Peroxiredoxin Q of Arabidopsis thaliana is attached to the thylakoids and functions in context of photosynthesis.

Peroxiredoxin Q (Prx Q) is one out of 10 peroxiredoxins encoded in the genome of Arabidopsis thaliana, and one out of four that are targeted to plastids. Peroxiredoxin Q functions as a monomeric protein and represents about 0.3% of chloroplast proteins. It attaches to the thylakoid membrane and is detected in preparations enriched in photosystem II complexes. Peroxiredoxin Q decomposes peroxides using thioredoxin as an electron donor with a substrate preference of H(2)O(2) > cumene hydroperoxide >> butyl hydroperoxide >> linoleoyl hydroperoxide and insignificant affinity towards complex phospholipid hydroperoxide. Plants with decreased levels of Prx Q did not have an apparently different phenotype from wildtype at the plant level. However, similar to antisense 2-cysteine (2-Cys) Prx plants [Baier, M. et al. (2000)Plant Physiol., 124, 823-832], Prx Q-deficient plants had a decreased sensitivity to oxidants in a leaf slice test as indicated by chlorophyll a fluorescence measurements. Increased fluorescence ratios of photosystem II to I at 77 K and modified transcript levels of plastid- and nuclear-encoded proteins show that regulatory mechanisms are at work to compensate for the lack of Prx Q. Apparently Prx Q attaches to photosystem II and has a specific function distinct from 2-Cys peroxiredoxin in protecting photosynthesis. Its absence causes metabolic changes that are sensed and trigger appropriate compensatory responses.

The mitochondrial type II peroxiredoxin F is essential for redox homeostasis and root growth of Arabidopsis thaliana under stress.

Peroxiredoxins (Prx) have recently moved into the focus of plant and animal research in the context of development, adaptation, and disease, as they function both in antioxidant defense by reducing a broad range of toxic peroxides and in redox signaling relating to the adjustment of cell redox and antioxidant metabolism. At-PrxII F is one of six type II Prx identified in the genome of Arabidopsis thaliana and the only Prx that is targeted to the plant mitochondrion. Therefore, it might be assumed to have functions similar to the human 2-Cys Prx (PRDX3) and type II Prx (PRDX5) and yeast 1-Cys Prx that likewise have mitochondrial localizations. This paper presents a characterization of PrxII F at the level of subcellular distribution, activity, and reductive regeneration by mitochondrial thioredoxin and glutaredoxin. By employing tDNA insertion mutants of A. thaliana lacking expression of AtprxII F (KO-AtPrxII F), it is shown that under optimal environmental conditions the absence of PrxII F is almost fully compensated for, possibly by increases in activity of mitochondrial ascorbate peroxidase and glutathione-dependent peroxidase. However, a stronger inhibition of root growth in KO-AtPrxII F seedlings as compared with wild type is observed under stress conditions induced by CdCl2 as well as after administration of salicylhydroxamic acid, an inhibitor of cyanide-insensitive respiration. Simultaneously, major changes in the abundance of both nuclear and mitochondria-encoded transcripts were observed. These results assign a principal role to PrxII F in antioxidant defense and possibly redox signaling in plants cells.

Characterization of plastidial thioredoxins from Arabidopsis belonging to the new y-type.

The plant plastidial thioredoxins (Trx) are involved in the light-dependent regulation of many enzymatic activities, owing to their thiol-disulfide interchange activity. Three different types of plastidial Trx have been identified and characterized so far: the m-, f-, and x-types. Recently, a new putative plastidial type, the y-type, was found. In this work the two isoforms of Trx y encoded by the nuclear genome of Arabidopsis (Arabidopsis thaliana) were characterized. The plastidial targeting of Trx y has been established by the expression of a TrxGFP fusion protein. Then both isoforms were produced as recombinant proteins in their putative mature forms and purified to characterize them by a biochemical approach. Their ability to activate two plastidial light-regulated enzymes, NADP-malate dehydrogenase (NADP-MDH) and fructose-1,6-bisphosphatase, was tested. Both Trx y were poor activators of fructose-1,6-bisphosphatase and NADP-MDH; however, a detailed study of the activation of NADP-MDH using site-directed mutants of its regulatory cysteines suggested that Trx y was able to reduce the less negative regulatory disulfide but not the more negative regulatory disulfide. This property probably results from the fact that Trx y has a less negative redox midpoint potential (-337 mV at pH 7.9) than thioredoxins f and m. The y-type Trxs were also the best substrate for the plastidial peroxiredoxin Q. Gene expression analysis showed that Trx y2 was mainly expressed in leaves and induced by light, whereas Trx y1 was mainly expressed in nonphotosynthetic organs, especially in seeds at a stage of major accumulation of storage lipids.

cDNA cloning of 12 subunits of the V-type ATPase from Mesembryanthemum crystallinum and their expression under stress.

The vacuolar-type ATPase (V-ATPase) and the vacuolar H(+)-pyrophosphatase are electrogenic proton pumps at plant endomembranes that create the proton motive force required for secondary activated transport and metabolite accumulation during development and adaptation to a variety of adverse growth conditions. Twelve distinct vacuolar H(+)-ATPase (VHA) subunits are suggested to constitute the functional V-ATPase complex. Starting from the available expressed sequence tag (EST) sequences and by homology screening, the complete set of 12 VHA subunits was cloned as cDNAs from the halophyte Mesembryanthemum crystallinum, vha-A-H, -a,-c, -d and -e. Transcript levels of all 12 VHA subunits as well as of tonoplast pyrophosphatase and P-ATPase were analysed in root and leaf tissue under conditions of osmotic (700 mM mannitol), heat and cold stress, and salinity. Distinct coordinated changes of stress-induced expression were observed for most subunits in roots and leaves, with mostly paralleled changes in transcript levels of all subunits. In some cases, contrasting responses were seen for vha-B and -c transcript amounts.

Divergent light-, ascorbate-, and oxidative stress-dependent regulation of expression of the peroxiredoxin gene family in Arabidopsis.

Peroxiredoxins (prxs) are peroxidases with broad substrate specificity. The seven prx genes expressed in Arabidopsis shoots were analyzed for their expressional response to changing photon fluence rates, oxidative stress, and ascorbate application. The results reveal a highly variable and gene-specific response to reducing and oxidizing conditions. The steady-state transcript amounts of the chloroplast-targeted prxs, namely the two-cysteine (2-Cys) prxs, prx Q and prx II E, decreased upon application of ascorbate. prx Q also responded to peroxides and diamide treatment. prx II B was induced by tertiary butylhydroperoxide, but rather unaffected by ascorbate. The strongest responses were observed for prx II C, which was induced with all treatments. The two Arabidopsis 2-Cys Prxs and four Prx II proteins were expressed heterologously in Escherichia coli. In an in vitro test system, they all showed peroxidase activity, but could be distinguished by their ability to accept dithiothreitol and thioredoxin as electron donor in the regeneration reaction. The midpoint redox potentials (E(m)') of Prx II B, Prx II C, and Prx II E were around -290 mV and, thus, less negative than E(m)' of Prx II F, 2-Cys Prx A, and 2-Cys Prx B (-307 to -322 mV). The data characterize expression and function of the mitochondrial Prx II F and the chloroplast Prx II E for the first time, to our knowledge. Antibodies directed against 2-Cys Prx and Prx II C showed a slight up-regulation of Prx II protein in strong light and of 2-Cys Prx upon transfer both to high and low light. The results are discussed in context with the subcellular localization of the Prx gene products.