Torsten Jakob

Regulation and function of xanthophyll cycle-dependent photoprotection in algae

The xanthophyll cycle represents one of the important photoprotection mechanisms in plant cells. In the present review, we summarize current knowledge about the violaxanthin cycle of vascular plants, green and brown algae, and the diadinoxanthin cycle of the algal classes Bacillariophyceae, Xanthophyceae, Haptophyceae, and Dinophyceae. We address the biochemistry of the xanthophyll cycle enzymes with a special focus on protein structure, co-substrate requirements and regulation of enzyme activity. We present recent ideas regarding the structural basis of xanthophyll cycle-dependent photoprotection, including different models for the mechanism of non-photochemical quenching of chlorophyll a fluorescence. In a dedicated chapter, we also describe the unique violaxanthin antheraxanthin cycle of the Prasinophyceae, together with its implication for the mechanism of xanthophyll cycle-dependent heat dissipation. The interaction between the diadinoxanthin cycle and alternative electron flow pathways in the chloroplasts of diatoms is an additional topic of this review, and in the last chapter we cover aspects of the importance of xanthophyll cycle-dependent photoprotection for different algal species in their natural environments.

Unusual pH-dependence of diadinoxanthin de-epoxidase activation causes chlororespiratory induced accumulation of diatoxanthin in the diatom Phaeodactylum tricornutum

Summary Based on our recent findings that in the diatom Phaeodactylum tricornutum, chlororespiration in periods of prolonged darkness leads to the accumulation of diatoxanthin (DT), we have elaborated in detail the interdependence between the chlororespiratory proton gradient and the activation of diadinoxanthin de-epoxidase (DDE). The data clearly demonstrates that activation of DDE in Phaeodactylum occurs at higher pH-values compared to activation of violaxanthin de-epoxidase (VDE) in higher plants. In thylakoid membranes as well as in enzyme assays with isolated DDE, the de-epoxidation of diadinoxanthin (DD) is efficiently catalyzed at pH 7.2. In comparison, de-epoxidation of violaxanthin (Vx) in spinach thylakoids is observed below pH 6.5. Phaeodactylum thylakoids isolated from high light grown cells, that also contain the pigments of the violaxanthin cycle, show violaxanthin de-epoxidation at higher pH-values, thus suggesting that in Phaeodactylum, one de-epoxidase converts both diadinoxanthin and violaxanthin. We conclude that the activation of DDE at higher pH-values can explain how the low rates of chlororespiratory electron flow, that lead to the build-up of a rather small proton gradient, can induce the observed accumulation of diatoxanthin in the dark. Furthermore, we show that dark activation of diadinoxanthin de-epoxidation is not restricted to Phaeodactylum tricornutum but was also found in another diatom, Cyclotella meneghiana

Molecular dynamics of the diatom thylakoid membrane under different light conditions

During the last years significant progress was achieved in unraveling molecular characteristics of the thylakoid membrane of different diatoms. With the present review it is intended to summarize the current knowledge about the structural and functional changes within the thylakoid membrane of diatoms acclimated to different light conditions. This aspect is addressed on the level of the organization and regulation of light-harvesting proteins, the dissipation of excessively absorbed light energy by the process of non-photochemical quenching, and the lipid composition of diatom thylakoid membranes. Finally, a working hypothesis of the domain formation of the diatom thylakoid membrane is presented to highlight the most prominent differences of heterokontic thylakoids in comparison to vascular plants and green algae during the acclimation to low and high light conditions.

From photons to biomass and biofuels: evaluation of different strategies for the improvement of algal biotechnology based on comparative energy balances

Microalgal based biofuels are discussed as future sustainable energy source because of their higher photosynthetic and water use efficiency to produce biomass. In the context of climate CO2 mitigation strategies, algal mass production is discussed as a potential CO2 sequestration technology which uses CO2 emissions to produce biomass with high-oil content independent on arable land. In this short review, it is presented how complete energy balances from photon to harvestable biomass can help to identify the limiting processes on the cellular level. The results show that high productivity is always correlated with high metabolic costs. The overall efficiency of biomass formation can be improved by a photobioreactor design which is kinetically adapted to the rate-limiting steps in cell physiology. However, taking into account the real photon demand per assimilated carbon and the energy input for biorefinement, it becomes obvious that alternative strategies must be developed to reach the goal of a real CO2 sequestration.

Estimation of chlorophyll content and daily primary production of the major algal groups by means of multiwavelength-excitation PAM chlorophyll fluorometry: performance and methodological limits.

