VAZ Cycle Research Articles

Striking differences in frost hardiness and inability to cold acclimate in two Mougeotia species (Zygnematophyceae) from alpine and lowland habitats

AbstractZygnematophyceae, a class of freshwater green algae, exhibit distinctive seasonal dynamics. The increasing frequency of cold snaps during the growing season might challenge the persistence of some populations. The present study explored the frost hardiness of two Mougeotia species isolated from different elevations and habitats. Additionally, a phylogenetic (rbcL sequence), ultrastructural and physiological characterization was performed. Both species, grown under standard culture conditions and cold acclimated cultures (+4°C), were exposed to freezing temperatures down to −9°C. Furthermore, ultrastructural‐, hydrogen peroxide (H2O2)‐ and photosynthetic pigment analysis were performed on cells exposed to −2°C, with and without induced ice nucleation. The alpine M. disjuncta showed a higher frost hardiness (LT50 = −5.8°C), whereas the lowland M. scalaris was susceptible to ice. However, frost hardiness did not improve after cold acclimation in either species but rather decreased significantly in M. disjuncta (LT50 = −4.7°C). Despite darkness, prolonged sub‐zero temperatures or freezing induced the activation of the xanthophyll (VAZ) cycle in M. scalaris. Our results demonstrate that frost hardiness varies within the genus Mougeotia and that the VAZ cycle can be activated in the dark under subzero temperature‐ and freezing stress but does not necessarily increase frost hardiness. As highly frost hardy cell types are usually formed at the end of the growing season, the ability of young cells to survive ice formation in the upper subzero temperature range represents a crucial survival strategy in populations exposed to late spring frosts.

Chlorella vulgaris cultivation under super high light intensity: An application of the flashing light effect

Chlorella vulgaris was grown under super high illumination (7000 μmolPhotonPAR/m2/s). Continuous and flashing light patterns were applied to investigate potential gain originating from the flashing light effect (square pattern, duty cycle of 0.5). Three control configurations were tested, all under continuous light: 200 μmolPhotonPAR/m2/s (conventional cultivation conditions), 4000 μmolPhotonPAR/m2/s (about the same average amount of energy), and 7000 μmolPhotonPAR/m2/s (maximum incident light intensity, cells did not grow). The experiments were conducted in ultra-thin flat-panel photobioreactors to ensure isoactinic growth conditions. After acclimation, the monitored outcomes were: growth rate, macronutrient composition (proteins, carbohydrates, lipids), pigment content, and transient fluorescence (OJIP assays). The results showed that Chlorella vulgaris can grow at a similar rate under 200 μmolPhotonPAR/m2/s of continuous light and under 7000 μmolPhotonPAR/m2/s of flashing light for frequencies between 0.1 and 100 Hz. Acclimation mechanisms did not alter cell macronutrient composition, yet, chlorophyll contents decreased while carotenoids increased (allegedly linked to an increased expression of the VAZ cycle). OJIP tests also revealed potential up-regulation of the water-water cycle (featuring PTOX enzyme), which would allow faster repletion of the PQ pool, delaying photosynthetic apparatus saturation. Above 100 Hz (1000 Hz in this study), cells exhibited the same growth rate as under the equivalent amount of continuous light. It suggests that light alternation is too fast for the cells to perceive it. On the contrary, too low frequencies (0.01 Hz) showed lower performances than under the same average intensity. In this condition, during the light phase, cells are exposed to harmful conditions (continuous 7000 μmolPhotonPAR/m2/s), and the too-long dark phase only allows the cells some rest but does not bring any benefits. In terms of applicability, these results pave the way for the use of super high light (either artificial or by sunlight concentration) to overcome the energy limitation burdening photoautotrophic microalgae cultivation.

Xanthophyll cycles in the juniper haircap moss (Polytrichum juniperinum) and Antarctic hair grass (Deschampsia antarctica) on Livingston Island (South Shetland Islands, Maritime Antarctica)

The summer climate in Maritime Antarctica is characterised by high humidity and cloudiness with slightly above zero temperatures. Under such conditions, photosynthetic activity is temperature-limited and plant communities are formed by a few species. These conditions could prevent the operation of the photoprotective xanthophyll (VAZ) cycle as low irradiance reduces the excess of energy and low temperatures limit enzyme activity. The VAZ cycle regulates the dissipation of the excess of absorbed light as heat, which is the main mechanism of photoprotection in plants. To test whether this mechanism operates dynamically in Antarctic plant communities, we characterised pigment dynamics under natural field conditions in two representative species: the moss Polytrichum juniperinum and the grass Deschampsia antarctica. Pigment analyses revealed that the total VAZ pool was in the upper range of the values reported for most plant species, suggesting that they are exposed to a high degree of environmental stress. Despite cloudiness, there was a strong conversion of violaxanthin (V) to zeaxanthin (Z) during daytime. Conversely, the dark-induced enzymatic epoxidation back to V was not limited by nocturnal temperatures. In contrast with plants from other cold ecosystems, we did not find any evidence of overnight retention of Z or sustained reductions in photochemical efficiency. These results are of interest for modelling, remote sensing and upscaling of the responses of Antarctic vegetation to environmental challenges.

