Redox Strategies for Crop Improvement.

Page 1 of 63

Antioxidants & Redox Signaling Redox Strategies for Crop Improvement (doi: 10.1089/ars.2014.6033) This article has been peer-reviewed and accepted for publication, but has yet to undergo copyediting and proof correction. The final published version may differ from this proof.

1 Forum Review Article Redox Strategies for Crop Improvement Pavel Kerchev1,2a, Barbara De Smet1,2,3,4,5a, Cezary Waszczak1,2,3,4,5b, Joris Messens3,4,5 and Frank Van Breusegem1,2

1

Department of Plant Systems Biology, VIB, B–9052 Ghent, Belgium.

2

Department of Plant Biotechnology and Bioinformatics, Ghent University, B–9052 Ghent, Belgium.

3

Structural Biology Research Center, VIB, B-1050 Brussels, Belgium.

4

Brussels Center for Redox Biology, B-1050 Brussel, Belgium.

5

Structural Biology Brussels, Vrije Universiteit Brussel, B-1050 Brussel, Belgium.

a

equal contribution current address: Division of Plant Biology, Department of Biosciences, University of Helsinki, FI-00014 Helsinki, Finland b

Corresponding author: Frank Van Breusegem Address: Technologiepark 927, B-9052 Gent E-mail: [email protected] Tel. 32 9 331 39 20 Fax 32 9 331 38 09

Running head: Redox strategies for crop improvement

Word count: 6953 Reference numbers: 197 Number of greyscale illustrations: 7 Number of color illustrations: 1

1

Page 2 of 63


2 Abstract

Significance: Recently, the agro-biotech industry has been driven by overcoming the limitations imposed by fluctuating environmental stress conditions on crop productivity. A common theme among (a)biotic stresses is the perturbation of the redox homeostasis. Recent Advances: As a strategy to engineer stress tolerant crops, many approaches have been centered on restricting the negative impact of reactive oxygen species (ROS) accumulation. Critical issues: Here, we discuss the scientific background of the existing redox-based strategies to improve crop performance and quality. In this respect, a special focus goes to summarizing the current patent landscape, because this aspect is very often ignored, despite constituting the forefront of applied research. Future Directions: The current increased understanding of ROS acting as signaling molecules has opened new avenues to exploit redox biology for crop improvement required for sustainable food security.

2

Page 3 of 63


3

Introduction Plant metabolism comprises a plethora of redox-mediated reactions that are ultimately involved in nearly every aspect of plant growth, development, and defense (12, 25, 140). In order to maintain an optimal redox homeostasis, plants employ a wide spectrum of enzymatic and non-enzymatic antioxidants (48). Some of them, such as ascorbic acid and alphatocopherol, are also essential elements of the human diet and their chronic deficiencies can lead to severe health problems (55). During the Age of Discovery (15th – 16th century), sea explorers have been plagued by the lack of ascorbic acid, and millions faced slow death, which could have been prevented by fresh plant-based food (88). The plant redox systems hence played a crucial role in expanding our geographical knowledge and the boundaries of the unknown. Several centuries later, we are looking specifically at the redox components and mechanisms that orchestrate plant growth and development in an effort to find answers and solutions, which are urgently needed for our voyage into the uncertain future in which population growth and climate change will inevitably threaten food security. Here, we review plant redox systems that have already served or might be exploited in the future as a basis to engineer crops with enhanced resistance to adverse environmental conditions, to develop efficient strategies for weed control, and to provide means to increase food security, while combating nutritional deficiencies. We primarily focus on strategies that are already used in agriculture or have the potential to be adopted in the near future, and discuss the science behind these technologies.

Generation of Reactive Oxygen Species in the Plant Cell Today, the majority of living organisms use oxygen (O2) as a final electron acceptor to sustain their energy levels. This requirement is a direct consequence of the accumulation of oxygen in the primordial atmosphere mediated by photosynthetic organisms able to split water into O2,

3

Page 4 of 63


4 electrons and protons (39). The reactivity of O2, however, entails the formation of toxic O2 derivatives as by-products of normal metabolism (Fig. 1). These by-products, i.e. hydrogen peroxide (H2O2), superoxide radical (O2•-), hydroxyl radical (HO•), and singlet oxygen (1O2), are commonly referred to as reactive oxygen species (ROS) and their levels have to be tightly controlled in order to avoid excessive oxidative damage of cellular constituents (Fig. 3). ROS production in plant cells reflects their functional specialization and is largely dependent on the fluctuating external environment. In contrast to animal cells, in which mitochondria are the main sites of ROS production, plant mitochondria are just one of the cellular compartments that contribute to the overall ROS abundance (Fig. 1) (123). In addition, chloroplasts, the plant organelles that harbor the photosynthetic machinery and combine O2 production with active electron transport chains, are the most important source of ROS generation in photosynthesizing tissues (4). Peroxisomal ROS production is also indirectly linked to photosynthesis via the photorespiratory pathway initiated by the enzymatic promiscuity of RuBisCO (16). Photorespiratory reactions recycle toxic 2-phosphoglycolate resulting from the oxygenation reaction of RuBisCO via a series of enzymatic steps dispersed between chloroplasts, peroxisomes and mitochondria (Fig. 4) (11). The peroxisomal glycolate oxidase generates substantial amounts of H2O2 that under conditions favoring photorespiration pose a significant load on the cellular redox poise (129). The enzymatic activities of several ROSgenerating enzymes, such as plasma membrane-localized NADPH oxidases, and apoplastic peroxidases and polyamine oxidases, are another important determinant of ROS homeostasis in plants (Fig. 1) (2). The presence of NADPH oxidase homologs both in animals (NOX) and plants (Rboh) indicates a common evolutionary origin for these superoxide-producing enzymes. NOX activity in human blood phagocytic cells is crucial for eliminating invading pathogens, whereas plant NADPH oxidases are similarly involved in ROS production in response to pathogens (Fig. 2). Apart from their involvement in defense, different NOX and

4

Page 5 of 63


5 Rboh family members have been implicated in diverse developmental and signal transduction events mediated through ROS production (185). The Yin-Yang of ROS Homeostasis: Maintaining the Balance between Signaling and Oxidative Damage The accumulation of ROS has been demonstrated for multiple stress conditions like wounding (114), excess light (45), pathogen infection (79), low CO2 availability (108), but also during developmental processes like root growth (43), gravitropism (60), and pollen tube growth (61). Furthermore, the production of ROS is an intrinsic feature of multiple cellular processes like mitochondrial respiration, photosynthesis, photorespiration and β-fatty acid oxidation (123). Under conditions favoring photorespiration, for example, the rate of peroxisomal H2O2 production in photosynthesizing tissues exceeds the contribution of chloroplasts and mitochondria to the overall ROS levels (44). The peroxisomes of germinating seeds, on the other hand, are a significant source of H2O2 generated during fatty acid β-oxidation of mobilized storage lipids (8). Turnover of fatty acids that can be used as an energy source during developmental and also stress-induced senescence have been suggested to enhance the rate of fatty acid β-oxidation, but given the complexity of the senescence process, the impact of fatty acid β-oxidation on ROS homeostasis remains to be deconvoluted (20, 192). Given the central role of lipid metabolism, however, it might constitute a potentially beneficial avenue for crop improvement. Although frequently reported, obtaining an accurate estimation of ROS levels is inherently limited, and such results should be regarded with some caution (128). Often the low methodological resolution obscures the contribution of ROS within individual cellular compartments. So, it is hard to distinguish between different ROS species, and we cannot adequately capture the transient nature of their accumulation (128). To circumvent these

5

Page 6 of 63


6 limitations, biochemical (144) and molecular markers (46) have been developed with the use of established genetic models exhibiting elevated ROS levels. However, the use of single marker genes does not seem to provide a good proxy for the site and nature of ROS production, therefore the use of “ROS signatures” has recently been proposed (138). Plants lacking peroxisomal catalase (cat2) (Fig. 5), for example, have been used to deconvolute the transcriptional response triggered by peroxisomal H2O2 (170). In a similar approach, the Arabidopsis

fluorescent

(flu)

mutant,

which

accumulates

the

photosensitizer

protochlorophyllide, was used to extract a 1O2-related transcriptional (113) and metabolic signature (127). Obviously, cautionary interpretation is necessary as these responses can be influenced by timing, interconversion of ROS and site of production (68, 143, 155). Additionally, the ability of ROS to differentially oxidize lipids to specific hydroxy fatty acids was exploited to discriminate between peroxidation reactions mediated by individual ROS forms (162). The attack of 1O2 on carotenoids results in the formation of oxidation products, such as β-cyclocitral, which might not only serve as indicators of ROS accumulation, but also act as potent signaling components (130). Oxidation of amino acid residues by ROS serves as a crucial factor governing the stability and function of both metabolic enzymes and signaling proteins (57, 142). Thus far, the best-documented evidences of such post-translational modifications are those involving oxidation of cysteine and methionine residues (Fig. 6). In both cases, the targets of oxidation are sulfur atoms found in sulfhydryl (R-SH) and thioether (R-S-R’) groups of cysteines and methionines, respectively (30). Methionine sulfoxide (Met-SO), the oxidation product of methionine, is relatively stable and rarely undergoes further oxidation to methionine sulfone (Fig. 6) (30). In contrast, the initial oxidation product of cysteine, sulfenic acid (Cys-SOH), is highly unstable and, unless stabilized within the protein environment, can be further oxidized (Fig. 6) either reversibly to sulfinic acid (Cys-SO2H) or irreversibly to sulfonic acid (Cys-

6

Page 7 of 63


7 SO3H) (31, 137). The reduction of sulfinic acid back to cysteine has been debated, but the existance of an enzyme, sulfiredoxin (Srx), catalyzing this reaction in plants, supports this notion (135). The only two known AtSrx substrates (chloroplast 2-Cys Peroxiredoxins and mitochondrial PrxIIF), however, question the general occurrence of this process (56, 135). Alternatively, sulfenic acid might react with free protein thiols to form intra- or intermolecular disulfide bonds or with low molecular weight thiols like glutathione (GSH) leading to cysteine-S-glutathionylation (Fig. 6) (137, 169). S-glutathionylation has long been believed to serve as a mechanism protecting active cysteine residues from overoxidation and permanent protein damage. Only recently, S-glutathionylation gained recognition for its role in redox signaling (195). S-glutathionylation, formation of intra-/intermolecular disulfides, and methionine sulfoxide are readily reversible and their redox status is controlled by glutaredoxins (Grx), thioredoxins (Trx), and methionine sulfoxide reductases (Msr), respectively (Fig. 6) (30, 137). Apart from methionine and cysteine residues, side chains of other amino acids, such as proline, histidine, arginine, lysine, and threonine, might also be subjected to oxidative modifications leading to the formation of carbonyl derivatives (Fig. 6). Protein carbonylation is an irreversible process caused by hydroxyl radicals (13) generated during metal-catalyzed cleavage of H2O2 (186). Formation of carbonyl groups is widely recognized as a marker for protein oxidative damage and is mainly associated with their proteolytic degradation (58, 110, 179).

Redox Regulation of Plant Immunity Upon recognition of pathogens, plants trigger a broad spectrum of defense processes, which have been reviewed extensively, and include the hypersensitive response (HR) caused by ROS accumulation. The initial oxidative burst following pathogen infection has been

7

Page 8 of 63


8 largely seen as mediated by class III cell wall peroxidases and membrane-bound NADPH oxidases (Respiratory burst oxidase homologs, Rbohs) generating H2O2 and superoxide radicals, respectively (29, 161). The precise mechanism of the peroxidase-dependent oxidative burst is far from elucidated and is likely to require reductants such as phenolic compounds and NADH (O'Brien et al., 2012). Upon infection, the ROS profile is governed by the life style of the pathogen. Avirulent species induce a rapid, transient ROS formation followed by a sustained, higher amplitude of oxidative burst associated with HR and defense (91, 107). In contrast, during virulent and symbiotic interactions, only the initial ROS formation is detected (91). Plants with compromised Rbohs and apoplastic peroxidase activities display altered resistance to pathogens, exemplifying the important role of ROS accumulation during plantpathogen interaction (6, 194). One member of the Rboh protein family, RbohD, is further required for triggering and propagating a systemic signal initiated by various biotic and abiotic stimuli (98). The effective long-distance communication depends on apoplastic ROS accumulation along the signal path, since the application of the NADPH oxidase inhibitor diphenylene iodonium (DPI) is sufficient to block further spread of the signal. Interestingly, rbohd mutants are susceptible to green peach aphid (Myzus persicae) attack, implying a functional role for RbohD and systemic ROS signaling during plant-aphid interactions (98). Other invasive species, such as the parasitic nematode Heterodera schachtii, similarly trigger Rboh-dependent ROS production (145). Infected plants lacking RbohD and RbohF display increased cell death lesions in comparison to the wild type. Despite the primary role of Rbohs in ROS generation during plant defense, emerging evidence points toward alternative ROS sources that might be activated under such scenarios and function independently from Rbohs. Peroxisomal glycolate oxidases (GOXs), for example, modulate the outcome of the non-host resistance in tobacco and Arabidopsis, with