The performance and methodological limits of the Phyto-PAM chlorophyll fluorometer were investigated with laboratory grown algae cultures and natural phytoplankton from the rivers Saar and Saale. The Phyto-PAM is a 4-wavelength chlorophyll fluorometer with the functional combination of chlorophyll (Chl) estimation and assessment of photosynthetic activity, both differentiated into the main algal groups. The reliability of fluorescence-based Chl estimation strongly depends on the group specific calibration of the instrument and the resulting chlorophyll/fluorescence (Chl/F) ratios in reference algal cultures. A very high reliability of the Chl estimation was obtained in the case of constant Chl/F-ratios. Algae grown at different light intensities showed marked differences in Chl/F-ratios, reflecting differences in pigment composition and Chl a specific absorption (a*). When the Phyto-PAM was calibrated with laboratory grown diatoms, the Chl a in river grown diatoms was underestimated, due a lower content of accessory pigments and stronger pigment packaging. While this aspect presently limits the application of PAM fluorometry in limnology, this limitation may be overcome by future technical progress in the detection of dynamic changes in Chl/F-ratio via fluorescence-based measurements of the functional PS II absorption cross-section. Practically identical Chl/F-ratios were found for the diatom-dominated waters of the rivers␣Saar and Saale, suggesting that the same instrument calibration parameters may be applied for hydrographically similar surface waters. For this particular case, despite of the present methodological limitations, the potential of PAM fluorometry in limnology could be demonstrated. Light response curves were measured to estimate primary production with a spectrally resolved model in daily courses at two sampling sites. Fluorescence based primary production was closely correlated with measured oxygen evolution rates until midday. In the afternoon, at the water surface the fluorescence approach gave higher␣rates than the measured oxygen evolution. Possible explanations for the observed differences are discussed.

The regulation of xanthophyll cycle activity and of non-photochemical fluorescence quenching by two alternative electron flows in the diatoms Phaeodactylum tricornutum and Cyclotella meneghiniana.

Intact cells of diatoms are characterized by a rapid diatoxanthin epoxidation during low light periods following high light illumination while epoxidation is severely restricted in phases of complete darkness. The present study shows that rapid diatoxanthin epoxidation is dependent on the availability of the cofactor of diatoxanthin epoxidase, NADPH, which cannot be generated in darkness due to the inactivity of PSI. In the diatom Phaeodactylum tricornutum, NADPH production during low light is dependent on PSII activity, and addition of DCMU consequently abolishes diatoxanthin epoxidation. In contrast to P. tricornutum, DCMU does not affect diatoxanthin epoxidation in Cyclotella meneghiniana, which shows the same rapid epoxidation in low light both in the absence or presence of DCMU. Measurements of the reduction state of the PQ pool and PSI activity indicate that, in the presence of DCMU, NADPH production in C. meneghiniana occurs via alternative electron transport, which includes electron donation from the chloroplast stroma to the PQ pool and, in a second step, from PQ to PSI. Similar electron flow to PQ is also observed during high light illumination of DCMU-treated P. tricornutum cells. In contrast to C. meneghiniana, the electrons are not directed to PSI, but most likely to a plastoquinone oxidase. This chlororespiratory electron transport leads to the establishment of an uncoupler-sensitive proton gradient in the presence of DCMU, which induces diadinoxanthin de-epoxidation and NPQ. In C. meneghiniana, electron flow to the plastoquinone oxidase is restricted, and consequently, diadinoxanthin de-epoxidation and NPQ is not observed after addition of DCMU.

Blue light is essential for high light acclimation and photoprotection in the diatom Phaeodactylum tricornutum

The objective of the present study was to test the hypothesis that the acclimation to different light intensities in the diatom Phaeodactylum tricornutum is controlled by light quality perception mechanisms. Therefore, semi-continuous cultures of P. tricornutum were illuminated with equal amounts of photosynthetically absorbed radiation of blue (BL), white (WL), and red light (RL) and in combination of two intensities of irradiance, low (LL) and medium light (ML). Under LL conditions, growth rates and photosynthesis rates were similar for all cultures. However, BL cultures were found to be in an acclimation state with an increased photoprotective potential. This was deduced from an increased capacity of non-photochemical quenching, a larger pool of xanthophyll cycle pigments, and a higher de-epoxidation state of xanthophyll cycle pigments compared to WL and RL cultures. Furthermore, in the chloroplast membrane proteome of BL cells, an upregulation of proteins involved in photoprotection, e.g. the Lhcx1 protein and zeaxanthin epoxidase, was evident. ML conditions induced increased photosynthesis rates and a further enhanced photoprotective potential for algae grown under BL and WL. In contrast, RL cultures exhibited no signs of acclimation towards increased irradiance. The data implicate that in diatoms the photoacclimation to high light intensities requires the perception of blue light.