Pigment modulation in response to irradiance intensity in the fast-growing alga Picochlorum celeri

Picochlorum celeri has among the fastest photoautotrophic growth rates (~2 h doubling time in optimal conditions) reported to date for a marine alga. This study comprehensively analyzes the levels of Picochlorum celeri photosynthetic pigments, which can reach up to ~13% of the total particulate organic carbon (POC) under light-limiting conditions. The main Picochlorum celeri pigments identified include: chlorophyll a, chlorophyll b, lutein, β-carotene, canthaxanthin, violaxanthin, neoxanthin, zeaxanthin and antheraxanthin. The ketocarotenoid canthaxanthin can accumulate up to 120 mg L−1 in Picochlorum celeri liquid culture; ~12% of it was found in an extracellular polysaccharide matrix. Using a solar-simulating automated photobioreactor, we monitor the photoacclimation of cultures maintained in unshaded conditions (<0.5 μg mL−1 of total chlorophyll) through transitions from high irradiance (1000 μmoles photosynthetically active radiation (PAR) m−2 s−1) to low irradiance (60 μmoles PAR m−2 s−1), and conversely from low to high irradiance. Canthaxanthin and zeaxanthin accumulation are among the most rapid modulation responses when cultures are shifted from low to high irradiance. The violaxanthin, antherazanthin, and zeaxanthin (VAZ) pool is ~3-fold higher in high-light cultures, suggesting that the VAZ cycle combined with a dramatic reduction in chlorophyll levels are among the major mechanisms used in Picochlorum celeri to efficiently acclimate to high-irradiance levels. Responsive pigment modulation was also observed in denser cultures (~0.65 g L−1) grown under a diel cycle (pond-mimicking conditions). This research provides unique insights into the dynamics of pigment modulation and photoacclimation in response to changing irradiance in a biotechnologically promising alga and will inform future pigment engineering strategies to further improve light capture and biomass accumulation.

Kaolin Reduces ABA Biosynthesis Through the Inhibition of Neoxanthin Synthesis in Grapevines Under Water Deficit.

In many viticulture regions, multiple summer stresses are occurring with increased frequency and severity because of warming trends. Kaolin-based particle film technology is a technique that can mitigate the negative effects of intense and/or prolonged drought on grapevine physiology. Although a primary mechanism of action of kaolin is the increase of radiation reflection, some indirect effects are the protection of canopy functionality and faster stress recovery by abscisic acid (ABA) regulation. The physiological mechanism underlying the kaolin regulation of canopy functionality under water deficit is still poorly understood. In a dry-down experiment carried out on grapevines, at the peak of stress and when control vines zeroed whole-canopy net CO2 exchange rates/leaf area (NCER/LA), kaolin-treated vines maintained positive NCER/LA (~2 µmol m−2 s−1) and canopy transpiration (E) (0.57 µmol m−2 s−1). Kaolin-coated leaves had a higher violaxanthin (Vx) + antheraxanthin (Ax) + zeaxanthin (Zx) pool and a significantly lower neoxanthin (Nx) content (VAZ) when water deficit became severe. At the peak of water shortage, leaf ABA suddenly increased by 4-fold in control vines, whereas in kaolin-coated leaves the variation of ABA content was limited. Overall, kaolin prevented the biosynthesis of ABA by avoiding the deviation of the VAZ epoxidation/de-epoxidation cycle into the ABA precursor (i.e., Nx) biosynthetic direction. The preservation of the active VAZ cycle and transpiration led to an improved dissipation of exceeding electrons, explaining the higher resilience of canopy functionality expressed by canopies sprayed by kaolin. These results point out the interaction of kaolin with the regulation of the VAZ cycle and the active mechanism of stomatal conductance regulation.

Rapid formation of antheraxanthin and zeaxanthin in seconds in microalgae and its relation to non-photochemical quenching.