8

Page 9 of 63


9 plants lacking GOX activity being more susceptible to pathogens (136). Moreover, silencing of RbohD within the gox mutant background indicated independent roles in plant defense, because lack of RbohD had largely no additive effect on bacterial growth in the respective double mutants (136). The highly reactive nature of ROS is very capable to impose a direct, cytotoxic effect on invading pathogens (121). For example, the virulence of Ustillago maydis (the causing agent of corn snut) depends on its ability to detoxify plant-generated ROS (99). An U. maydis strain lacking an ortholog of the S. cerevisiae YAP1 (Yeast AP-1-like) transcription factor, which controls the response toward oxidative stress conditions (33, 34), is more susceptible to H2O2 produced by NADPH oxidases during oxidative burst (99). In contrast to biotrophic pathogens, necrotrophs utilize dead plant cells as source of nutrients and often induce ROS production during the infection process to stimulate the hypersensitive response-related cell death (49). Salicylic acid (SA) has a prominent role in plant-pathogen responses through a coordinated action of the NONEXPRESSER OF PR GENES 1 (NPR1) transcriptional coactivator and its interacting transcription factors from the TGA family (Fig. 7). The molecular properties of both NPR1 and its TGA-family interactors depend on changes in the cellular redox status that occur upon pathogen infection (104, 153). Under non-stressed conditions, NPR1 oligomers are stabilized by intramolecular disulfide bonds involving two cysteine residues (Cys82 and Cys216), and localized to the cytoplasm (Fig. 7). SA triggers fluctuations in the cellular redox status which lead to a Trxh3/h5-dependent reduction of these disulfides, NPR1 monomerization, and subsequent nuclear import (104, 153). This process is in direct competition with the oligomerization and disulfide formation induced by the S-nitrosylation of Cys156 (Fig. 7). The interplay between these two modifications tightly controls NPR1dependent signaling (153). Upon nuclear import, NPR1 interacts with TGA basic leucine-

9

Page 10 of 63


10 zipper transcription factors to promote the transcription of pathogenesis related (PR) genes. Clade 2 TGA transcription factors (TGA2/5/6) act as constitutive repressors of PR transcription. Binding of NPR1 to TGA2 results in co-activation of transcription, and depends on NPR1 Cys521 and Cys529 required for binding of SA via the transition metal copper (190). The interaction of NPR1 with clade 1 TGA transcription factors (TGA1/4) relies on their redox status. When oxidized, TGA1 and/or TGA4 form an intramolecular disulfide bridge (Cys260-Cys266) that hinders binding. Reduction of this disulfide stimulates the interaction with NPR1, and subsequent binding to the as-1 element for the transcription of PR genes (35). Recent data indicate that the two remaining TGA1 cysteine residues, Cys172 and Cys287, are also involved in regulatory disulfide formation (83). Furthermore, all four Cys residues were found to undergo S-nitrosylation/S-glutathionylation after treatment with GSNO (Fig. 7). These modifications serve to protect the Cys residues from disulfide formation and increase the DNA-binding activity of TGA1 (83). The recognized role of ROS accumulation for an effective immune response was harnessed in a technology that provides resistance to the economically important pathogen Sclerotinia sclerotiorum (15 - WO199904012). Transgenic sunflower, canola, and soybean plants overexpressing the H2O2-producing enzyme oxalate oxidase displayed an improved performance after infection under field conditions (15 - WO199904012). Moreover, introducing the transgenic trait into conventional pathogen-tolerant cultivars had a synergistic effect. Boosting H2O2 levels by introducing glucose oxidase (GO) from Aspergillus sp. also positively affected plant immunity and led to a broad-spectrum resistance against various fungal and bacterial pathogens (76 - WO1995014784, 189). For example, potato tubers with enhanced GO levels displayed tolerance against Erwinia carotovora, an economically important post-harvest pathogen causing potato soft rot (76 - WO1995014784, 189).

10

Page 11 of 63


11 Redox Homeostasis in Plant Growth and Development Until recently, developmental processes have been largely seen as orchestrated by hormonal cross-talk and transcriptional regulators (73, 174). The redox control of plant growth and development, however, is now being increasingly recognized in the light of ample evidence for an extensive network of interactions between ROS and hormonal (e.g. auxin, gibberellin, and abscisic acid) signaling pathways (1, 183). For example, the complex machinery involved in auxin transport, perception, and biosynthesis is influenced by the cellular redox homeostasis (158). One of the most strongly induced transcripts after H2O2 treatment of Arabidopsis plants encodes an UDP-Glucosyltransferase 74E2 (UGT74E2; 171), acting on indole-3-butyric acid (160, 167 - WO2008155139). Endogenous overexpression of UGTE74E2 in Arabidopsis altered the endogenous auxin pools and led to distinct morphological changes accompanied by enhanced drought and salt tolerance (160, 167 WO2008155139). The organ forming capacity of the shoot and root meristem depends on continuous cell proliferation that is affected by fluctuations of the redox potential (43, 86, 164). The root quiescent center (QC) is maintained in a highly oxidized state that might be linked to the cell cycle arrest observed in the stem cells (59, 84). Moreover, high levels of the lipid peroxidation product malondialdehyde were detected in the meristematic tissues, implying increased rates of ROS formation (141). Active rearrangements of the intracellular GSH pools, a major water-soluble antioxidant, have been observed during cell cycle progression, with GSH migrating to the nucleus at the G1 stage and oxidizing the cytosol (177, 178). The pivotal importance of GSH in plant development was further demonstrated by the inability of the GSH biosynthetic mutant root meristemless (rml) to maintain cell division (175). The low levels of GSH in the rml mutant (less than 5% of the wild-type content) could be mimicked by inhibiting GSH biosynthesis with buthione sulphoximine (BSO), which arrests root growth as

11

Page 12 of 63


12 well. The crucial role of redox homeostasis maintainance for proper development is also evident in Arabidopsis mutants lacking NADPH-thioredoxin reductases (NTRs), which display stunted growth phenotypes similar to those observed upon perturbation of GSH biosynthesis (132). These effects are likely to be partially mediated through the perturbation of polar auxin transport, since GSH depletion negatively impacts various auxin transporters (70). Supporting this view is the fact that the triple mutant rml ntra ntrb, impaired in GSH biosynthesis and thioredoxin reductase activity, phenotypically mimics auxin transport mutants (10). The agriculturally important processes of bud dormancy and bud release are also predominantly driven by redox mechanisms (excellently reviewed by 25). Synchronization of bud burst is of particular importance for economically relevant perennial plants, such as grapevine and kiwi, because it ensures uniform flowering and predictable harvesting times. Current agricultural practices applied to simultaneously break bud dormancy include physical and chemical treatments, such as cyanamide (NH2CN) (53). Transcriptional responses triggered by NH2CN suggest that it might exert its action via the perturbation of the redox homeostasis, since numerous NH2CN-induced genes, such as glutathione-S-transferases (GSTs), catalase, superoxide dismutase (SOD), ascorbate peroxidase, and glutathione peroxidase, are also responsive to H2O2 (53). Various transcription factors, important for plant development, are regulated via redox mechanisms (38). PERIANTHIA (PAN/TGA8), a member of the TGA family transcription factors, controls floral development via an interaction with ROXY1, a floral monothiol glutaredoxin required for petal development (80). Class II, III, and IV members of the plantspecific homeodomain-leucine zipper (HD-ZIP) transcription factor family modulate apical embryo development, meristem function, and epidermal cell fate through the formation of intermolecular disulfides impairing their DNA-binding activity (24, 163). The TCP

12

Page 13 of 63


13 transcription factors implemented in the control of various developmental processes contain a highly conserved cysteine residue within their DNA-binding/dimerization domain that mediates the oxidative inhibition of DNA-binding via the formation of intramolecular disulfide bonds. Treatments with H2O2 and oxidized glutathione (GSSG) inhibit binding of TCP15 to DNA (176). Given the tightly regulated redox homeostasis required for plant development, it is not surprising that external treatments resulting in redox perturbations might trigger distinct morphological changes. Both GSH and its biosynthetic inhibitor, BSO, modulate the flowering time of Arabidopsis (111 - WO2001080638, 112). Similar effects were achieved by using H2O2 and the redox regulators KCN and NaN3 (111 - WO2001080638). Moreover, ROS has been shown to regulate cell elongation (43, 105). In cotton (Gossypium hirsutum), fiber length is an important quality parameter for fiber production that can be modified by altering ROS homeostasis. Exogenous H2O2 and Ca2+ starvation are crucial determinants of early cotton fiber elongation, which is also fine-tuned by the ROS-dependent calcium sensor GhCaM7 (156). The redox requirement of maize anther development has been exploited as a means to control archesporial cell abundance. By lowering the partial pressure of O2 or exposing developing anthers to reducing agents, the inventors achieved a higher pollen production. In contrast, oxidizing conditions led to a lower pollen production or to male sterile plants. Moreover, this approach might be used for the generation of hybrid monocot plants (64, 65 - WO2013123051). A beneficial impact of ROS signaling on plant growth and development is exemplified by the effect of a class of pesticides called strobilurins. First introduced on the market as fungicides, they quickly attracted attention due to the significantly higher yield observed in strobilurin-sprayed crops that could not be attributed to their fungicidal properties related to the inhibition of fungal mitochondrial electron transport at the level of Complex III (9). Even

13

Page 14 of 63


14 though the exact molecular mechanisms triggered by strobilurins are not entirely elucidated, a transient inhibition of plant mitochondria is believed to alter the cellular redox homeostasis, which causes a long-lasting positive impact on growth (14, 36). Examples of strobilurin compounds in commercially available products that combine both fungicidal and yieldenhancing properties are pyraclostrobin and kresoxim-methyl.

Engineering Stress Tolerance in Plants by Reducing Oxidative Stress Prolonged drought, heat waves, suboptimal temperatures, and saline soils are among the common environmental scenarios that negatively affect crop yield. Such suboptimal conditions often lead to over-reduction of the chloroplast electron transport chain, resulting in excessive ROS formation (4). Singlet oxygen is typically produced at photosystem II (PSII) following energy transfer from excited chlorophyll molecules to the ground state triplet oxygen (3O2) (72), whereas PSI contributes to the generation of O2•- via the Mehler reaction (5). Excessive superoxide levels in the chloroplasts are controlled by iron (Fe) and copper/zinc (Cu/Zn) SOD isoforms. Cu/Zn SOD also resides in the cytosol and peroxisomes, whereas manganese (Mn) SOD can be found in both mitochondria and peroxisomes (3). The crucial antioxidant role of SOD in dismutating highly reactive O2•- into O2 and H2O2 has attracted a significant attention as a tool to restrict oxidative damage during stress conditions (17 - WO199002804, 148, 168, 181). Targeting MnSOD, otherwise localized in the mitochondria, to the chloroplasts in rice has resulted in an overall increase of SOD activity and an improved performance under cold and paraquat-induced stress (103). Stacking of multiple antioxidant enzymes has also been used to engineer transgenic plants with superior abilities to withstand stress conditions. Simultaneous overexpression of ascorbate peroxidase (APX), dehydroascorbate reductase (DHAR), and Cu/Zn SOD led to an increased survival under multiple abiotic stresses (77). Moreover, driving APX and SOD under the control of the oxidative stress-inducible promoter of the sweet potato peroxidase

14

Page 15 of 63


15 (pSWPA2) resulted in the conditional activation of the transgenes under ROS-promoting scenarios such as cold, heat, and salt stress (74 - WO2006054815). Similarly, overexpression of Cu/Zn SOD, cytosolic APX, or their combination, conferred higher tolerance to in vitro salt stress in plums (37) and to drought conditions in tobacco (41). During the evolution of multicellular organisms, the O2 environment has served as a selective force, predominantly retaining O2-resistant cellular components (39, 150). Nevertheless, certain O2-sensitive elements such as iron-sulfur (Fe-S) clusters are still present as building blocks in plant proteins (168). One such example is ferredoxin (Fdx), the major electron carrier protein in chloroplasts, which is reduced at the level of PSI by the electron flow via the chloroplast electron transport chain (184). In a reaction catalyzed by ferredoxinNADP+ reductase (FNR), the reducing equivalent carried by Fdx is transferred to NADPH, which is required for numerous biosynthetic processes, including CO2 assimilation, among others. Other Fdx-dependent enzymes involved in sulfur and nitrogen assimilation, and amino acid and fatty acid synthesis also rely on reducing equivalents carried by Fdx (54). Importantly, light activation of some key enzymes of the Calvin cycle is mediated via the reduction of regulatory disulfide bonds by thioredoxin/ferredoxin-thioredoxin reductase and requires a flow of reducing equivalents through Fdx (142). Despite its crucial role, Fdx integrity is highly vulnerable and its activity quickly declines under adverse environmental conditions in all photosynthetic species, including microorganisms (159). In contrast, certain eukaryotic algae and all prokaryotes can counteract the negative effects of Fdx depletion by inducing the expression of alternative electron carriers containing flavin mononucleotide (42). These carriers named flavodoxins, however, are not present in the genome of higher plants (197). Despite their absence, many Fdx-dependent plant enzymes can accept electrons from flavodoxin in vitro, reflecting their common ancestral origin. Based on these evolutionary conserved properties, Palatnik et al. (116) designed transgenic plants with enhanced tolerance

15

Page 16 of 63


16 to oxidative stress-promoting conditions. Expression of chloroplast-targeted cyanobacterial flavodoxin in tobacco was sufficient to compensate for the Fdx decline and to provide protection against adverse environmental conditions, whereas cytosolic expression did not have any beneficial effects (116 - WO2003035881, 159). Plants overexpressing flavodoxin in the chloroplasts sustained predominantly reduced ascorbate and GSH pools under stress, and displayed improved performance under heat, cold, drought, and UV radiation. Interestingly, the transgenic plants also accumulated more biomass in the field in comparison to control plants (116 - WO2003035881). Overexpression of flavodoxin has direct implications for disease resistance and was shown to alleviate symptoms of fungal, bacterial, and viral attacks (116 - WO2003035881). Furthermore, bacterial flavodoxin prevented the decline of nitrogen fixation in nodules of Medicago truncatula (32). Since nodules are highly susceptible to drought and high salinity, flavodoxin overexpression might serve as a way to sustain legume symbiotic performance under unfavorable conditions. The broader agricultural application of flavodoxin was also shown in the important crop species rice, corn, alfalfa, sugarcane, cotton, canola, and wheat (134 - WO2013150402). The transgenic plants displayed improved traits such as increased biomass, higher seed yield, higher sugar content, and enhanced tolerance against various abiotic stresses. Due to the losses in carbon fixation associated with photorespiratory metabolism, multiple efforts have been focused on reducing the rate of photorespiration in plants. Next to yet unsuccessful efforts toward the modification of RuBisCO oxygenase activity (119), independent strategies targeting downstream photorespiratory reactions have been described. This seminal approach (52 - WO2003100066, 63, 71 - WO2011095528) is based on heterologous overexpression of E. coli enzymes constituting the glycolate catabolic pathway (Fig. 7 A). In this strategy, glycolate is converted to glycerate directly in the chloroplast, which significantly reduces the photorespiratory flux through peroxisomes and mitochondria.

