Induction Of Non-photochemical Quenching Research Articles

Interactions between the Photosystem II Subunit PsbS and Xanthophylls Studied in Vivo and in Vitro

The photosystem II subunit PsbS is essential for excess energy dissipation (qE); however, both lutein and zeaxanthin are needed for its full activation. Based on previous work, two models can be proposed in which PsbS is either 1) the gene product where the quenching activity is located or 2) a proton-sensing trigger that activates the quencher molecules. The first hypothesis requires xanthophyll binding to two PsbS-binding sites, each activated by the protonation of a dicyclohexylcarbodiimide-binding lumen-exposed glutamic acid residue. To assess the existence and properties of these xanthophyll-binding sites, PsbS point mutants on each of the two Glu residues PsbS E122Q and PsbS E226Q were crossed with the npq1/npq4 and lut2/npq4 mutants lacking zeaxanthin and lutein, respectively. Double mutants E122Q/npq1 and E226Q/npq1 had no qE, whereas E122Q/lut2 and E226Q/lut2 showed a strong qE reduction with respect to both lut2 and single glutamate mutants. These findings exclude a specific interaction between lutein or zeaxanthin and a dicyclohexylcarbodiimide-binding site and suggest that the dependence of nonphotochemical quenching on xanthophyll composition is not due to pigment binding to PsbS. To verify, in vitro, the capacity of xanthophylls to bind PsbS, we have produced recombinant PsbS refolded with purified pigments and shown that Raman signals, previously attributed to PsbS-zeaxanthin interactions, are in fact due to xanthophyll aggregation. We conclude that the xanthophyll dependence of qE is not due to PsbS but to other pigment-binding proteins, probably of the Lhcb type.

Involvement of phycobilisome diffusion in energy quenching in cyanobacteria.

Nonphotochemical quenching (NPQ) of excitation energy is a well-established phenomenon in green plants, where it serves to protect the photosynthetic apparatus from photodamage under excess illumination. The induction of NPQ involves a change in the function of the light-harvesting apparatus, with the formation of quenching centers that convert excitation energy into heat. Recently, a comparable phenomenon was demonstrated in cyanobacteria grown under iron-starvation. Under these conditions, an additional integral membrane chlorophyll-protein, IsiA, is synthesized, and it is therefore likely that IsiA is required for NPQ in cyanobacteria. We have previously used fluorescence recovery after photobleaching to show that phycobilisomes diffuse rapidly on the membrane surface, but are immobilized when cells are immersed in high-osmotic strength buffers, apparently because the interaction between phycobilisomes and reaction centers is stabilized. Here, we show that when cells of the cyanobacterium Synechocystis sp. PCC 6803 subjected to prolonged iron-deprivation are immersed in 1 m phosphate buffer, NPQ can still be induced as normal by high light. However, the formation of the quenched state is irreversible under these conditions, suggesting that it involves the coupling of free phycobilisomes to an integral-membrane complex, an interaction that is stabilized by 1 m phosphate. Fluorescence spectra are consistent with this idea. Fluorescence recovery after photobleaching measurements confirm that the induction of NPQ in the presence of 1 m phosphate is accompanied by immobilization of the phycobilisomes. We propose as a working hypothesis that a major component of the fluorescence quenching observed in iron-starved cyanobacteria arises from the coupling of free phycobilisomes to IsiA.

Regulation of photochemical activity in cultured symbiotic dinoflagellates under nitrate limitation and deprivation

The effects of nitrate limitation and nitrate starvation on the photochemical efficiency of photosystem II (Fv/F m) were examined in batch cultures of two species of symbiotic dinoflagellates, Symbiodinium kawagutii and S. pilosum. F v/F m values were determined along growth curves and show that the F v/F m values are negatively correlated with external nitrogen concentrations in cultures of both species. Changes in growth irradiances in the batch cultures due to increments of the cell densities were estimated S. kawagutii cultures showed a negative correlation between F v/F m and growth irradiance. These results indicate that F v/F m is dependent on the light history of the cultures and on the individual sensitivity of each species, and independent of their nutrient status. Nitrate starvation was analyzed by measuring changes in the quantum yield of fluorescence (Fv/F m), electron transport rate (ETR) and non-photochemical quenching (NPQ) at five time points along the growth curves under three conditions: control (C), without nitrogen (N−), and with ammonia (N+) as a nitrogen source sufficient to meet daily nitrogen requirements. Cells collected during the exponential growth phase and exposed to N− and N+ showed significant reductions in their maximum ETR relative to controls (20% in S. pilosum and 40% in S. kawagutii). The loss of electron transport capacity is consistent with a sink limitation rather than the result of nitrogen starvation. Under nitrate-starvation, the induction of NPQ resulted in effective protection against photosystem II damage in S. pilosum. In contrast, S. kawagutii cells failed to induced NPQ resulting in a concomitant increase in the excitation pressure over photosystem II leading to damage. Collectively the data indicate that F v/F m is not a robust indicator of nitrogen limitation in symbiotic dinoflagellates and that protection against photosystem II damage under sink limitations, is largely dependent on the differential capacities of each species to induce NPQ.