Investigation of the quenching efficiency of diatoxanthin in cells of Phaeodactylum tricornutum (Bacillariophyceae) with different pool sizes of xanthophyll cycle pigments

A. Schumann, R. Goss, T. Jakob and C. Wilhelm. 2007. Investigation of the quenching efficiency of diatoxanthin in cells of Phaeodactylum tricornutum (Bacillariophyceae) with different pool sizes of xanthophyll cycle pigments. Phycologia 46: 113–117. DOI: 10.2216/06-30.1 In the present study, the content of diadinoxanthin (Dd) + diatoxanthin (Dt) in the diatom Phaeodactylum tricornutum was manipulated by low-light (LL) and high-light (HL) growth conditions. Cells grown in HL almost doubled the pool size of Dd cycle pigments compared to cells from a LL culture. HL-grown cultures exhibited a stronger de-epoxidation and higher amounts of Dt during actinic HL illumination. However, the correlation between Dt and the corresponding nonphotochemical quenching (NPQ) revealed that the quenching efficiency of Dt was drastically decreased in these HL-grown cells compared to cells from the LL cultivation. The correlation between the de-epoxidation state of the Dd cycle and NPQ implied that in HL-grown cells a part of the Dt molecules had lost their ability to contribute to the NPQ. A precise quantification of the amount of free Dd + Dt by anion exchange chromatography and high-performance liquid chromatography showed that only 10% of the additionally synthesized Dd cycle pigments existed as free pigment in the thylakoid membrane and that most of the Dd + Dt was still bound to the light-harvesting proteins. The possible function of the additional free Dt in photo-protection and the antenna-bound Dt in the synthesis of the light-harvesting pigment fucoxanthin is discussed.

A New Multicomponent NPQ Mechanism in the Diatom Cyclotella meneghiniana

In the present study we report that in the diatom Cyclotella meneghiniana the diatoxanthin-dependent non-photochemical quenching of chlorophyll fluorescence (NPQ) is heterogeneous and consists of three different components. (i) A transient NPQ component that generates immediately upon illumination, depends on the transthylakoid proton gradient as well as on the light intensity, and is modulated by the initial diatoxanthin content of the cells. It is located in the antenna complexes of C. meneghiniana and is comparable with the transient NPQ observed in vascular plants. (ii) A steady-state NPQ component is observed during later stages of the high-light illumination and depends on the diatoxanthin content formed by the light-activated diadinoxanthin cycle. (iii) A fast relaxing NPQ component is seen upon a transition of high-light-illuminated cells to complete darkness. This component relaxes within a time frame of tens of seconds and its extent is correlated with the amount of diatoxanthin formed during the phase of actinic illumination. It cannot be observed in dithiothreitol-treated cells where the de-epoxidation of diadinoxanthin to diatoxanthin is suppressed. The fast relaxing component can be interpreted as a relaxation of part of the steady-state NPQ. The different diatoxanthin-dependent components are characterized by different quenching efficiencies of diatoxanthin. Diatoxanthin involved in the transient NPQ exhibits a 2-fold higher quenching efficiency compared with diatoxanthin participating in the steady-state NPQ. It is proposed that the different quenching efficiencies of diatoxanthin are caused by the existence of different diatoxanthin pools within the antenna system of C. meneghiniana.

Aureochrome 1a is involved in the photoacclimation of the diatom Phaeodactylum tricornutum.

Aureochromes constitute a family of blue light (BL) receptors which are found exclusively in heterokont algae such as diatoms (Bacillariophyceae) and yellow-green algae (Xanthophyceae). Previous studies on the diatom Phaeodactylum tricornutum indicate that the formation of a high light acclimated phenotype is mediated by the absorption of BL and that aureochromes might play an important role in this process. P. tricornutum possesses four genes encoding aureochromes. In this study we confirm the nuclear localisation of the PtAUREO1a, 1b and 2 proteins. Furthermore we studied the physiology of light quality acclimation in genetically transformed P. tricornutum cell lines with reduced expression of the aureochrome 1a gene. The results demonstrate that the AUREO1a protein has a distinct function in light acclimation. However, rather unexpectedly AUREO1a seems to repress high light acclimation which resulted in a state of ‘hyper’ high light acclimation in aureo1a silenced strains. This was indicated by characteristic changes of several photosynthetic parameters, including increased maximum photosynthesis rates, decreased chlorophyll a contents per cell and increased values of non-photochemical quenching in AUREO1a silenced strains compared to wild type cultures. Strikingly, AUREO1a silenced strains exhibited phenotypic differences compared to wild type cells during cultivation under BL as well as under red light (RL) conditions. Therefore, AUREO1a might influence the RL signalling process, suggesting an interaction of AUREO1a with RL perception pathways.