The Effect of Zeaxanthin as the Only Xanthophyll on the Structure and Function of the Photosynthetic Apparatus in Arabidopsis thaliana

In green plants, the xanthophyll carotenoid zeaxanthin is synthesized transiently under conditions of excess light energy and participates in photoprotection. In the Arabidopsis lut2 npq2 double mutant, all xanthophylls were replaced constitutively by zeaxanthin, the only xanthophyll whose synthesis was not impaired. The relative proportions of the different chlorophyll antenna proteins were strongly affected with respect to the wild-type strain. The major antenna, LHCII, did not form trimers, and its abundance was strongly reduced as was CP26, albeit to a lesser extent. In contrast, CP29, CP24, LHCI proteins, and the PSI and PSII core complexes did not undergo major changes. PSII-LHCII supercomplexes were not detectable while the PSI-LHCI supercomplex remained unaffected. The effect of zeaxanthin accumulation on the stability of the different Lhc proteins was uneven: the LHCII proteins from lut2 npq2 had a lower melting temperature as compared with the wild-type complex while LHCI showed increased resistance to heat denaturation. Consistent with the loss of LHCII, light-state 1 to state 2 transitions were suppressed, the photochemical efficiency in limiting light was reduced and photosynthesis was saturated at higher light intensities in lut2 npq2 leaves, resulting in a photosynthetic phenotype resembling that of high light-acclimated leaves. Zeaxanthin functioned in vivo as a light-harvesting accessory pigment in lut2 npq2 chlorophyll antennae. As a whole, the in vivo data are consistent with the results obtained by using recombinant Lhc proteins reconstituted in vitro with purified zeaxanthin. While PSII photoinhibition was similar in wild type and lut2 npq2 exposed to high light at low temperature, the double mutant was much more resistant to photooxidative stress and lipid peroxidation than the wild type. The latter observation is consistent with an antioxidant and lipid protective role of zeaxanthin in vivo

Oil accumulation in the model green alga Chlamydomonas reinhardtii: characterization, variability between common laboratory strains and relationship with starch reserves

International audienceBackground: When cultivated under stress conditions, many microalgae species accumulate both starch and oil (triacylglycerols). The model green microalga Chlamydomonas reinhardtii has recently emerged as a model to test genetic engineering or cultivation strategies aiming at increasing lipid yields for biodiesel production. Blocking starch synthesis has been suggested as a way to boost oil accumulation. Here, we characterize the triacylglycerol (TAG) accumulation process in Chlamydomonas and quantify TAGs in various wild-type and starchless strains. Results: In response to nitrogen deficiency, Chlamydomonas reinhardtii produced TAGs enriched in palmitic, oleic and linoleic acids that accumulated in oil-bodies. Oil synthesis was maximal between 2 and 3 days following nitrogen depletion and reached a plateau around day 5. In the first 48 hours of oil deposition, a~80% reduction in the major plastidial membrane lipids occurred. Upon nitrogen re-supply, mobilization of TAGs started after starch degradation but was completed within 24 hours. Comparison of oil content in five common laboratory strains (CC124, CC125, cw15, CC1690 and 11-32A) revealed a high variability, from 2 μg TAG per million cell in CC124 to 11 μg in 11-32A. Quantification of TAGs on a cell basis in three mutants affected in starch synthesis (cw15sta1-2, cw15sta6 and cw15sta7-1) showed that blocking starch synthesis did not result in TAG over-accumulation compared to their direct progenitor, the arginine auxotroph strain 330. Moreover, no significant correlation was found between cellular oil and starch levels among the twenty wild-type, mutants and complemented strains tested. By contrast, cellular oil content was found to increase steeply with salt concentration in the growth medium. At 100 mM NaCl, oil level similar to nitrogen depletion conditions could be reached in CC124 strain. Conclusion: A reference basis for future genetic studies of oil metabolism in Chlamydomonas is provided. Results highlight the importance of using direct progenitors as control strains when assessing the effect of mutations on oil content. They also suggest the existence in Chlamydomonas of complex interplays between oil synthesis, genetic background and stress conditions. Optimization of such interactions is an alternative to targeted metabolic engineering strategies in the search for high oil yields