Significance of the lipid phase in the dynamics and functions of the xanthophyll cycle as revealed by PsbS overexpression in tobacco and in-vitro de-epoxidation in monogalactosyldiacylglycerol micelles.

The dynamics of the xanthophyll cycle relative to non-photochemical quenching (NPQ) were examined in tobacco plants overexpressing violaxanthin de-epoxidase (VDE), PsbS and PsbS+VDE for effects on NPQ and violaxanthin (V) de-epoxidation over a range of light intensities. Induction of de-epoxidation and NPQ increased in overexpressed VDE and PsbS plants, respectively. Surprisingly, under low light, overexpressing PsbS enhanced de-epoxidation in addition to NPQ. The effect was hypothesized as due to PsbS binding zeaxanthin (Z) or inducing the binding of Z within the quenching complex, thus shifting the equilibrium toward higher de-epoxidation states. Studies in model systems show that Z can stereospecifically inhibit VDE activity against violaxanthin. This effect, observed under conditions of limiting lipid concentration, was interpreted as product feedback inhibition. These results support the hypothesis that the capacity of the thylakoid lipid phase for xanthophylls is limited and modulates xanthophyll-cycle activity, in conjunction with the release of V and binding of Z by pigment-binding proteins. These modulating factors are incorporated into a lipid-matrix model that has elements of a signal transduction system wherein the light-generated protons are the signal, VDE the signal receptor, Z the secondary messenger, the lipid phase the transduction network, and Z-binding proteins the targets.

Strategies providing success in a variable habitat: III. Dynamic control of photosynthesis in Cladophora glomerata

AbstractDiurnal patterns of photosynthesis were studied in July and April populations of Cladophora glomerata (L.) Kütz. from open and from shaded sites. Summer samples exposed to full sunlight showed decreased efficiency of open photosystem II at noon, and only slight differences were found between samples that had grown at open or at shaded sites. Electron transport rate was limited at highest fluence rates in shade plants, and non‐photochemical quenching (NPQ) revealed faster regulation in samples from open sites. Daily course of de‐epoxidation was not linearly correlated with the course of NPQ. The comparison of samples from open and from shaded sites revealed a higher capacity of thermal energy dissipation and an increase in the total amount of xanthophyll‐cycle pigments (21%) in samples from open sites. In April, down‐regulation of the efficiency of open photosystem II was related to lower water temperature, and hence, increased excitation pressure. In April the pool size of xanthophyll‐cycle pigments was increased by 21% in comparison with summer and suggested higher levels of thermal energy dissipation via de‐epoxidized xanthophylls. In both, summer and spring the amount of xanthophyll‐cycle pigments was 20% higher in samples from open sites. Acclimation of C. glomerata to growth light conditions was further shown by experimental induction of NPQ, indicating NPQ increases of 23%, and increases of 77% in the reversible component of NPQ in open site samples. The effect of temperature on photosynthetic rate was non‐linear, and different optimum temperatures of electron transport rate and oxygen evolution were exhibited.

Genetic manipulation of carotenoid biosynthesis and photoprotection.

There are multiple complementary and redundant mechanisms to provide protection against photo-oxidative damage, including non-photochemical quenching (NPQ). NPQ dissipates excess excitation energy as heat by using xanthophylls in combination with changes to the light-harvesting complex (LHC) antenna. The xanthophylls are oxygenated carotenoids that in addition to contributing to NPQ can quench singlet or triplet chlorophyll and are necessary for the assembly and stability of the antenna. We have genetically manipulated the expression of the epsilon-cyclase and beta-carotene hydroxylase carotenoid biosynthetic enzymes in Arabidopsis thaliana. The epsilon-cyclase overexpression confirmed that lut2 (lutein deficient) is a mutation in the epsilon-cyclase gene and demonstrated that lutein content can be altered at the level of mRNA abundance with levels ranging from 0 to 180% of wild-type. Also, it is clear that lutein affects the induction and extent of NPQ. The deleterious effects of lutein deficiency on NPQ in Arabidopsis and Chlamydomonas are additive, no matter what the genetic background, whether npq1 (zeaxanthin deficient), aba1 or antisense beta-hydroxylase (xanthophyll cycle pool decreased). Additionally, increasing lutein content causes a marginal, but significant, increase in the rate of induction of NPQ despite a reduction in the xanthophyll cycle pool size.