The violaxanthin (V)-antheraxanthin (A)-zeaxanthin (Z) (VAZ) cycle was deemed a non-second-scale process of photoprotection in higher plants and microalgae, but the validity of this view has not been confirmed. To test this view, we explored responses of the VAZ cycle and the relationship between the VAZ cycle and non-photochemical quenching (NPQ) under highlight at second and minute scales in Heterosigma akashiwo and Platymonas sp. Both A and Z were generated in H. akashiwo during 15s of light exposure, whereas only A rapidly accumulated within 15s of exposure in Platymonas sp. The above results, together with a time-dependent sigmoidal relationship between the VAZ cycle (de-epoxidation state, A/Chl a, and Z/Chl a) and NPQ, proved that the VAZ cycle was a second-scale process related to NPQ. In addition, we found that not all NPQ was dependent on the VAZ cycle and suggested that NPQ model should be carefully modified due to the species-specific proportions of de-epoxidation-dependent NPQ.

Engineering the lutein epoxide cycle into Arabidopsis thaliana

Although sunlight provides the energy necessary for plants to survive and grow, light can also damage reaction centers of photosystem II (PSII) and reduce photochemical efficiency. To prevent damage, plants possess photoprotective mechanisms that dissipate excess excitation. A subset of these mechanisms is collectively referred to as NPQ, or nonphotochemical quenching of chlorophyll a fluorescence. The regulation of NPQ is intrinsically linked to the cycling of xanthophylls that affects the kinetics and extent of the photoprotective response. The violaxanthin cycle (VAZ cycle) and the lutein epoxide cycle (LxL cycle) are two xanthophyll cycles found in vascular plants. The VAZ cycle has been studied extensively, owing in large part to its presence in model plant species where mutants are available to aid in its characterization. In contrast, the LxL cycle is not found in model plants, and its role in photosynthetic processes has been more difficult to define. To address this challenge, we introduced the LxL cycle into Arabidopsis thaliana and functionally isolated it from the VAZ cycle. Using these plant lines, we showed an increase in dark-acclimated PSII efficiency associated with Lx accumulation and demonstrated that violaxanthin deepoxidase is responsible for the light-driven deepoxidation of Lx. Conversion of Lx to L was reversible during periods of low light and occurred considerably faster than rates previously described in nonmodel species. Finally, we present clear evidence of the LxL cycle's role in modulating a rapid component of NPQ that is necessary to prevent photoinhibition in excess light.

Epiphytes Modulate Posidonia oceanica Photosynthetic Production, Energetic Balance, Antioxidant Mechanisms, and Oxidative Damage

Epiphytes impose physical barriers to light penetration into seagrass leaves causing shading, which may decrease the production of oxygen reactive species (ROS), but also constitute a physical aggression that may trigger the production of ROS, leading to oxidative damage. Here we investigate the effects of epiphytes on Posidonia oceanica under both interactive perspectives, light attenuation and oxidative stress. Specifically the role of epiphytes in net photosynthesis, chlorophyll a and b, photoprotection (Violaxanthin+Anteraxanthin+Zeaxanthin cycle), soluble sugar and starch contents, enzymatic (ascorbate peroxidase (APX) and dehydroascorbate reductase (DHAR)) and global (trolox equivalent antioxidant capacity (TEAC)) and oxygen radical antioxidant capacity (ORAC)) antioxidant responses, phenolics and oxidative damage (malondialdehyde) are tested. Leaves with epiphytes showed higher chlorophyll b and lower content in VAZ cycle carotenoids. Epiphyte shading was the probable reason for the lower VAZ de-epoxidation-ratio of leaves with epiphytes. In spite of being shaded, leaves with epiphytes showed higher antioxidant levels, indicating that epiphytes trigger the production of ROS. Both ORAC and TEAC and also APX and DHAR activities were higher in leaves with epiphytes, indicating that this response was related with its presence. Malondialdehyde concentrations also suggest oxidative damage caused by epiphytes. We conclude that the epiphyte load causes oxidative stress in P. oceanica and the mechanisms to scavenge ROS were not completely effective to avoid cell damage.

The non-foliar hypoxic photosynthetic syndrome: evidence for enhanced pools and functionality of xanthophyll cycle components and active cyclic electron flow in fruit chlorenchyma.