16

Page 17 of 63


17 Plants that utilize this photorespiratory bypass exhibited improved biomass production and increased sugar content (52 - WO2003100066, 62 - WO2010012796, 63, 71 WO2011095528, 109). Similarly to the first approach, a recent strategy (89) introduced a complete glycolate catabolic cycle in chloroplasts (Fig. 7 B). This cycle comprises glycolate oxidase, malate synthase and catalase, and was designed to metabolize glycolate into malate that is catalyzed by endogenous enzymes into pyruvate with the release of CO2 (89, 93 WO2009103782). This approach remains controversial, since no 3-PGA is recycled, and therefore the reaction depletes the intermediates of the Calvin cycle (122). Nevertheless, a growth improvement was reported for transgenic plants generated in this study (89). Initially associated with responses to high temperature, heat shock transcription factors (HSFs) are currently implicated in various other ROS-promoting conditions (152, 166). Importantly, HSFs play vital roles in the adaptation to oxidative stress (31, 117), which can be exploited in crop engineering. That said, the ectopic expression of HSF3 increases seed yield under control and drought stress conditions, and enhances tolerance toward biotrophic pathogens (106). Similarly, the overexpression of sunflower (Helianthus annuus) heat shock transcription factor (HaHSFA9) in Arabidopsis increases tolerance toward oxidative stress and dehydration (126). In response to drought, plants accumulate abscisic acid (ABA), required for activation of survival programs, stomatal closure, and reduction of water loss (7, 22). Changes of the ABA level are perceived by the PYR/PYL/RCAR (PYRABACTIN RESISTANCE/PYR1 LIKE/REGULATORY COMPONENT OF ABA RECEPTOR) receptor proteins (87, 118). Conformational changes triggered by ABA lead to interaction between the receptors and the PP2C (PROTEIN PHOSPHATASE 2C) protein phosphatase family members ABI1 (ABA INSENSITIVE 1) and ABI2 resulting in their inactivation. Both ABI1 and ABI2 have been shown to be negatively regulated by oxidative modifications, which amplify the ABA

17

Page 18 of 63


18 signaling cascade (Fig. 8) (95, 96). In the case of ABI2, GLUTATHIONE PEROXIDASE 3 (GPX3) relays oxidizing equivalents via thiol-disulfide exchange and inhibits the phosphatase activity of ABI2 (97). The inactivation of PP2C releases SnRK type 2 family of kinases (SnRK2), which phosphorylate transcription factors mediating ABA-related gene expression and ion channels orchestrating stomatal movements (78, 139). Among the targets of OST1, a member of the SnRK2 family, are the ROS-producing NADPH oxidases RbohD/F (147). Furthermore, OST1 phosphorylates SLAC1 (SLOW ANION CHANNEL ASSOCIATED 1), which controls the polarity of the guard cell membrane and hence the opening of K+ efflux channels governing stomatal movement (78). SLAC1 is further regulated by phosphorylation mediated by calcium (Ca2+)-dependent protein kinase (CPK) family members upon increase in cytosolic Ca2+ levels (47), which has been shown to be dependent on ABA-induced H2O2 accumulation (120). Several functionally redundant CPKs, such as CPK23, CPK21, CPK6, are activated by elevated Ca2+ levels (18, 47). Moreover, calcineurin B-like (CBL) calcium sensors CBL1 and CBL9 in complex with CIPK 23, a member of the CBL interacting protein kinases family, could also activate SLAC1 (90). Similarly, CPKs and CIPK26 can induce oxidative burst by the phosphorylation of Rbohs (40, 69). Among the upstream SLAC1 regulators, the activity of CPK21 is inhibited by H2O2 following formation of an intramolecular disulfide bond, which can be reduced by Trx h1 (165). Overexpression of constitutively active ABA PYL4 receptors leads to improved water use, drought tolerance, and enhanced ABA sensitivity (124). Conserved PYL4 homologs exist in many crop species including maize, rice, barley, soybean, alfalfa, and potato, implying that PYL4 overexpression might be an interesting avenue to engineer drought resistant crops. Putative membrane-associated phospholipid hydroperoxide glutathione peroxidase (PHGPx) has been proposed to act as a scavenger of lipid peroxidation products in analogy with glutathione peroxidase that removes excess of H2O2 with the formation of GSSG and

18

Page 19 of 63


19 water (94 - WO2010108836). Transgenic Arabidopsis plants expressing chloroplast-targeted glutathione peroxidase from S. cerevisiae (YIR037W) displayed an increased yield under water limiting conditions (94 - WO2010108836). When the protein was targeted to the mitochondria, however, a protective effect was not observed. In animals, GSH can be degraded by the action of γ-glutamyl transpeptidase (GGT) (82). The Arabidopsis genome contains four putative GGT genes (GGT1, GGT2, GGT3, and GGT4) (151). Knockouts of GGT3 show a reduced number of siliques and a reduced seed yield (92). Expressing γ-glutamyl transpeptidase (slr1269) from Synechocystis sp. in Arabidopsis

with

chloroplast

and

mitochondrial

signals,

negatively

affected

the

photosynthetic performance and plant biomass. However, when the protein was not targeted to any subcellular compartment, the transgenic plants displayed higher biomass in comparison to controls under cycling drought conditions (94 - WO2010108836). An alternative strategy based on boosting cellular GSH levels through the overexpression of the GSH-synthesizing enzymes, γ-glutamylcysteine synthetase and GSH synthetase, enhances the oxidative stress tolerance of lettuce and tomatoes (26 - WO2002033105). Although modifications of the GSH pool might be beneficial in certain cases and are relatively straightforward to generate, it remains to be proven if such approaches can be employed in crop improving strategies. Crucial for the viability of such technologies will be to test their effect in the field under fluctuating environmental conditions. The pentose phosphate pathway is induced under abiotic stress conditions and generates reducing equivalents (NADPH) and pentose sugars. One of the steps in the nonoxidative phase of this metabolic pathway is a reversible reaction between sedoheptulose-7phosphate and glyceraldehyde-3-phosphate catalyzed by transaldolase. The reaction products are erythrose-4-phosphate and fructose-6-phosphate, the latter being a glycolytic intermediate linking both pathways. Arabidopsis plants overexpressing bacterial Transaldolase A (b2464)

19

Page 20 of 63


20 performed better under drought conditions. Interestingly, under optimal watering conditions, these plants accumulated higher biomass (94 - WO2010108836). Glutaredoxins are ubiquitous oxidoreductases that play a major role in the regulation of S-glutathionylation, a reversible post-translational modification affecting protein stability and redox signaling (Fig. 6) (195). Arabidopsis plants expressing PvGRX5, encoding a glutaredoxin from fern Pterris vittata, displayed enhanced tolerance to heat during seed germination and at the mature stage (131 - WO2010021798). Additionally, these transgenic plants were more tolerant to drought, which was associated with reduced oxidative damage of proteins, such as HSP70, OEC33, RubisCO’s large subunit, RubisCO-activase, and chlorophyll a/b binding protein. These proteins have previously been shown to be extremely sensitive to oxidation (131 - WO2010021798).

Oxidative Stress-Based Strategies for Crop Protection Modern agriculture relies heavily on herbicides to control invasive weed species that might severely impair crop yield. Given the high importance of ROS metabolism and their immediate toxicity, not surprisingly, nearly half of the herbicides exert their effect either by directly boosting ROS levels or indirectly triggering ROS overproduction, eventually leading to cell death. Perhaps, the most notorious example of a ROS-generating herbicide is paraquat. Being among the most toxic and most widely used herbicides for decades, paraquat application is gradually declining, because of environmental concerns and tougher regulations (67). It acts quickly and non-selectively by transferring electrons from PSI to O2, which leads to the generation of O2•- (Fig. 1) (75). The precise mechanism involves the reduction of paraquat by ferredoxin and a subsequent electron transfer to O2 with paraquat being recycled back to its initial redox state, ready to accept a new electron (75). Thus, the potency of paraquat is a

20

Page 21 of 63


21 direct consequence of the architecture and the reactions executed at PSI, which functions as a plastocyanin-ferredoxin oxidoreductase. The triazine herbicides are widely applied blockers of PSII with atrazine being possibly the most well-known representative of this class (182). Their physicochemical properties allow them to occupy the plastoquinone binding site at the D1 protein, a crucial component of PSII (23). Since triazines are non-reducible, the electron flow from pheophytin is blocked, thus preventing dissipation of the excited energy state. The resulting high-energy (triplet) chlorophyll molecules readily form 1O2 and trigger oxidative stress (Fig. 1). (3-(3,4dichlorophenyl)-1,1-dimethylurea), commonly known as DCMU, is another chemically distinct herbicide that exerts its effect via the same mechanism (157). The protoporphyinogen oxidase (protox)-inhibiting herbicides kill plants by generating 1O2 in the chloroplasts (51). Blocking the metabolic flux at this chlorophyll biosynthetic enzyme results in the accumulation of protoporphyrinogen-IX, which is a powerful photosensitizer generating 1O2 in illuminated leaves (Fig. 1) (51). Strategies to develop Protox herbicide resistant plants have been reviewed previously and include identification and engineering of insensitive plant protoporphyrinogen oxidases (PPOs) or introduction of the Bacillus subtilis PPO coding sequence (81, 180 - US6084155). Bromoxynil and other structurally related 3,5-disubstituted 4-hydroxy benzonitriles act as PSII blockers at the level of plastoquinone reduction, thus provoking oxidative stress in high light conditions (Fig. 1) (28). The possibility to detoxify bromoxynil via nitrilasemediated hydrolysis served as the basis to develop bromoxynil-tolerant transgenic canola (Brassica napus). This was achieved by introducing a nitrilase-encoding gene from Klebsiella pneumonia (27, 149 - WO1987004181). The combined application of bromoxynil and bromoxynil-tolerant canola allows an effective control against broadleaf weed species threatening this crop.

21

Page 22 of 63


22 Discovered in an effort to generate novel chemical weapons during World War II, 2,4dichlorophenoxyacetic acid (2,4-D) was quickly recognized as a powerful selective herbicide and is currently one of the most widely used herbicides worldwide (188). 2,4-D belongs to the group of the synthetic phenoxyacetic auxins that largely mimic the effects of the naturally occurring auxin indole-3-acetic acid. Auxin is often regarded as a master regulator of plant physiology due to its involvement in virtually every aspect of plant growth and development (173). Binding of 2,4-D and other synthetic auxins to the receptor transport inhibitor response TIR1 and its homologs activates the auxin signaling machinery (66). At low concentrations, these chemicals stimulate growth, which makes them powerful growth regulators used in agriculture and tissue culture technologies (50). Applied at herbicidal rates, the synthetic auxins cause profound alterations of cellular morphology that eventually result in the disintegration of membrane structures and cell death. These effects are accompanied with increased rates of ROS formation. Thus, ROS-triggered processes resulting in progressive chlorosis, wilting, and necrosis, mediate cell collapse upon auxin herbicides treatment (50). Recently, metabolic resistance to 2,4-D and structurally related herbicides was engineered by transforming plants with bacterial aryloxyalkanoate dioxygenase (188). Given the low-cost and high selectivity of 2,4-D, this technology might play a major role in the production of herbicide-resistant crops in the future.