Involvement of CDSP 32, a drought-induced thioredoxin, in the response to oxidative stress in potato plants

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The Plastidic 2-Cysteine Peroxiredoxin Is a Target for a Thioredoxin Involved in the Protection of the Photosynthetic Apparatus against Oxidative Damage

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The Arabidopsis Plastidic Methionine Sulfoxide Reductase B Proteins. Sequence and Activity Characteristics, Comparison of the Expression with Plastidic Methionine Sulfoxide Reductase A, and Induction by Photooxidative Stress

Two types of methionine (Met) sulfoxide reductases (Msr) catalyze the reduction of Met sulfoxide (MetSO) back to Met. MsrA, well characterized in plants, exhibits an activity restricted to the Met-S-SO-enantiomer. Recently, a new type of Msr enzyme, called MsrB, has been identified in various organisms and shown to catalytically reduce the R-enantiomer of MetSO. In plants, very little information is available about MsrB and we focused our attention on Arabidopsis (Arabidopsis thaliana) MsrB proteins. Searching Arabidopsis genome databases, we have identified nine open reading frames encoding proteins closely related to MsrB proteins from bacteria and animal cells. We then analyzed the activity and abundance of the two chloroplastic MsrB proteins, MsrB1 and MsrB2. Both enzymes exhibit an absolute R-stereospecificity for MetSO and a higher catalytic efficiency when using protein-bound MetSO as a substrate than when using free MetSO. Interestingly, we observed that MsrB2 is reduced by thioredoxin, whereas MsrB1 is not. This feature of MsrB1 could result from the lack of the catalytical cysteine (Cys) corresponding to Cys-63 in Escherichia coli MsrB that is involved in the regeneration of Cys-117 through the formation of an intramolecular disulfide bridge followed by thioredoxin reduction. We investigated the abundance of plastidial MsrA and B in response to abiotic (water stress, photooxidative treatment) and biotic (rust fungus) stresses and we observed that MsrA and B protein levels increase in response to the photooxidative treatment. The possible role of plastidic MsrB in the tolerance to oxidative damage is discussed

Characterization of Nda2, a Plastoquinone-reducing Type II NAD(P)H Dehydrogenase in Chlamydomonas Chloroplasts

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Heavy metal accumulation by recombinant mammalian metallothionein within Escherichia coli protects against elevated metal exposure.

International audienceMetallothioneins (MTs) are ubiquitous metal-binding, cysteine-rich, small proteins known to provide protection against toxic heavy metals such as cadmium. In an attempt to increase the ability of bacterial cells to accumulate heavy metals, sheep MTII was produced in fusion with the maltose binding protein (MBP) and localized to the cytoplasmic or periplasmic compartments of Escherichia coli. For all metals tested, higher levels of bioaccumulation were measured with strains over-expressing MBP-MT in comparison with control strains. A marked bioaccumulation of Cd, As, Hg and Zn was observed in the strain over-expressing MBP-MT in the cytoplasm, whereas Cu was accumulated to higher levels when MBP-MT was over-expressed in the periplasm. Metal export systems may also play a role in this bioaccumulation. To illustrate this, we over-expressed MBP-MT in the cytoplasm of two mutant strains of E. coli affected in metal export. The first, deficient in the transporter ZntA described to export numerous divalent metal ions, showed increasing quantities of Zn, Cd, Hg and Pb being bioaccumulated. The second, strain LF20012, deficient in As export, showed that As was bioaccumulated in the form of arsenite. Furthermore, high quantities of accumulated metals, chelated by MBP-MT in the cytoplasm, conferred greater metal resistance levels to the cells in the presence of added toxic metals, such as Cd or Hg, while other metals showed toxic effects when the export systems were deficient. The strain over-expressing MBP-MT in the cytoplasm, in combination, with disruption of metal export systems, could be used to develop strategies for bioremediation