Green fruits display a high engagement in CEF and enhanced VAZ cycle activity as a response to the demands imposed by their internal aerial conditions, particularly low O 2 , due to gas exchange limitations. In the present study, we used HPLC analysis, post-illumination changes in fluorescence yield under varying O2 and CO2 partial pressures and absorbance changes at 820 nm induced by far-red light to assess the carotenoid composition, the functionality of the xanthophyll cycle (VAZ) and the possibility of an active cyclic e (-) flow (CEF) in the fully exposed green fruits from Nerium oleander and Rosa sp. Equally exposed, mature leaves served as controls. Compared to leaves, fruits display less total chlorophylls and carotenoids but higher Car/Chl ratio, mainly shaped by the increased pools of the VAZ cycle components, in both species. The enhanced VAZ pool size in fruits is combined with a higher mid-day de-epoxidation state (DEPS). Moreover, fruits exhibit considerably lower levels of oxidizable P700, a faster re-reduction of PSI and significantly higher relative magnitude of CEF, irrespective of the O2/CO2 levels applied. We conclude that the higher VAZ investment may serve the enhanced heat dissipation needs in fruits, in the presence of a suppressed linear e (-) flow. In addition, the elevated potential of CEF may replenish the ATP lost due to hypoxia and concurrently facilitate the development of adequate non-photochemical quenching (NPQ), through its contribution to ΔpH increase. Since other non-foliar green organs exhibit a similar photosynthetic pattern, we argue that this may reflect a common strategy for green tissues under similar micro-environmental conditions, particularly hypoxia.

Short- and long-term physiological responses of grapevine leaves to UV-B radiation

The present study aimed at evaluating the short- and long-term effects of UV-B radiation on leaves of grapevine Vitis vinifera (cv. Tempranillo). Grapevine fruit-bearing cuttings were exposed to two doses of supplemental biologically effective UV-B radiation (UV-BBE) under glasshouse-controlled conditions: 5.98 and 9.66kJm−2d−1. The treatments were applied either for 20d (from mid-veraison to ripeness) or 75d (from fruit set to ripeness). A 0kJm−2d−1 UV-B treatment was included as control. The main effects of UV-B were observed after the short-term exposure (20d) to 9.66kJm−2d−1. Significant decreases in net photosynthesis, stomatal conductance, sub-stomatal CO2 concentration, the actual photosystem II (PSII) efficiency, total soluble proteins and de-epoxidation state of the VAZ cycle were observed, whereas the activities of several antioxidant enzymes increased significantly. UV-B did not markedly affect dark respiration, photorespiration, the maximum potential PSII efficiency (Fv/Fm), non-photochemical quenching (NPQ), as well as the intrinsic PSII efficiency. However, after 75d of exposure to 5.98and 9.66kJm−2d−1 UV-B most photosynthetic and biochemical variables were unaffected and there were no sign of oxidative damage in leaves. The results suggest a high long-term acclimation capacity of grapevine to high UV-B levels, associated with a high accumulation of UV-B absorbing compounds in leaves, whereas plants seemed to be tolerant to moderate doses of UV-B.

Resposta antioxidante, formação de fitoquelatinas e composição de pigmentos fotoprotetores em Brachiaria decumbens Stapf submetida à contaminação com Cd e Zn

-1 of Cd and 500 and 2000 mmol L -1 of Zn. Metal content of shoots and roots was determined, as well as alterations in photosynthetic and photoprotective pigments, antioxidant metabolites and phytochelatin synthesis. Plants concentrated elevated levels of Cd and Zn, especially in roots. Zinc exposure negatively affected chlorophyll and β-carotene content, whereas the highest dose of Cd reduced VAZ cycle pigments and tocopherol levels in plant shoots. Cadmium was the maximum inducer of the phytochelatin synthesis pathway.

Leaf Growth and Photosynthetic Performance of Two Co-existing Oak Species in Contrasting Growing Seasons

Ecophysiological investigations of Quercus petraea and Quercus cerris were performed at the Sikfőkút research site in the dry and humid growing seasons of 2003 and 2004. The results suggested that leaf growth and the photosynthetic apparatus of Q. petraea exhibited higher sensitivity to drought in 2003 than that of Q. cerris. In leaves of Q. petraea, chlorophyll content showed larger inter-annual and within-canopy variability than in those of Q. cerris. Fully developed leaves of Q petraea showed lower SLM which indicated higher leaf cell wall elasticity allowing them to maintain a water spending strategy, while high specific leaf mass (SLM) values reflected a water saving strategy for Q. cerris. Water use efficiency of Q. cerris was higher than in the case of Q. petraea, which may provide an advantage for this species in dry periods. In the contrasting years the final leaf area and leaf mass of both species were determined by the amount of rainfall and temperature conditions during the period of early exponential phase of leaf growth. As indicated by the low values of the Fv/Fm chlorophyll fluorescence parameter the photosynthetic apparatus of both species exhibited high susceptibility to abiotic stress factors in early spring. A large VAZ cycle pool indicated that zeaxanthin dependent heat dissipation was the main contributor to photoprotection of photosynthetic apparatus in young leaves but in fully developed leaves the relatively high light saturated ETR and low Pmax as well as the maintenance of high Fv/Fm even in severe dry periods reflected the potential involvement of photorespiratory electron transport in photoprotection of both species in summer. Drought in 2003 may have resulted in serious depletion of dry matter reserves influencing the vitality of trees in following year. Q. petraea showed lower photochemical activity in the successive vegetation period after the dry year than Q. cerris which suggested lower tolerance to drought in the long term.