Biofortification and Food Security Food shortage and unequal distribution of food supplies pose imminent threats to the well-being of billions of people worldwide. Especially vulnerable are poor communities in developing countries relying predominantly on plant-based food. Their diet often cannot deliver the recommended daily vitamin allowances leading to health problems and even death. Severe deficiencies of vitamin A take thousands of children’s lives each year worldwide

22

Page 23 of 63


23 (187). In an effort to overcome million personal tragedies originating from a diet poor in vitamin A, researchers have developed a transgenic rice variety accumulating high amounts of β-carotene (pro-vitamin A) in the rice endosperm (115, 193). This gives the rice seeds a characteristic golden color and hence the name “Golden Rice”. The pro-vitamin A levels in the optimized Golden Rice 2 allow only 100 g (50 g for children) rice per day to cover the daily requirements of vitamin A (155). The introduction of the carotenoid biosynthetic machinery in the rice endosperm was achieved by combining maize phytoene synthase (psy) with the carotene desaturase (crtI) from Erwinia uredovora (115). Current regulatory measures pause the adoption of the Golden Rice technology in countries that need solutions to overcome nutrient deficiencies. β-carotene and other plant carotenoids fulfill important functions in plants (21). By serving as accessory pigments of the photosynthetic machinery, they absorb light and transfer its energy to the photosynthetic reaction centers (146). Moreover, carotenoids scavenge high-energy chlorophyll molecules and 1O2 formed during the photosynthetic activity (125). Oxidation products of carotenoids such as b-cyclocitral and b-ionene are potent signaling molecules that govern plant acclimation to adverse environmental conditions (130). There is an increasing interest to enrich edible plants with metabolites known to have beneficial health effects. Introduction of only two genes, Delila (Del) and Rosea1 (Ros1), from Antirrhinum majus in tomato boosted anthocyanin production and gave the fruits a deep purple color (19). Del and Ros1 encode basic helix-loop-helix and MYB-related transcription factors, respectively. Anthocyanin functions in plants include among others pigmentation important to attract pollinators, protection against UV radiation, and ROS scavenging (154). Increasing food production, one of the main goals of the agro-biotech industry, is only relevant if it is adequately used and not wasted. Currently, a significant amount of food is lost due to short shelf life, which restricts its logistical distribution and long-term storage. Tight

23

Page 24 of 63


24 control of ROS levels has been observed during the early ripening stages of tomato and guava, whereas oxidative stress progressively increased during maturation, implying a major role of ROS in fruit development (100, 101). Moreover, tomato and guava cultivars with a long shelf life displayed higher antioxidant capacity in comparison to cultivars with a short shelf life. Transgenic broccoli expressing isopentenyltransferase (ipt) accumulate antioxidant proteins such as SOD and APX, which is accompanied with delayed post-harvest yellowing (85). Taken together, these observations suggest that redox control during fruit maturation and storage could be exploited as a strategy to extend the shelf life. This is supported by the fact that the “purple tomatoes” described above displayed a prolonged shelf life possibly due to the antioxidant properties of the accumulated anthocyanins (196). Post-harvest physiological deterioration (PPD) observed in cassava roots, a staple food for millions in the tropics, significantly affects their quality within the first three days after harvest, making them unpalatable and impossible to sell (133). PPD has been associated with ROS accumulation and increased abundance of various proteins related to GSH homeostasis (133, 172). Delay of PPD and shelf-life increase have been observed in cassava cultivars containing higher levels of β-carotene (102). By overexpressing alternative oxidase from Arabidopis, Liu et al. (85) were able to extend the cassava shelf life to 10-21 days both under field and greenhouse conditions. Control of ROS levels through the overexpression of Cu/Zn SOD and catalase similarly delayed the onset of PPD with at least ten days (191). Cytosolic glutathione peroxidase overexpressors also displayed postponed PPD symptoms and lower levels of H2O2 (172).

Concluding Remarks Over the years, scientists have attempted to elucidate the role of ROS in plant development and stress responses. The gathered knowledge has repeatedly been used to

24

Page 25 of 63


25 improve plant traits such as biomass, stress tolerance, and nutritional value. We expect that the growing knowledge about ROS homeostasis, perception, and signaling will further stimulate efforts toward improving plant traits and future application of this knowledge into crop species.

Acknowledgments This work was supported by Ghent University (Multidisciplinary Research Partnership “Biotechnology for a Sustainable Economy”, Grant 01MRB510W), VIB and Marie Curie (OMICS@VIB PCOFUND-GA-2010-267139), FWO (Fonds Wetenschappelijk Onderzoek – Vlaanderen; “Sulfenomics: oxidatieve schakelaars in planten”), European Cooperation in Science and Technology (COST Action BM1203 “EU-ROS”), and the Interuniversity Attraction Poles Program (IUAP P7/29 “MARS”), initiated by the Belgian Science Policy Office. This work was also supported by the European Cooperation in Science and Research (COST Action BM1203 / EU‐ROS) and by FWO through a Pegasus postdoctoral grant funded to P.K, by IWT through a predoctoral fellowship to B.D.S and by VIB through a predoctoral scholarship (International PhD Program in Life Sciences) granted to C.W.

Author Disclosure Statement The authors report no conﬂict of interest.

Abbreviations used in the text 1

O2 = singlet oxygen

µE = microEinstein (µmol · m-2 · s-1) 2,4-D = dichlorophenoxyacetic acid 2OG = 2-oxoglutarate 2-PG = 2-phosphoglycolate

25

Page 26 of 63


26 3

O2 = state triplet oxygen

3-PGA = 3-phosphoglyceric acid ABA = abscisic acid ABI = abscisic acid insensitive AOX = alternative oxidase APX = ascorbate peroxidase ASC = ascorbate ATP = adenosine triphosphate BSO = buthionine sulphoximine Ca2+ = calcium ions CAT = catalase cat2 = catalase 2 CBL = calcineurin B-like CIPK = CBL interacting protein kinases family CO2 = carbon dioxide CPK = CALCIUM-DEPENDENT PROTEIN KINASE crtI = carotene desaturase Cu = copper Cys = cysteine Cys-SO2H = sulfinic acid Cys-SO3H = sulfonic acid Cys-SOH = sulfenic acid cETC = chloroplastic electron transport chains DCMU = 3-(3,4-dichlorophenyl)-1,1-dimethylurea Del = Delila DHA = dehydroascorbate DHAR = dehydroascorbate reductase DPI = diphenylene iodonium Fdx = ferredoxin Fe = iron Fe-S = iron-sulfur cluster flu = fluorescent FNR = ferredoxin-NADP+ reductase FTR = ferrodoxin-thioredoxin reductase 26

Page 27 of 63


27 Fv'/Fm' = PSII maximum efficiency G3P = glyceraldehyde 3-phosphate GCL = glyoxylate carboxylase GDC = glycine decarboxylase GDH = glycolate dehydrogenase GGT = γ-glutamyl transpeptidase GhCaM7 = Gossypium hirsutum Calmodulin 7 GLYK = glycerate kinase GO = glucose oxidase GOX = glycolate oxidase GPX = glutathione peroxidase GPX3 = GLUTATHIONE PEROXIDASE 3 GR = glutathione reductase Grx = glutaredoxin GSH = reduced glutathione GSNO = S-Nitrosoglutathione GSSG = oxidized glutathione GST = glutathione S-transferase H2O2 = hydrogen peroxide HD-ZIP = homeodomain-leucine zipper HO· = hydroxyl radical HPR = hydroxypyruvate reductase HR = hypersensitive response HSF = heat shock transcription factor HSP = heat shock protein ipt = isopentenyltransferase KCN = potassium cyanide MDA = monodehydroascorbate MDAR = monodehydroascorbate reductase ME = malic enzyme mETCs = mitochondrial electron transport chains Met-SO = methionine sulfoxide Mn = manganese MS = malate synthase 27

Page 28 of 63


28 Msr = methionine sulfoxide reductase NADP+ = oxidized nicotinamide adenine dinucleotide phosphate NADPH = reduced nicotinamide adenine dinucleotide phosphate NaN3 = sodium azide NH2CN = hydrogen cyanamide NPR1 = NONEXPRESSER OF PR GENES 1 NTR = NADPH-thioredoxin reductase O2 = oxygen O2•- = superoxide radical OEC33 = oxygen-evolving complex 33 kDa protein OST1 = OPEN STOMATA 1 PAN = PERIANTHIA PDH = pyruvate dehydrogenase PGLP = 2-phosphoglycolate phosphatase PHGPx = phospholipid hydroperoxide glutathione peroxidase PP2C = PROTEIN PHOSPHATASE 2C PPD = post-harvest physiological deterioration PPO = protoporphyrinogen oxidase ppm = parts per million PR = pathogenesis related protox = protoporphyinogen oxidase Prx = peroxiredoxin PSI = photosystem I PSII = photosystem II pSWPA2 = promoter of the sweet potato peroxidase 2 psy = phytoene synthase PTOX = plastoquinol terminal oxidase PYR/PYL/RCAR

=

PYRABACTIN

RESISTANCE/PYR1

COMPONENT OF ABA RECEPTOR QC = quiescent center Rboh = RESPIRATORY BURST OXIDASE HOMOLOG rlm = root meristemless ROS = reactive oxygen species Ros1 = Rosea1 28

LIKE/REGULATORY

Page 29 of 63


29 R-SH = sulfhydryl R-S-R’ = thioether RuBisCO = Ribulose-1,5-bisphosphate carboxylase/oxygenase RuBP = ribulose-1,5-phosphate SA = salicylic acid SGT = serine-glyoxylate aminotransferase SHMT = serine hydroxymathyl transferase SLAC1 = SLOW ANION CHANNEL-ASSOCIATED 1 SnRK = SnRK type 2 family of kinases SOD = superoxide dismutase Srx = sulfiredoxin TCP = TEOSINTE BRANCHED1/CYLOIDEA/PROLIFERATING CELL FACTOR TGA = TGACG SEQUENCE-SPECIFIC BINDING PROTEIN TIR1 = TRANSPORT INHIBITOR RESPONSE 1 Trx = thioredoxin TSR = tartronic semialdehyde reductase UGT74E2 = UDP-GLUCOSYL TRANSFERASE 74E2 UV = ultraviolet YAP1 = Yeast AP-1-like Zn = zinc

29

Page 30 of 63


30 References 1.

Achard P, Renou J-P, Berthomé R, Harberd NP, and Genschik P. Plant DELLAs restrain growth and promote survival of adversity by reducing the levels of reactive oxygen species. Curr Biol 18: 656-660, 2008.

2.

Allan AC and Fluhr R. Two distinct sources of elicited reactive oxygen species in tobacco epidermal cells. Plant Cell 9: 1559-1572, 1997.

3.

Alscher RG, Erturk N, and Heath LS. Role of superoxide dismutases (SODs) in controlling oxidative stress in plants. J Exp Bot 53: 1331-1341, 2002.

4.

Asada K. Production and scavenging of reactive oxygen species in chloroplasts and their functions. Plant Physiol 141: 391-396, 2006.

5.

Asada K, Kiso K, and Yoshikawa K. Univalent reduction of molecular oxygen by spinach chloroplasts on illumination. J Biol Chem 249: 2175-2181, 1974.

6.

Asai S, Ohta K, and Yoshioka H. MAPK signaling regulates nitric oxide and NADPH oxidase-dependent oxidative bursts in Nicotiana benthamiana. Plant Cell 20: 13901406, 2008.

7.

Assmann SM. OPEN STOMATA1 opens the door to ABA signaling in Arabidopsis guard cells. Trends Plant Sci 8: 151-153, 2003.

8.

Baker A, Graham IA, Holdsworth M, Smith SM, and Theodoulou FL. Chewing the fat: β-oxidation in signalling and development. Trends Plant Sci 11: 124-132, 2006.

9.

Bartlett DW, Clough JM, Godwin JR, Hall AA, Hamer M, and Parr-Dobrzanski B. The strobilurin fungicides. Pest Manag Sci 58: 649-662, 2002.

10.

Bashandy T, Guilleminot J, Vernoux T, Caparros-Ruiz D, Ljung K, Meyer Y, and Reichheld J-P. Interplay between the NADP-linked thioredoxin and glutathione systems in Arabidopsis auxin signaling. Plant Cell 22: 376-391, 2010.

30

Page 31 of 63


31 11.

Bauwe H, Hagemann M, Kern R, and Timm S. Photorespiration has a dual origin and manifold links to central metabolism. Curr Opin Plant Biol 15: 269-275, 2012.

12.

Baxter A, Mittler R, and Suzuki N. ROS as key players in plant stress signalling. J Exp Bot 65: 1229-1240, 2014.

13.

Berlett BS and Stadtman ER. Protein oxidation in aging, disease, and oxidative stress. J Biol Chem 272: 20313-20316, 1997.

14.

Bertelsen JR, de Neergaard E, and Smedegaard-Petersen V. Fungicidal effects of azoxystrobin and epoxiconazole on phyllosphere fungi, senescence and yield of winter wheat. Plant Pathol 50: 190-205, 2001.

15.

Bidney DL, Charne DG, Coughlan SL, Falak I, Mancl MK, Nazarian KP, Scelonge CJ, and Yalpani N. Production of pathogen resistant plants (WO199904012). USA, Pioneer Hi-Bred International, Inc., 1999.

16.

Bowes G, Ogren WL, and Hageman RH. Phosphoglycolate production catalyzed by ribulose diphosphate carboxylase. Biochem Biophys Res Commun 45: 716-722, 1971.

17.

Bowler C, Inzé D, Van Camp W, Alliotte T, Van Montagu M, and Botterman J. Stress-tolerant plants (WO1990002804). Belgium, Plant Genetic Systems NV, 1990.

18.

Brandt B, Brodsky DE, Xue S, Negi J, Iba K, Kangasjärvi J, Ghassemian M, Stephan AB, Hu H, and Schroeder JI. Reconstitution of abscisic acid activation of SLAC1 anion channel by CPK6 and OST1 kinases and branched ABI1 PP2C phosphatase action. Proc Natl Acad Sci USA 109: 10593-10598, 2012.

19.

Butelli E, Titta L, Giorgio M, Mock H-P, Matros A, Peterek S, Schijlen EGWM, Hall RD, Bovy AG, Luo J, and Martin C. Enrichment of tomato fruit with healthpromoting anthocyanins by expression of select transcription factors. Nat Biotechnol 26: 1301-1308, 2008.

31

Page 32 of 63


32 20.

