to 3 months, to test the capability of mature shade leaves to acclimate to full solar visible and UV radiation. Photosynthetic pigments and the antioxidant, ascorbate, were analysed in leaves of two pioneer and two late-succession species. Seedlings of one or two of these species were used to assess further acclimative responses. Sun-exposure for 0.5 or 1 h d-1 resulted in strongly decreased α-carotene and increased ß-carotene and lutein levels. The pool size of xanthophyll-cycle pigments (sum of viola-, anthera- and zeaxanthin) was increased and their turnover was enhanced. These changes were associated with an increase in the capacity of non-photochemical fluorescence quenching and its 'energy-dependent' component, qE, and with reduced susceptibility to photoinhibition of PSII. Prolonged exposure to full direct sunlight (approximately 4 or 9 h d-1) resulted in a marked decrease of chlorophyll a + b content and increase in chlorophyll a / b ratios and the pool of xanthophyll-cycle pigments (based on chlorophyll), leading to extremely high zeaxanthin levels during high-light periods. Contents of ascorbate and UV-B-absorbing substances were substantially increased. PSI activity exhibited a response to full sunlight that is characteristic of sun leaves. Rates of net photosynthetic CO2 assimilation under saturating light were increased. The data show that mature shade leaves of seedlings of both early- and late-succession tree species can substantially acclimate to full-sunlight conditions by employing similar physiological mechanisms.

Sudden exposure to solar UV-B radiation reduces net CO(2) uptake and photosystem I efficiency in shade-acclimated tropical tree seedlings.

Tree seedlings developing in the understory of the tropical forest have to endure short periods of high-light stress when tree-fall gaps are formed, and direct solar radiation, including substantial UV light, reaches the leaves. In experiments simulating the opening of a tree-fall gap, the response of photosynthesis in leaves of shade-acclimated seedlings (Anacardium excelsum, Virola surinamensis, and Calophyllum longifolium) to exposure to direct sunlight (for 20-50 min) was investigated in Panama (9 degrees N). To assess the effects of solar UV-B radiation (280-320 nm), the sunlight was filtered through plastic films that selectively absorbed UV-B or transmitted the complete spectrum. The results document a strong inhibition of CO(2) assimilation by sun exposure. Light-limited and light-saturated rates of photosynthetic CO(2) uptake by the leaves were affected, which apparently occurred independently of a simultaneous inhibition of potential photosystem (PS) II efficiency. The ambient UV-B light substantially contributed to these effects. The photochemical capacity of PSI, measured as absorbance change at 810 nm in saturating far-red light, was not significantly affected by sun exposure of the seedlings. However, a decrease in the efficiency of P700 photooxidation by far-red light was observed, which was strongly promoted by solar UV-B radiation. The decrease in PSI efficiency may result from enhanced charge recombination in the reaction center, which might represent an incipient inactivation of PSI, but contributes to thermal dissipation of excessive light energy and thereby to photoprotection.

Capacity of protection against ultraviolet radiation in sun and shade leaves of tropical forest plants.

Protection of leaves of tropical forest plants against UV-A and -B radiation was studied in three lowland forests, a montane cloud forest and a mangrove stand in Panama. Leaves were classified as sun or shade leaves according to their chlorophyll a / b ratio, pool size of xanthophyll cycle pigments and α- and ß-carotene contents. The capacity of the leaves for protection against UV radiation was assessed by estimating epidermal UV-A shielding, by a non-invasive fluorometric method, and by the absorbance of ethanolic / aqueous leaf extracts in the UV spectral region. In all sun leaves tested, UV-A shielding by the adaxial epidermis was high, usually above 90%, whereas in shade leaves the epidermal UV-A shielding was markedly lower and varied widely between species. In most cases UV-A shielding by the abaxial epidermis was lower than by the adaxial epidermis. UV absorbance of the leaf extracts was generally higher in sun than in shade leaves, and the absorbance was much higher in the UV-B spectral region at 305 nm than in the UV-A region at 375 nm. The data demonstrate that sun leaves of tropical plants are well protected against solar UV-A and UV-B radiation. However, UV-induced damage may occur when shade leaves become exposed to full solar radiation.

Study of tobacco transformants to assess the role of chloroplastic NAD(P)H dehydrogenase in photoprotection of photosystems I and II.

Nicotiana tabacum L. wild-type plants and transformants (DeltandhCKJ), deficient in functional NAD(P)H dehydrogenase (NDH), were subjected to high light at 20 degrees C and 4 degrees C for 2 h to examine a possible role of NDH-mediated cyclic electron flow in protecting photosystems I and II from photoinhibition. Photochemical activity of photosystem I (PSI) was assessed by means of P700 absorbance changes at 810 nm. In addition, potential photosystem II (PSII) efficiency was determined by measuring the 'dark-adapted' ratio of variable to maximum chlorophyll fluorescence, F(v)/ F(m). Both photosystems were more susceptible to photoinhibition at 4 degrees C than at 20 degrees C. However, the degree of photoinhibition was not less in the wild type than in the NDH-deficient plants. To evaluate the efficiency of P700 oxidation in far-red light, a saturation constant, K(s), was determined, representing the far-red irradiance at which half of the maximum P700 absorbance change was reached. In photoinhibited leaves, a decrease in the efficiency of P700 oxidation (increase in K(s)) was observed. The increase in K(s) was more pronounced at 4 degrees C than at 20 degrees C, but not significantly different between wild-type and DeltandhCKJ plants. Re-reduction kinetics of oxidised P700 in the dark were accelerated to a similar extent in photoinhibited samples of both genotypes and at the two temperatures tested. The data indicate that NDH-mediated cyclic electron flow does not protect PSI against short-term light stress. It is proposed that the observed increase in K(s) represents a protective mechanism that is based on accelerated charge recombination in PSI and facilitates thermal dissipation of excessive light energy.