Seasonal changes in the xanthophyll cycle and antioxidants in sun-exposed and shaded parts of the crown of Cryptomeria japonica in relation to rhodoxanthin accumulation during cold acclimation.

Xanthophyll rhodoxanthin, which is present in sun-exposed needles of certain gymnosperms in winter, may have a photoprotective role during long-term cold acclimation. To examine how cold acclimation processes vary within tree crowns and to examine putative correlations between xanthophyll cycle pigments (VAZ), rhodoxanthin and the water-water cycle in photoprotection, we monitored seasonal changes in the activities of two key antioxidant enzymes (ascorbate peroxidase (APX) and glutathione reductase (GR)), pigment composition and chlorophyll fluorescence parameters in sun and shade needles of crowns of the gymnosperm Cryptomeria japonica D. Don. Although APX and GR activities in both sun and shade needles were higher in winter than in summer when assayed at 20 degrees C, differences between seasons were less pronounced when enzymatic activities in summer and winter were assayed at 20 and 5 degrees C, respectively. These results suggest that increases in the potential activity of antioxidant enzymes in winter is an adaptation that helps counterbalance reductions in absolute enzyme activity caused by low temperature, and thus allows the photoprotective capacity of the water-water cycle in C. japonica to be maintained at a roughly constant value throughout the year. In shade needles, the concentration of VAZ increased in winter, but no rhodoxanthin accumulated. Photosynthetic activity was maintained in winter. In sun needles, however, the electron transport rate (ETR) and photochemical quenching (q(P)) decreased to their lowest values in December, just before the accumulation of rhodoxanthin, which coincided with the highest amount of VAZ. Changes in rhodoxanthin concentration mirrored changes in VAZ concentration from January to March. Winter values of ETR and q(P) were comparable with summer values after accumulation of rhodoxanthin, indicating that rhodoxanthin may play a more important role than the VAZ cycle in protecting the photosynthetic apparatus from photodamage in winter. Photosynthetic activity may be modulated, as a result of the interception of light by rhodoxanthin, to match the extent to which absorbed light energy can be utilized in winter when the VAZ cycle is unable to operate effectively because of low temperatures.

Regulation of the xanthophyll cycle pool size in duckweed (Lemna minor) plants.

Duckweed (Lemna minor L.) plants grown under high light are characterized, when compared to low light acclimated plants, by a higher xanthophyll cycle (VAZ) pool content, but also by a higher proportion of photoconvertible violaxanthin and a superior ability to synthesize VAZ pigments. When duckweed plants were transferred to a high light environment a general response was the quick adjustment of the carotenoid composition, mainly xanthophyll cycle pigments. These changes resulted from a balance between a process of continuous light-independent carotenoid degradation and a light-induced accumulation. The use of norflurazon, an inhibitor of carotenogenesis, allowed us to demonstrate that the observed light induced increase of the VAZ pool was mainly caused by de novo synthesis through carotenogenesis. The extent of light-induced carotenogenesis was proportional to the light treatment and also to the operation of the VAZ cycle since it was partly abolished by treatments leading to a low activity of the VAZ cycle, such as low light, DTT or DCMU. These results suggest that not only the light itself, but also a mechanism triggered by a factor associated with the de-epoxidation state of the VAZ cycle controls carotenogenesis at some point before phytoene formation in the terpenoid biosynthesis pathway.

PROTEINS, PHOTOSYNTHETIC PIGMENTS AND POLAR LIPIDS CHANGES OF THYLAKOID BIOMEMBRANE FROM PRUNUS AFFECTED BY IRON CHLOROSIS

ISHS III International Symposium on Mineral Nutrition of Deciduous Fruit Trees PROTEINS, PHOTOSYNTHETIC PIGMENTS AND POLAR LIPIDS CHANGES OF THYLAKOID BIOMEMBRANE FROM PRUNUS AFFECTED BY IRON CHLOROSIS