Castillo MC and León J. Expression of the β-oxidation gene 3-ketoacyl-CoA thiolase 2 (KAT2) is required for the timely onset of natural and dark-induced leaf senescence in Arabidopsis. J Exp Bot 59: 2171-2179, 2008.

21.

Cazzonelli CI. Carotenoids in nature: insights from plants and beyond. Funct Plant Biol 38: 833-847, 2011.

22.

Christmann A, Weiler EW, Steudle E, and Grill E. A hydraulic signal in root-to-shoot signalling of water shortage. Plant J 52: 167-174, 2007.

23.

Cobb

AH

and

Reade

JPH.

Herbicides

and

Plant

Physiology,

Second

EditionHerbicides and Plant Physiology, Second Edition. Oxford, United Kingdom: Wiley-Blackwell.2010. 24.

Comelli RN and Gonzalez DH. Conserved homeodomain cysteines confer redox sensitivity and influence the DNA binding properties of plant class IIIHD-Zip proteins. Arch Biochem Biophys 467: 41-47, 2007.

25.

Considine MJ and Foyer CH. Redox regulation of plant development. Antioxid Redox Signal 21: 1305-1326, 2014.

26.

Creissen GP and Mullineaux PM. Reducing oxidative stress of plants by increasing glutathione content (WO2002033105). United Kingdom, Plant Bioscience Ltd., 2002.

27.

Cuthbert JL, McVetty PBE, Freyssinet G, and Freyssinet M. Comparison of the performance of bromoxynil-resistant and susceptible near-isogenic populations of oilseed rape. Can J Plant Sci 81: 367-372, 2001.

28.

Cutulle MA, Armel GR, Brosnan JT, Best MD, Kopsell DA, Bruce BD, Bostic HE, and Layton DS. Synthesis and evaluation of heterocyclic analogues of bromoxynil. J Agric Food Chem 62: 329-336, 2014.

32

Page 33 of 63


33 29.

Daudi A, Cheng Z, O'Brien JA, Mammarella N, Khan S, Ausubel FM, and Bolwell GP. The apoplastic oxidative burst peroxidase in Arabidopsis is a major component of pattern-triggered immunity. Plant Cell 24: 275-287, 2012.

30.

Davies MJ. The oxidative environment and protein damage. Biochim Biophys Acta Proteins Proteomics 1703: 93-109, 2005.

31.

Davletova S, Rizhsky L, Liang H, Shengqiang Z, Oliver DJ, Coutu J, Shulaev V, Schlauch K, and Mittler R. Cytosolic ascorbate peroxidase 1 is a central component of the reactive oxygen gene network of Arabidopsis. Plant Cell 17: 268-281, 2005.

32.

de la Peña TC, Redondo FJ, Manrique E, Lucas MM, and Pueyo JJ. Nitrogen fixation persists under conditions of salt stress in transgenic Medicago truncatula plants expressing a cyanobacterial flavodoxin. Plant Biotechnol J 8: 954-965, 2010.

33.

Delaunay A, Isnard A-D, and Toledano MB. H2O2 sensing through oxidation of the Yap1 transcription factor. EMBO J 19: 5157-5166, 2000.

34.

Delaunay A, Pflieger D, Barrault M-B, Vinh J, and Toledano MB. A thiol peroxidase is an H2O2 receptor and redox-transducer in gene activation. Cell 111: 471-481, 2002.

35.

Després C, Chubak C, Rochon A, Clark R, Bethune T, Desveaux D, and Fobert PR. The Arabidopsis NPR1 disease resistance protein is a novel cofactor that confers redox regulation of DNA binding activity to the basic domain/leucine zipper transcription factor TGA1. Plant Cell 15: 2181-2191, 2003.

36.

Diaz-Espejo A, Cuevas MV, Ribas-Carbo M, Flexas J, Martorell S, and Fernández JE. The effect of strobilurins on leaf gas exchange, water use efficiency and ABA content in grapevine under field conditions. J Plant Physiol 169: 379-386, 2012.

37.

Diaz-Vivancos P, Faize M, Barba-Espin G, Faize L, Petri C, Hernández JA, and Burgos L. Ectopic expression of cytosolic superoxide dismutase and ascorbate

33

Page 34 of 63


34 peroxidase leads to salt stress tolerance in transgenic plums. Plant Biotechnol J 11: 976-985, 2013. 38.

Dietz K-J. Redox regulation of transcription factors in plant stress acclimation and development. Antioxid Redox Signal 21: 1356-1372, 2014.

39.

Dismukes GC, Klimov VV, Baranov SV, Kozlov YN, DasGupta J, and Tyryshkin A. The origin of atmospheric oxygen on Earth: The innovation of oxygenic photosynthesis. Proc Natl Acad Sci USA 98: 2170-2175, 2001.

40.

Drerup MM, Schlücking K, Hashimoto K, Manishankar P, Steinhorst L, Kuchitsu K, and Kudla J. The calcineurin B-like calcium sensors CBL1 and CBL9 together with their interacting protein kinase CIPK26 regulate the Arabidopsis NADPH oxidase RBOHF. Mol Plant 6: 559-569, 2013.

41.

Faize M, Burgos L, Faize L, Piqueras A, Nicolas E, Barba-Espin G, Clemente-Moreno MJ, Alcobendas R, Artlip T, and Hernandez JA. Involvement of cytosolic ascorbate peroxidase and Cu/Zn-superoxide dismutase for improved tolerance against drought stress. J Exp Bot 62: 2599-2613, 2011.

42.

Falk S, Samson G, Bruce D, Huner NPA, and Laudenbach DE. Functional analysis of the iron-stress induced CP 43 ′ polypeptide of PS II in the cyanobacterium Synechococcus sp. PCC 7942. Photosynth Res 45: 51-60, 1995.

43.

Foreman J, Demidchik V, Bothwell JHF, Mylona P, Miedema H, Torres MA, Linstead P, Costa S, Brownlee C, Jones JDG, Davies JM, and Dolan L. Reactive oxygen species produced by NADPH oxidase regulate plant cell growth. Nature 422: 442-446, 2003.

44.

Foyer CH and Noctor G. Redox regulation in photosynthetic organisms: signaling, acclimation, and practical implications. Antioxid Redox Signal 11: 861-905, 2009.

34

Page 35 of 63


35 45.

Fryer MJ, Ball L, Oxborough K, Karpinski S, Mullineaux PM, and Baker NR. Control of Ascorbate Peroxidase 2 expression by hydrogen peroxide and leaf water status during excess light stress reveals a functional organisation of Arabidopsis leaves. Plant J 33: 691-705, 2003.

46.

Gadjev I, Vanderauwera S, Gechev TS, Laloi C, Minkov IN, Shulaev V, Apel K, Inzé D, Mittler R, and Van Breusegem F. Transcriptomic footprints disclose specificity of reactive oxygen species signaling in Arabidopsis. Plant Physiol 141: 436-445, 2006.

47.

Geiger D, Scherzer S, Mumm P, Marten I, Ache P, Matschi S, Liese A, Wellmann C, Al-Rasheid KAS, Grill E, Romeis T, and Hedrich R. Guard cell anion channel SLAC1 is regulated by CDPK protein kinases with distinct Ca2+ affinities. Proc Natl Acad Sci USA 107: 8023-8028, 2010.

48.

Gill SS and Tuteja N. Reactive oxygen species and antioxidant machinery in abiotic stress tolerance in crop plants. Plant Physiol Biochem 48: 909-930, 2010.

49.

Govrin EM and Levine A. The hypersensitive response facilitates plant infection by the necrotrophic pathogen Botrytis cinerea. Curr Biol 10: 751-757, 2000.

50.

Grossmann K. Auxin herbicides: current status of mechanism and mode of action. Pest Manag Sci 66: 113-120, 2010.

51.

Grossmann K, Niggeweg R, Christiansen N, Looser R, and Ehrhardt T. The herbicide saflufenacil (KixorTM) is a new inhibitor of protoporphyrinogen IX oxidase activity. Weed Sci 58: 1-9, 2010.

52.

Hain R, Berg D, Peterhänsel C, Kreuzaler F, Bari R, Weier D, Hirsch H-J, and Rademacher T. A method for production of plants with suppressed photorespiration and improved CO2 fixation (WO2003100066). Germany, Bayer CropScience AG, 2003.

35

Page 36 of 63


36 53.

Halaly T, Pang X, Batikoff T, Crane O, Keren A, Venkateswari J, Ogrodovitch A, Sadka A, Lavee S, and Or E. Similar mechanisms might be triggered by alternative external stimuli that induce dormancy release in grape buds. Planta 228: 79-88, 2008.

54.

Hanke G and Mulo P. Plant type ferredoxins and ferredoxin-dependent metabolism. Plant Cell Environ 36: 1071-1084, 2013.

55.

Herrmann W and Obeid R. Vitamins in the prevention of human diseasesVitamins in the prevention of human diseases. Berlin/New York: Walter de Gruyter GmbH & Co.2011.

56.

Iglesias-Baena I, Barranco-Medina S, Sevilla F, and Lázaro J-J. The dual-targeted plant sulfiredoxin retroreduces the sulfinic form of atypical mitochondrial peroxiredoxin. Plant Physiol 155: 944-955, 2011.

57.

Jacques S, Ghesquière B, Van Breusegem F, and Gevaert K. Plant proteins under oxidative attack. Proteomics 13: 932-940, 2013.

58.

Jain V, Kaiser W, and Huber SC. Cytokinin inhibits the proteasome-mediated degradation of carbonylated proteins in Arabidopsis leaves. Plant Cell Physiol 49: 843-852, 2008.

59.

Jiang K, Meng YL, and Feldman LJ. Quiescent center formation in maize roots is associated with an auxin-regulated oxidizing environment. Development 130: 14291438, 2003.

60.

Joo JH, Bae YS, and Lee JS. Role of auxin-induced reactive oxygen species in root gravitropism. Plant Physiol 126: 1055-1060, 2001.

61.

Kaya H, Nakajima R, Iwano M, Kanaoka MM, Kimura S, Takeda S, Kawarazaki T, Senzaki E, Hamamura Y, Higashiyama T, Takayama S, Abe M, and Kuchitsu K. Ca2+activated reactive oxygen species production by Arabidopsis RbohH and RbohJ is essential for proper pollen tube tip growth. Plant Cell 26: 1069-1080, 2014.

36

Page 37 of 63


37 62.

Kebeish R, Kreuzaler F, Metzlaff M, Niessen M, Peterhänsel C, and Van Rie J. A method for increasing photosynthetic carbon fixation in rice (WO2010012796). Germany and Belgium, Bayer CropScience AG, 2010.

63.

Kebeish R, Niessen M, Thiruveedhi K, Bari R, Hirsch H-J, Rosenkranz R, Stäbler N, Schönfeld B, Kreuzaler F, and Peterhänsel C. Chloroplastic photorespiratory bypass increases photosynthesis and biomass production in Arabidopsis thaliana. Nat Biotechnol 25: 593-599, 2007.

64.

Kelliher T and Walbot V. Hypoxia triggers meiotic fate acquisition in maize. Science 337: 345-348, 2012.

65.

Kelliher T and Walbot V. Method for modulating the number of archesporial cells in developing anther (WO2013123051). USA, The Board of Trustees of the Leland Stanford Junior University, 2013.

66.

Kepinski S and Leyser O. The Arabidopsis F-box protein TIR1 is an auxin receptor. Nature 435: 446-451, 2005.

67.

Kervégant M, Merigot L, Glaizal M, Schmitt C, Tichadou L, and de Haro L. Paraquat poisonings in France during the European ban: experience of the Poison Control Center in Marseille. J Med Toxicol 9: 144-147, 2013.

68.

Kim C, Meskauskiene R, Apel K, and Laloi C. No single way to understand singlet oxygen signalling in plants. EMBO Rep 9: 435-439, 2008.

69.

Kobayashi M, Ohura I, Kawakita K, Yokota N, Fujiwara M, Shimamoto K, Doke N, and Yoshioka H. Calcium-dependent protein kinases regulate the production of reactive oxygen species by potato NADPH oxidase. Plant Cell 19: 1065-1080, 2007.

70.

Koprivova A, Mugford ST, and Kopriva S. Arabidopsis root growth dependence on glutathione is linked to auxin transport. Plant Cell Rep 29: 1157-1167, 2010.

37

Page 38 of 63


38 71.

Kreuzaler F, Noelke G, Peterhänsel C, and Schillberg S. A method for increasing photosynthetic carbon fixation using glycolate dehydrogenase multi-subunit fusion protein (WO2011095528). Germany, Bayer Cropscience AG, 2011.

72.

Krieger-Liszkay A, Fufezan C, and Trebst A. Singlet oxygen production in photosystem II and related protection mechanism. Photosynth Res 98: 551-564, 2008.

73.

Krogan NT and Long JA. Why so repressed? Turning off transcription during plant growth and development. Curr Opin Plant Biol 12: 628-636, 2009.

74.

Kwak S-S, Kwon S-Y, Lee H-S, Tang L, Lim S, and Lee B-H. Recombinant expression vector for production of plants having multiple stress tolerances, and method for preparing multiple stress-tolerant plants using the same (WO2006054815). Korea, Korea Research Institute of Bioscience and Biotechnology, 2006.

75.

Lascano R, Muñoz N, Robert G, Rodriguez M, Melchiorre M, Trippi V, and Quero G. Paraquat: an oxidative stress inducer, In: Herbicides – Properties, Synthesis and Control of Weeds,edited by MNAE-L Hasaneen: InTech, 2012, pp. 135-148.

76.

Lawrence EB, Levine EB, and Shah DM. Method of controlling plant pathogens (WO1995014784). USA, Monsanto Co, 1995.

77.

Lee S-H, Ahsan N, Lee K-W, Kim D-H, Lee D-G, Kwak S-S, Kwon S-Y, Kim T-H, and Lee B-H. Simultaneous overexpression of both CuZn superoxide dismutase and ascorbate peroxidase in transgenic tall fescue plants confers increased tolerance to a wide range of abiotic stresses. J Plant Physiol 164: 1626-1638, 2007.

78.

Lee SC, Lan W, Buchanan BB, and Luan S. A protein kinase-phosphatase pair interacts with an ion channel to regulate ABA signaling in plant guard cells. Proc Natl Acad Sci USA 106: 21419-21424, 2009.

38

Page 39 of 63


39 79.

Levine A, Tenhaken R, Dixon R, and Lamb C. H2O2 from the oxidative burst orchestrates the plant hypersensitive disease resistance response. Cell 79: 583-593, 1994.

80.

Li S, Lauri A, Ziemann M, Busch A, Bhave M, and Zachgo S. Nuclear activity of ROXY1, a glutaredoxin interacting with TGA factors, is required for petal development in Arabidopsis thaliana. Plant Cell 21: 429-441, 2009.

81.

Li X and Nicholl D. Development of PPO inhibitor-resistant cultures and crops. Pest Manag Sci 61: 277-285, 2005.

82.

Lieberman MW, Barrios R, Carter BZ, Habib GM, Lebovitz RM, Rajagopalan S, Sepulveda AR, Shi Z-Z, and Wan D-F. γ-Glutamyl-transpeptidase - What does the organization and expression of a multipromoter gene tell us about its functions? Am J Pathol 147: 1175-1185, 1995.

83.

Lindermayr C, Sell S, Müller B, Leister D, and Durner J. Redox regulation of the NPR1-TGA1 system of Arabidopsis thaliana by nitric oxide. Plant Cell 22: 28942907, 2010.

84.

Liso R, Innocenti AM, Bitonti MB, and Arrigoni O. Ascorbic acid-induced progression of quiescent centre cells from G1 to S phase. New Phytol 110: 469-471, 1988.

85.

Liu M-S, Li H-C, Chang Y-M, Wu M-T, and Chen L-FO. Proteomic analysis of stress-related proteins in transgenic broccoli harboring a gene for cytokinin production during postharvest senescence. Plant Sci 181: 288-299, 2011.

86.

Lu D, Wang T, Persson S, Mueller-Roeber B, and Schippers JHM. Transcriptional control of ROS homeostasis by KUODA1 regulates cell expansion during leaf development. Nat Commun 5: 3767, 2014.

39

Page 40 of 63


40 87.

Ma Y, Szostkiewicz I, Korte A, Moes D, Yang Y, Christmann A, and Grill E. Regulators of PP2C phosphatase activity function as abscisic acid sensors. Science 324: 1064-1068, 2009.

88.

Magiorkinis E, Beloukas A, and Diamantis A. Scurvy: Past, present and future. Eur J Intern Med 22: 147-152, 2011.

89.

Maier A, Fahnenstich H, von Caemmerer S, Engqvist MKM, Weber APM, Flügge UI, and Maurino VG. Transgenic introduction of a glycolate oxidative cycle into A. thaliana chloroplasts leads to growth improvement. Front Plant Sci 3: 38, 2012.

90.

Maierhofer T, Diekmann M, Offenborn JN, Lind C, Bauer H, Hashimoto K, AlRasheid KAS, Luan S, Kudla J, Geiger D, and Hedrich R. Site- and kinase-specific phosphorylation-mediated activation of SLAC1, a guard cell anion channel stimulated by abscisic acid. Sci Signal 7: ra86, 2014.

91.

Mandal S, Das RK, and Mishra S. Differential occurrence of oxidative burst and antioxidative mechanism in compatible and incompatible interactions of Solanum lycopersicum and Ralstonia solanacearum. Plant Physiol Biochem 49: 117-123, 2011.

92.

Martin MN, Saladores PH, Lambert E, Hudson AO, and Leustek T. Localization of members of the γ-glutamyl transpeptidase family identifies sites of glutathione and glutathione S-conjugate hydrolysis. Plant Physiol 144: 1715-1732, 2007.

93.

Maurino VG and Flügge U-I. Means for improving agrobiological traits in a plant by providing a plant cell comprising in its chloroplasts enzymatic activities for converting glycolate into malate (WO2009103782). Germany, Universität Zu Köln, 2009.

94.

McKersie B. Transgenic plants with altered redox mechanisms and increased yield (WO2010108836). Germany, BASF Plant Science Company GmbH, 2010.

95.

Meinhard M and Grill E. Hydrogen peroxide is a regulator of ABI1, a protein phosphatase 2C from Arabidopsis. FEBS Lett 508: 443-446, 2001.

40

Page 41 of 63


41 96.

Meinhard M, Rodriguez PL, and Grill E. The sensitivity of ABI2 to hydrogen peroxide links the abscisic acid-response regulator to redox signalling. Planta 214: 775-782, 2002.

97.

Miao Y, Lv D, Wang P, Wang X-C, Chen J, Miao C, and Song C-P. An Arabidopsis glutathione peroxidase functions as both a redox transducer and a scavenger in abscisic acid and drought stress responses. Plant Cell 18: 2749-2766, 2006.

98.

Miller G, Schlauch K, Tam R, Cortes D, Torres MA, Shulaev V, Dangl JL, and Mittler R. The plant NADPH oxidase RBOHD mediates rapid systemic signaling in response to diverse stimuli. Sci Signal 2: ra45, 2009.

99.

Molina L and Kahmann R. An Ustilago maydis gene involved in H2O2 detoxification is required for virulence. Plant Cell 19: 2293-2309, 2007.

100.

Mondal K, Malhotra SP, Jain V, and Singh R. Oxidative stress and antioxidant systems in Guava (Psidium guajava L.) fruits during ripening. Physiol Mol Biol Plants 15: 327-334, 2009.

101.

Mondal K, Sharma NS, Malhotra SP, Dhawan K, and Singh R. Antioxidant systems in ripening tomato fruits. Biol Plant 48: 49-53, 2004.

102.

Morante N, Sánchez T, Ceballos H, Calle F, Pérez JC, Egesi C, Cuambe CE, Escobar AF, Ortiz D, Chávez AL, and Fregene M. Tolerance to postharvest physiological deterioration in cassava roots. Crop Sci 50: 1333-1338, 2010.

103.

Morawala-Patell V. Rice conferring resistance to environmental stress by targetting mnsod to the chloroplast (WO2004053136). India, Avestha Gengraine Tech Pvt Ltd, 2004.

104.

Mou Z, Fan W, and Dong X. Inducers of plant systemic acquired resistance regulate NPR1 function through redox changes. Cell 113: 935-944, 2003.

41

Page 42 of 63


42 105.

Müller K, Linkies A, Vreeburg RAM, Fry SC, Krieger-Liszkay A, and LeubnerMetzger G. In vivo cell wall loosening by hydroxyl radicals during cress seed germination and elongation growth. Plant Physiol 150: 1855-1865, 2009.

106.

Mullineaux P and Bechthold U. Plant responses (WO2008110848). United Kingdom, Wivenhoe Technology Limited, 2008.

107.

Mur LAJ, Laarhoven LJJ, Harren FJM, Hall MA, and Smith AR. Nitric oxide interacts with salicylate to regulate biphasic ethylene production during the hypersensitive response. Plant Physiol 148: 1537-1546, 2008.

108.

Noctor G, Veljovic-Jovanovic S, Driscoll S, Novitskaya L, and Foyer CH. Drought and oxidative load in the leaves of C3 plants: a predominant role for photorespiration? Ann Bot 89: 841-850, 2002.

109.

Nölke G, Houdelet M, Kreuzaler F, Peterhänsel C, and Schillberg S. The expression of a recombinant glycolate dehydrogenase polyprotein in potato (Solanum tuberosum) plastids strongly enhances photosynthesis and tuber yield. Plant Biotechnol J 12: 734742, 2014.

110.

Nyström T. Role of oxidative carbonylation in protein quality control and senescence. EMBO J 24: 1311-1317, 2005.

111.

Ogawa K, Henmi K, and Tasaka Y. Cell- or organ-differentiation controllers and method of controlling morphogenesis by using the same (WO2001080638). Japan, Okayama Prefecture, 2001.

112.

Ogawa K, Tasaka Y, Mino M, Tanaka Y, and Iwabuchi M. Association of glutathione with flowering in Arabidopsis thaliana. Plant Cell Physiol 42: 524-530, 2001.

113.

op den Camp RGL, Przybyla D, Ochsenbein C, Laloi C, Kim C, Danon A, Wagner D, Hideg E, Göbel C, Feussner I, Nater M, and Apel K. Rapid induction of distinct stress

42

Page 43 of 63


43 responses after the release of singlet oxygen in Arabidopsis. Plant Cell 15: 2320-2332, 2003. 114.

Orozco-Cardenas M and Ryan CA. Hydrogen peroxide is generated systemically in plant leaves by wounding and systemin via the octadecanoid pathway. Proc Natl Acad Sci USA 96: 6553-6557, 1999.

115.

Paine JA, Shipton CA, Chaggar S, Howells RM, Kennedy MJ, Vernon G, Wright SY, Hinchliffe E, Adams JL, Silverstone AL, and Drake R. Improving the nutritional value of Golden Rice through increased pro-vitamin A content. Nat Biotechnol 23: 482-487, 2005.

116.

Palatnik JF, Fillat-Castejon MF, Carrillo N, Valle EM, and Tognetti VB. Stress tolerant plants (WO2003035881). United Kingdom, Plant Bioscience Ltd, 2003.

117.

Panchuk II, Volkov RA, and Schöffl F. Heat stress- and heat shock transcription factor-dependent expression and activity of ascorbate peroxidase in Arabidopsis. Plant Physiol 129: 838-853, 2002.

118.

Park S-Y, Fung P, Nishimura N, Jensen DR, Fujii H, Zhao Y, Lumba S, Santiago J, Rodrigues A, Chow T-fF, Alfred SE, Bonetta D, Finkelstein R, Provart NJ, Desveaux D, Rodriguez PL, McCourt P, Zhu J-K, Schroeder JI, Volkman BF, and Cutler SR. Abscisic acid inhibits type 2C protein phosphatases via the PYR/PYL family of START proteins. Science 324: 1068-1071, 2009.

119.

Parry MAJ, Andralojc PJ, Scales JC, Salvucci ME, Carmo-Silva AE, Alonso H, and Whitney SM. Rubisco activity and regulation as targets for crop improvement. J Exp Bot 64: 717-730, 2013.

120.

Pei Z-M, Murata Y, Benning G, Thomine S, Klüsener B, Allen GJ, Grill E, and Schroeder JI. Calcium channels activated by hydrogen peroxide mediate abscisic acid signalling in guard cells. Nature 406: 731-734, 2000.

43

Page 44 of 63


44 121.

Peng M and Kuc J. Peroxidase-generated hydrogen peroxide as a source of antifungal activity in vitro and on tobacco leaf disks. Phytopathology 82: 696-699, 1992.

122.

Peterhansel C, Blume C, and Offermann S. Photorespiratory bypasses: how can they work? J Exp Bot 64: 709-715, 2013.

123.

Petrov VD and Van Breusegem F. Hydrogen peroxide - a central hub for information flow in plant cells. AoB Plants 2012: pls014, 2012.

124.

Pizzio GA, Rodriguez L, Antoni R, Gonzalez-Guzman M, Yunta C, Merilo E, Kollist H, Albert A, and Rodriguez PL. The PYL4 A194T mutant uncovers a key role of PYR1-LIKE4/PROTEIN PHOSPHATASE 2CA interaction for abscisic acid signaling and plant drought resistance. Plant Physiol 163: 441-455, 2013.

125.

Polívka T and Sundström V. Ultrafast dynamics of carotenoid excited states - From solution to natural and artificial systems. Chem Rev 104: 2021-2071, 2004.

126.

Prieto-Dapena P, Castaño R, Almoguera C, and Jordano J. The ectopic overexpression of a seed-specific transcription factor, HaHSFA9, confers tolerance to severe dehydration in vegetative organs. Plant J 54: 1004-1014, 2008.

127.

Przybyla D, Göbel C, Imboden A, Hamberg M, Feussner I, and Apel K. Enzymatic, but not non-enzymatic, 1O2-mediated peroxidation of polyunsaturated fatty acids forms part of the EXECUTER1-dependent stress response program in the flu mutant of Arabidopsis thaliana. Plant J 54: 236-248, 2008.

128.

Queval G, Hager J, Gakière B, and Noctor G. Why are literature data for H 2O2 contents so variable? A discussion of potential difficulties in the quantitative assay of leaf extracts. J Exp Bot 59: 135-146, 2008.

129.

Queval G, Issakidis-Bourguet E, Hoeberichts FA, Vandorpe M, Gakière B, Vanacker H, Miginiac-Maslow M, Van Breusegem F, and Noctor G. Conditional oxidative stress responses in the Arabidopsis photorespiratory mutant cat2 demonstrate that

44

Page 45 of 63


45 redox state is a key modulator of daylength-dependent gene expression, and define photoperiod as a crucial factor in the regulation of H2O2-induced cell death. Plant J 52: 640-657, 2007. 130.

Ramel F, Birtic S, Ginies C, Soubigou-Taconnat L, Triantaphylidès C, and Havaux M. Carotenoid oxidation products are stress signals that mediate gene responses to singlet oxygen in plants. Proc Natl Acad Sci USA 109: 5535-5540, 2012.

131.

Rathinasabapathi B and Sundaram S. Improved crop stress tolerance, yield and quality via glutaredoxin overexpression (WO2010021798). USA, University Of Florida Research Foundation, Inc., 2010.

132.

Reichheld J-P, Khafif M, Riondet C, Droux M, Bonnard G, and Meyer Y. Inactivation of thioredoxin reductases reveals a complex interplay between thioredoxin and glutathione pathways in Arabidopsis development. Plant Cell 19: 1851-1865, 2007.

133.

Reilly K, Gómez-Vásquez R, Buschmann H, Tohme J, and Beeching JR. Oxidative stress responses during cassava post-harvest physiological deterioration. Plant Mol Biol 56: 625-641, 2004.

134.

Reuzeau C. Plants having one or more enhanced yield-related traits and method for making the same (WO2013150402). Germany, Switzerland and China, BASF Plant Science Company GmbH, BASF Schweiz AG, BASF (China) Company Limited, 2013.

135.

Rey P, Bécuwe N, Barrault M-B, Rumeau D, Havaux M, Biteau B, and Toledano MB. The Arabidopsis thaliana sulfiredoxin is a plastidic cysteine-sulfinic acid reductase involved in the photooxidative stress response. Plant J 49: 505-514, 2007.

136.

Rojas CM, Senthil-Kumar M, Wang K, Ryu C-M, Kaundal A, and Mysore KS. Glycolate oxidase modulates reactive oxygen species-mediated signal transduction

45

Page 46 of 63


46 during nonhost resistance in Nicotiana benthamiana and Arabidopsis. Plant Cell 24: 336-352, 2012. 137.

Roos G and Messens J. Protein sulfenic acid formation: From cellular damage to redox regulation. Free Radic Biol Med 51: 314-326, 2011.

138.

Rosenwasser S, Fluhr R, Joshi JR, Leviatan N, Sela N, Hetzroni A, and Friedman H. ROSMETER: a bioinformatic tool for the identification of transcriptomic imprints related to reactive oxygen species type and origin provides new insights into stress responses. Plant Physiol 163: 1071-1083, 2013.

139.

Sato A, Sato Y, Fukao Y, Fujiwara M, Umezawa T, Shinozaki K, Hibi T, Taniguchi M, Miyake H, Goto DB, and Uozumi N. Threonine at position 306 of the KAT1 potassium channel is essential for channel activity and is a target site for ABAactivated SnRK2/OST1/SnRK2.6 protein kinase. Biochem J 424: 439-448, 2009.

140.

Scheler C, Durner J, and Astier J. Nitric oxide and reactive oxygen species in plant biotic interactions. Curr Opin Plant Biol 16: 534-539, 2013.

141.

Schmid-Siegert E, Loscos J, and Farmer EE. Inducible malondialdehyde pools in zones of cell proliferation and developing tissues in Arabidopsis. J Biol Chem 287: 8954-8962, 2012.

142.

Schürmann P and Buchanan BB. The ferredoxin/thioredoxin system of oxygenic photosynthesis. Antioxid Redox Signal 10: 1235-1274, 2008.

143.

Sewelam N, Jaspert N, Van Der Kelen K, Tognetti VB, Schmitz J, Frerigmann H, Stahl E, Zeier J, Van Breusegem F, and Maurino VG. Spatial H2O2 signaling specificity: H2O2 from chloroplasts and peroxisomes modulates the plant transcriptome differentially. Mol Plant 7: 1191-1210, 2014.

144.

Shulaev V and Oliver DJ. Metabolic and proteomic markers for oxidative stress. New tools for reactive oxygen species research. Plant Physiol 141: 367-372, 2006.

46

Page 47 of 63


47 145.

Siddique S, Matera C, Radakovic ZS, Shamim Hasan M, Gutbrod P, Rozanska E, Sobczak M, Torres MA, and Grundler FM. Parasitic worms stimulate host NADPH oxidases to produce reactive oxygen species that limit plant cell death and promote infection. Sci Signal 7: ra33, 2014.

146.

Siefermann-Harms D. The light-harvesting and protective functions of carotenoids in photosynthetic membranes. Physiol Plant 69: 561-568, 1987.

147.

Sirichandra C, Gu D, Hu H-C, Davanture M, Lee S, Djaoui M, Valot B, Zivy M, Leung J, Merlot S, and Kwak JM. Phosphorylation of the Arabidopsis AtrbohF NADPH oxidase by OST1 protein kinase. FEBS Lett 583: 2982-2986, 2009.

148.

Slooten L, Capiau K, Van Camp W, Van Montagu M, Sybesma C, and InzéD. Factors affecting the enhancement of oxidative stress tolerance in transgenic tobacco overexpressing manganese superoxide dismutase in the chloroplasts. Plant Physiol 107: 737-750, 1995.

149.

Stalker M. Haloarylnitrile degrading gene, its use, and cells containing the same (WO1987004181). USA, Calgene Inc, 1987.

150.

Stamati K, Mudera V, and Cheema U. Evolution of oxygen utilization in multicellular organisms and implications for cell signalling in tissue engineering. J Tissue Eng 2: 2041731411432365, 2011.

151.

Storozhenko S, Belles-Boix E, Babiychuk E, Hérouart D, Davey MW, Slooten L, Van Montagu M, Inzé D, and Kushnir S. γ-Glutamyl transpeptidase in transgenic tobacco plants. Cellular localization, processing, and biochemical properties. Plant Physiol 128: 1109-1119, 2002.

152.

Swindell WR, Huebner M, and Weber AP. Transcriptional profiling of Arabidopsis heat shock proteins and transcription factors reveals extensive overlap between heat and non-heat stress response pathways. BMC Genomics 8: 125, 2007.

47

Page 48 of 63


48 153.

Tada Y, Spoel SH, Pajerowska-Mukhtar K, Mou Z, Song J, Wang C, Zuo J, and Dong X. Plant immunity requires conformational changes of NPR1 via S-nitrosylation and thioredoxins. Science 321: 952-956, 2008.

154.

Tanaka Y, Sasaki N, and Ohmiya A. Biosynthesis of plant pigments: anthocyanins, betalains and carotenoids. Plant J 54: 733-749, 2008.

155.

Tang G, Qin J, Dolnikowski GG, Russell RM, and Grusak MA. Golden Rice is an effective source of vitamin A. Am J Clin Nutr 89: 1776-1783, 2009.

156.

Tang W, Tu L, Yang X, Tan J, Deng F, Hao J, Guo K, Lindsey K, and Zhang X. The calcium sensor GhCaM7 promotes cotton fiber elongation by modulating reactive oxygen species (ROS) production. New Phytol 202: 509-520, 2014.

157.

Tischer W and Strotmann H. Relationship between inhibitor binding by chloroplasts and inhibition of photosynthetic electron transport. Biochim Biophys Acta - Bioenerg 460: 113-125, 1977.

158.

Tognetti VB, Mühlenbock P, and Van Breusegem F. Stress homeostasis - the redox and auxin perspective. Plant Cell Environ 35: 321-333, 2012.

159.

Tognetti VB, Palatnik JF, Fillat MF, Melzer M, Hajirezaei M-R, Valle EM, and Carrillo N. Functional replacement of ferredoxin by a cyanobacterial flavodoxin in tobacco confers broad-range stress tolerance. Plant Cell 18: 2035-2050, 2006.

160.

Tognetti VB, Van Aken O, Morreel K, Vandenbroucke K, van de Cotte B, De Clercq I, Chiwocha S, Fenske R, Prinsen E, Boerjan W, Genty B, Stubbs KA, InzéD, and Van Breusegem F. Perturbation of indole-3-butyric acid homeostasis by the UDPglucosyltransferase UGT74E2 modulates Arabidopsis architecture and water stress tolerance. Plant Cell 22: 2660-2679, 2010.

48

Page 49 of 63


49 161.

Torres MA, Dangl JL, and Jones JDG. Arabidopsis gp91phox homologues AtrbohD and AtrbohF are required for accumulation of reactive oxygen intermediates in the plant defense response. Proc Natl Acad Sci USA 99: 517-522, 2002.

162.

Triantaphylidès C, Krischke M, Hoeberichts FA, Ksas B, Gresser G, Havaux M, Van Breusegem F, and Mueller MJ. Singlet oxygen is the major reactive oxygen species involved in photooxidative damage to plants. Plant Physiol 148: 960-968, 2008.

163.

Tron AE, Bertoncini CW, Chan RL, and Gonzalez DH. Redox regulation of plant homeodomain transcription factors. J Biol Chem 277: 34800-34807, 2002.

164.

Tsukagoshi H, Busch W, and Benfey PN. Transcriptional regulation of ROS controls transition from proliferation to differentiation in the root. Cell 143: 606-616, 2010.

165.

Ueoka-Nakanishi H, Sazuka T, Nakanishi Y, Maeshima M, Mori H, and Hisabori T. Thioredoxin h regulates calcium dependent protein kinases in plasma membranes. FEBS J 280: 3220-3231, 2013.

166.

Vacca RA, de Pinto MC, Valenti D, Passarella S, Marra E, and De Gara L. Production of reactive oxygen species, alteration of cytosolic ascorbate peroxidase, and impairment of mitochondrial metabolism are early events in heat shock-induced programmed cell death in tobacco Bright-Yellow 2 cells. Plant Physiol 134: 11001112, 2004.

167.

Van Aken O, Van Breusegem F, Tognetti V, and De Clercq I. Methods and means for the production of plants with improved stress resistance (WO2008155139). Belgium, VIB VZW and Universiteit Gent, 2009.

168.

Van Breusegem F, Slooten L, Stassart J-M, Moens T, Botterman J, Van Montagu M, and InzéD. Overproduction of Arabidopsis thaliana FeSOD confers oxidative stress tolerance to transgenic maize. Plant Cell Physiol 40: 515-523, 1999.

49

Page 50 of 63


50 169.

Van Laer K, Hamilton CJ, and Messens J. Low-molecular-weight thiols in thioldisulfide exchange. Antioxid Redox Signal 18: 1642-1653, 2013.

170.

Vandenabeele S, Vanderauwera S, Vuylsteke M, Rombauts S, Langebartels C, Seidlitz HK, Zabeau M, Van Montagu M, InzéD, and Van Breusegem F. Catalase deficiency drastically affects gene expression induced by high light in Arabidopsis thaliana. Plant J 39: 45-58, 2004.

171.

Vanderauwera S, Zimmermann P, Rombauts S, Vandenabeele S, Langebartels C, Gruissem W, Inzé D, and Van Breusegem F. Genome-wide analysis of hydrogen peroxide-regulated gene expression in Arabidopsis reveals a high light-induced transcriptional cluster involved in anthocyanin biosynthesis. Plant Physiol 139: 806821, 2005.

172.

Vanderschuren H, Nyaboga E, Poon JS, Baerenfaller K, Grossmann J, HirschHoffmann M, Kirchgessner N, Nanni P, and Gruissem W. Large-scale proteomics of the cassava storage root and identification of a target gene to reduce postharvest deterioration. Plant Cell 26: 1913-1924, 2014.

173.

Vanneste S and Friml J. Auxin: a trigger for change in plant development. Cell 136: 1005-1016, 2009.

174.

Vanstraelen M and Benková E. Hormonal interactions in the regulation of plant development. Annu Rev Cell Dev Biol 28: 463-487, 2012.

175.

Vernoux T, Wilson RC, Seeley KA, Reichheld J-P, Muroy S, Brown S, Maughan SC, Cobbett CS, Van Montagu M, Inzé D, May MJ, and Sung ZR. The ROOT MERISTEMLESS1/CADMIUM SENSITIVE2 gene defines a glutathione-dependent pathway involved in initiation and maintenance of cell division during postembryonic root development. Plant Cell 12: 97-109, 2000.

50

Page 51 of 63


51 176.

Viola IL, Güttlein LN, and Gonzalez DH. Redox modulation of plant developmental regulators from the class I TCP transcription factor family. Plant Physiol 162: 14341447, 2013.

177.

Vivancos PD, Dong Y, Ziegler K, Markovic J, Pallardó FV, Pellny TK, Verrier PJ, and Foyer CH. Recruitment of glutathione into the nucleus during cell proliferation adjusts whole-cell redox homeostasis in Arabidopsis thaliana and lowers the oxidative defence shield. Plant J 64: 825-838, 2010.

178.

Vivancos PD, Wolff T, Markovic J, PallardóFV, and Foyer CH. A nuclear glutathione cycle within the cell cycle. Biochem J 431: 169-178, 2010.

179.

Volkov RA, Panchuk II, Mullineaux PM, and Schöffl F. Heat stress-induced H2O2 is required for effective expression of heat shock genes in Arabidopsis. Plant Mol Biol 61: 733-746, 2006.

180.

Volrath SL, Johnson MA, Ward ER, and Heifetz PB. Herbicide-tolerant protoporphyrinogen oxidase ("protox") genes (US6084155). Switzerland, Novartis AG, 2000.

181.

Wang F-Z, Wang Q-B, Kwon S-Y, Kwak S-S, and Su W-A. Enhanced drought tolerance of transgenic rice plants expressing a pea manganese superoxide dismutase. J Plant Physiol 162: 465-472, 2005.

182.

Wang L, Samac DA, Shapir N, Wackett LP, Vance CP, Olszewski NE, and Sadowsky MJ. Biodegradation of atrazine in transgenic plants expressing a modified bacterial atrazine chlorohydrolase (atzA) gene. Plant Biotechnol J 3: 475-486, 2005.

183.

Wang PT and Song C-P. Guard-cell signalling for hydrogen peroxide and abscisic acid. New Phytol 178: 703-718, 2008.

184.

Whatley FR, Tagawa K, and Arnon DI. Separation of the light and dark reactions in electron transfer during photosynthesis. Proc Natl Acad Sci USA 49: 266-270, 1963.

51

Page 52 of 63


52 185.

Wingler K, Hermans JJR, Schiffers P, Moens AL, Paul M, and Schmidt HHHW. NOX1, 2, 4, 5: counting out oxidative stress. Br J Pharmacol 164: 866-883, 2011.

186.

Wong CM, Marcocci L, Liu L, and Suzuki YJ. Cell signaling by protein carbonylation and decarbonylation. Antioxid Redox Signal 12: 393-404, 2010.

187.

World Health Organization (WHO). Micronutrient deficiencies: Vitamin a deficiency (http://www.who.int/nutrition/topics/vad/en/).

188.

Wright TR, Shan G, Walsh TA, Lira JM, Cui C, Song P, Zhuang M, Arnold NL, Lin G, Yau K, Russell SM, Cicchillo RM, Peterson MA, Simpson DM, Zhou N, Ponsamuel J, and Zhang Z. Robust crop resistance to broadleaf and grass herbicides provided by aryloxyalkanoate dioxygenase transgenes. Proc Natl Acad Sci USA 107: 20240-20245, 2010.

189.

Wu G, Shortt BJ, Lawrence EB, Levine EB, Fitzsimmons KC, and Shah DM. Disease resistance conferred by expression of a gene encoding H2O2-generating glucose oxidase in transgenic potato plants. Plant Cell 7: 1357-1368, 1995.

190.

Wu Y, Zhang D, Chu JY, Boyle P, Wang Y, Brindle ID, De Luca V, and Després C. The Arabidopsis NPR1 protein is a receptor for the plant defense hormone salicylic acid. Cell Reports 1: 639-647, 2012.

191.

Xu J, Duan X, Yang J, Beeching JR, and Zhang P. Enhanced reactive oxygen species scavenging by overproduction of superoxide dismutase and catalase delays postharvest physiological deterioration of cassava storage roots. Plant Physiol 161: 1517-1528, 2013.

192.

Yang Z and Ohlrogge JB. Turnover of fatty acids during natural senescence of Arabidopsis, Brachypodium, and switchgrass and in Arabidopsis β-oxidation mutants. Plant Physiol 150: 1981-1989, 2009.

52

Page 53 of 63


53 193.

Ye X, Al-Babili S, Klöti A, Zhang J, Lucca P, Beyer P, and Potrykus I. Engineering the provitamin A (β-carotene) biosynthetic pathway into (carotenoid-free) rice endosperm. Science 287: 303-305, 2000.

194.

Yoshioka H, Numata N, Nakajima K, Katou S, Kawakita K, Rowland O, Jones JDG, and Doke N. Nicotiana benthamiana gp91phox homologs NbrbohA and NbrbohB participate in H2O2 accumulation and resistance to Phytophthora infestans. Plant Cell 15: 706-718, 2003.

195.

Zaffagnini M, Bedhomme M, Marchand CH, Morisse S, Trost P, and Lemaire SD. Redox regulation in photosynthetic organisms: focus on glutathionylation. Antioxid Redox Signal 16: 567-586, 2012.

196.

Zhang Y, Butelli E, De Stefano R, Schoonbeek H-j, Magusin A, Pagliarani C, Wellner N, Hill L, Orzaez D, Granell A, Jones JDG, and Martin C. Anthocyanins double the shelf life of tomatoes by delaying overripening and reducing susceptibility to gray mold. Curr Biol 23: 1094-1100, 2013.

197.

Zurbriggen MD, Tognetti VB, and Carrillo N. Stress-inducible flavodoxin from photosynthetic microorganisms. The mystery of flavodoxin loss from the plant genome. IUBMB Life 59: 355-360, 2007.

53

Page 54 of 63


54 Figure legends

Figure 1. Main sites of ROS production in the plant cell. NADPH oxidases (Respiratory Burst Oxidase Homologs, Rbohs) and apoplastic peroxidases actively produce ROS within the apoplastic space needed for effective defense against invading pathogens and cell wall lignification. When illuminated, the electron transport chains in the chloroplasts (cETCs) generate singlet oxygen (1O2) and superoxide (O2•-) at the level of photosystem II (PSII) and PSI, respectively. Superoxide is further converted to H2O2 by the action of iron (Fe) and copper/zinc (Cu/Zn) superoxide dismutases (SODs). The levels of H2O2 are controlled by the glutathione (GSH) – ascorbate (ASC) cycle. Dashed lines indicate the cyclic electron flow around PSI affecting the reduction state of cETC together with plastoquinol terminal oxidase (PTOX). Electron flow via the mitochondrial electron transport chains (mETC) results in O2•production, which is processed to H2O2 by manganese (Mn) SOD. Plants possess a mitochondrial alternative oxidase (AOX), which is activated under various stress conditions and serves as a terminal oxidase believed to alleviate ROS accumulation. A significant part of the peroxisomal/glyoxysomal matrtix is occupied by catalase removing H2O2 generated during photorespiration and fatty acid β-oxidation. Many herbicides target crucial chloroplast components which lead to excessive ROS formation and cell death. Depicted are the sites of action of DCMU (3-(3,4-dichlorophenyl)-1,1-dimethylurea), triazine, paraquat, bromoxynil and protoporphyrinogen oxidase (PPO, Protox) inhibiting herbicides.

54

Page 55 of 63


55

Figure 2. Simplified depiction of ROS producing mechanisms specific for plants and shared between plants and mammals. NOX, NADPH oxidase; Rboh, Respiratory Burst Oxidase Homologs; GOX, Glycolate Oxidase; PSII, Photosystem I; PSI, photosystem I;1O2, singlet oxygen; O2•-, superoxide; H2O2, hydrogen peroxide.

55

Page 56 of 63


56

Figure 3. Major enzymatic antioxidant systems scavenging H2O2 in plants. The levels of H2O2 are tightly controlled by catalase (CAT), ascorbate peroxidase (APX), glutathione peroxidase (GPX), and by thioredoxin (Trx) or glutaredoxin (Grx)-dependent peroxiredoxins (Prx). The enzymatic activity of APX results in oxidation of ascorbate (ASC) to monodehydroascorbate

(MDA)

which

could

be

reduced

back

to

ASC

by

monodehydroascorbate reductase (MDAR) or further disproportionates into ASC and dehydroascorbate (DHA). Dehydroascorbate reductase (DHAR) reduces DHA to ASC by exploiting reducing equivalents from glutathione (GSH). The resulting oxidized glutathione (GSSG) is reduced by glutathione reductase (GR).

56

Page 57 of 63


57

Figure 4. Chloroplastic photorespiratory bypasses. A) Strategy applied by Kebeish et al. (63) utilizes glycolate dehydrogenase (GDH), glyoxylate carboxylyase (GCL) and tartronic semialdehyde reductase (TSR) to convert the photorespiratory glycolate to glycerate within chloroplasts. B) Glycolate catabolic pathway described by Maier et al. (89). A series of reactions involving transgenically introduced glycolate oxidase (GOX), catalase (CAT) and malate synthase (MS) leads to formation of malate that is subsequently processed by endogenous malic enzyme (ME) and pyruvate dehydrogenase (PDH) with the release of CO2 and NAD(P)H. Asterisks (*) indicate enzymes introduced by transgenesis. RuBisCo, ribulose1,5-biphosphate carboxylase/oxygenase; PGLP, 2-phosphoglycolate phosphatase; GGT, glutamate-glyoxylate aminotransferase; GDC, glycine decarboxylase; SHMT, serine hydroxymethyltransferase; SGT serine-glyoxylate aminotransferase; HPR, hydroxypyruvate

57

Page 58 of 63


58 reductase;

GLYK,

glycerate

kinase;

RuBP,

ribulose

1,5-biphosphate;

2-PG,

2-

phosphoglycolate; 2OG, 2-oxoglutarate; 3-PGA, 3-phosphoglycerate; G3P, glyceraldehyde 3phosphate. “(To see this illustration in color the reader is referred to the web version of this article at www.liebertonline.com/ars)”

58

Page 59 of 63


59

Figure 5. Arabidopsis mutants lacking CATALASE 2 have been widely used to study the effects of elevated peroxisomal H2O2 levels. Three-week-old plants (16-h/8-h light/dark; light intensity 120-130 μmol·m−2·s−1), grown under high CO2 levels (3000 ppm) in order to suppress the flux via the photorespiratory pathway, were transferred to conditions favoring photorespiration (ambient air and high light irradiation (1100-1200 μmol·m−2·s−1)). Representative images demonstrate formation of cell death lesions and reduction of PSII maximum efficiency (Fv'/Fm') upon induction of the photorespiratory pathway. (Waszczak et al., unpublished data).

59

Page 60 of 63


60

Figure 6. Oxidative post-translational modifications of proteins on specific methionine (Met), cysteine (Cys) and lysine (Lys) amino acid residues. Oxidative attack on methionine leads to the formation of methionine sulfoxide (Met-SO) and rarely methionine sulfone. Met-SO can be reduced by methionine sulfoxide reductases (MsrA/B) that rely on thioredoxins as electron source. Exposure of redox-sensitive cysteine residues to ROS leads to reversible sulfenic acid formation that, unless stabilized within the protein environment, reacts with nearby thiols of the same protein to form intramolecular disulfide bonds, or with thiols from other proteins or glutathione (GSH) to form mixed disulfide bonds. These thiol modifications are reversible and can be reduced either by glutaredoxin (Grx), or by thioredoxin (Trx). In the presence of high levels of ROS, overoxidation to sulfinic or sulfonic acid can occur. While in some cases sulfinic acids can be reduced by sulfiredoxin (Srx), formation of sulfonic acids leads to irreversible protein damage.

60

Page 61 of 63


61

Figure 7. Schematic representation of NPR1-based plant immune signaling. Pathogentriggered oxidative burst mediated by salicylic acid (SA) accumulation leads to monomerization of NPR1, which under non-stressed conditions resides in the cytosol as an oligomer stabilized by disulfide bonds. The monomerization is mediated by thioredoxin (Trx). Upon nuclear import, NPR1 interacts with TGA transcription factors to activate the expression of defense genes. Formation of an intramolecular disulfide bridge on TGA1 hinders its binding to DNA. Reduction of its disulfide bond enhances the interaction with NPR1 and DNA binding. The cysteine residues of TGA1 can undergo S-nitrosylation (SNO) or S-glutathionylation (S-SG), which promotes its DNA binding activity.

61

Page 62 of 63


62

Figure 8. Redox-mediated events leading to stomatal closure. Perception of abscisic acid (ABA) by PYR/PYL/RCAR (PYRABACTIN RESISTANCE/PYR1 LIKE/REGULATORY COMPONENT OF ABA RECEPTOR) receptor proteins inactivates the PP2C (PROTEIN PHOSPHATASE 2C) protein phosphatase family members ABI1 (ABA insensitive 1) and ABI2. Oxidative modifications mediated by GLUTATHIONE PEROXIDASE 3 (GPX3) have been show to similarly inactivate their phosphatase activity. The inhibition of PP2C activates SnRK type 2 family of kinases (SnRK2) which regulate ABA-related gene expression and ion channels. The SnRK2 family member OST1 phosphorylates the ROS-producing enzymes RbohD/F. The ABA-triggered increase of cytosolic Ca2+ levels, which is dependent on H2O2 accumulation, activates Ca2+-dependent protein kinases (CPKs) and calcineurin B-like (CBL) calcium sensors leading to the stimulation of slow anion channel associated 1 (SLAC1).

62

Antioxidants & Redox Signaling Redox Strategies for Crop Improvement (doi: 10.1089/ars.2014.6033) This article has been peer-reviewed and accepted for publication, but has yet to undergo copyediting and proof correction. The final published version may differ from this proof. Page 63 of 63

63

63

Molecular identification of sex in Simarouba glauca by RAPD markers for crop improvement strategies.

Cisgenics - a sustainable approach for crop improvement.

Untapping root system architecture for crop improvement.

Call for crop improvement analysis articles.

Physiological phenotyping of plants for crop improvement.

Barley genetic variation: implications for crop improvement.

Plant genetic engineering for crop improvement.

Crop improvement through tissue culture.

Green biotechnology, nanotechnology and bio-fortification: perspectives on novel environment-friendly crop improvement strategies.

Lessons from Domestication: Targeting Cis-Regulatory Elements for Crop Improvement.

Genetic improvement for root growth angle to enhance crop production.

Genome editing for crop improvement: Challenges and opportunities.

Perspectives and Challenges of Microbial Application for Crop Improvement.

Emerging paradigms in genomics-based crop improvement.

Emerging strategies for enhancing crop resistance to microbial pathogens.

Quality improvement strategies for critical care nursing.

Connecting Biochemical Photosynthesis Models with Crop Models to Support Crop Improvement.

Regulating Subcellular Metal Homeostasis: The Key to Crop Improvement.

Lipase improvement: goals and strategies.

Plant exomics: concepts, applications and methodologies in crop improvement.

RNA interference: concept to reality in crop improvement.

RiceMetaSys for salt and drought stress responsive genes in rice: a web interface for crop improvement.

Mortality meetings in geriatric medicine: strategies for improvement.

Future-proof crops: challenges and strategies for climate resilience improvement.