The redox-sensitive transcription factor Rap2.4a controls nuclear expression of 2-Cys peroxiredoxin A and other chloroplast antioxidant enzymes

Shaikhali J, Heiber I, Seidel T, et al. The redox-sensitive transcription factor Rap2.4a controls nuclear expression of 2-Cys peroxiredoxin A and other chloroplast antioxidant enzymes. BMC Plant Biology. 2008;8(1):48

The redox imbalanced Mutants of Arabidopsis Differentiate Signaling Pathways for Redox Regulation of Chloroplast Antioxidant Enzymes

A network of enzymatic and nonenzymatic antioxidants protects chloroplasts from photooxidative damage. With all enzymatic components being nuclear encoded, the control of the antioxidant capacity depends on chloroplast-to-nucleus redox signaling. Using an Arabidopsis (Arabidopsis thaliana) reporter gene line expressing luciferase under control of the redox-sensitive 2-cysteine peroxiredoxin A (2CPA) promoter, six mutants with low 2CPA promoter activity were isolated, of which five mutants show limitations in redox-box regulation of the 2CPA promoter. In addition to 2CPA, the transcript levels for other chloroplast antioxidant enzymes were decreased, although a higher oxidation status of the ascorbate pool, a higher reduction state of the plastoquinone pool, and an increased oxidation status of the 2-Cys peroxiredoxin pool demonstrated photooxidative stress conditions. Greening of the mutants, chloroplast ultrastructure, steady-state photosynthesis, and the responses to the stress hormone abscisic acid were wild type like. In the rosette state, the mutants were more sensitive to low CO(2) and to hydrogen peroxide. Comparison of gene expression patterns and stress sensitivity characterizes the mutants as redox imbalanced in the regulation of nuclear-encoded chloroplast antioxidant enzymes and differentiates redox signaling cascades

Mitochondrial Malate Dehydrogenase Lowers Leaf Respiration and Alters Photorespiration and Plant Growth in Arabidopsis[W][OA]

Malate dehydrogenase (MDH) catalyzes a reversible NAD+-dependent-dehydrogenase reaction involved in central metabolism and redox homeostasis between organelle compartments. To explore the role of mitochondrial MDH (mMDH) in Arabidopsis (Arabidopsis thaliana), knockout single and double mutants for the highly expressed mMDH1 and lower expressed mMDH2 isoforms were constructed and analyzed. A mmdh1mmdh2 mutant has no detectable mMDH activity but is viable, albeit small and slow growing. Quantitative proteome analysis of mitochondria shows changes in other mitochondrial NAD-linked dehydrogenases, indicating a reorganization of such enzymes in the mitochondrial matrix. The slow-growing mmdh1mmdh2 mutant has elevated leaf respiration rate in the dark and light, without loss of photosynthetic capacity, suggesting that mMDH normally uses NADH to reduce oxaloacetate to malate, which is then exported to the cytosol, rather than to drive mitochondrial respiration. Increased respiratory rate in leaves can account in part for the low net CO2 assimilation and slow growth rate of mmdh1mmdh2. Loss of mMDH also affects photorespiration, as evidenced by a lower postillumination burst, alterations in CO2 assimilation/intercellular CO2 curves at low CO2, and the light-dependent elevated concentration of photorespiratory metabolites. Complementation of mmdh1mmdh2 with an mMDH cDNA recovered mMDH activity, suppressed respiratory rate, ameliorated changes to photorespiration, and increased plant growth. A previously established inverse correlation between mMDH and ascorbate content in tomato (Solanum lycopersicum) has been consolidated in Arabidopsis and may potentially be linked to decreased galactonolactone dehydrogenase content in mitochondria in the mutant. Overall, a central yet complex role for mMDH emerges in the partitioning of carbon and energy in leaves, providing new directions for bioengineering of plant growth rate and a new insight into the molecular mechanisms linking respiration and photosynthesis in plants

Plastidial glutaredoxins: glutathione-dependent enzymes involved in detoxification, redox signalling and iron-sulfur cluster biogenesis.

International audienceGlutaredoxins (Grxs) are oxidoreductases structurally related to thioredoxins (Trxs). They have two major proposed biochemical roles, the reduction of disulfide bonds and the binding of iron-sulfur (Fe-S) clusters, both of which usually involve glutathione. In photosynthetic organisms, Grxs are distributed into six classes. Several Grxs from classes I and II can indeed exist either as apoforms which display deglutathionylation activity or as holoforms which bind Fe-S clusters. Using biochemical, spectroscopic and structural approaches, we have characterized four plastidial Grxs, showing that three of these can bind an Fe-S cluster. Site-directed mutagenesis experiments and resolution of the crystal structure of class I Grxs (GrxC5 and S12) revealed the critical role of some active site residues for cluster formation and for protein activity. The deglutathionylation activity of these two Grxs proceeds through a monothiol mechanism, which is important for the regeneration of the thiol-peroxidase and methionine sulfoxide reductase families. The two plastidial class II Grxs (GrxS14 and S16) can bind more labile Fe-S clusters that can serve for the maturation of other proteins as demonstrated by in vitro Fe-S cluster transfer experiments and by the complementation of a yeast strain deleted for the mitochondrial Grx5. Using yeast two hybrid and bimolecular fluorescence complementation, we have shown that class II but not class I Grxs interact with plastidial BolA proteins, including SufE1, a sulfurtransferase required for the maturation of plastidial Fe-S proteins.Hence, we propose (i) that, based on its peculiar catalytic and thermodynamic properties, GrxS12 and potentially GrxC5 could act as redox sensors, and (ii) that GrxS14 and GrxS16 could participate to the maturation of iron-sulfur proteins either by being directly involved in Fe-S cluster trafficking or by having regulatory roles in particular via their interaction with BolA proteins

Plastidial glutaredoxins: glutathione-dependent enzymes involved in detoxification, redox signalling and iron-sulfur cluster biogenesis.

International audienceGlutaredoxins (Grxs) are oxidoreductases structurally related to thioredoxins (Trxs). They have two major proposed biochemical roles, the reduction of disulfide bonds and the binding of iron-sulfur (Fe-S) clusters, both of which usually involve glutathione. In photosynthetic organisms, Grxs are distributed into six classes. Several Grxs from classes I and II can indeed exist either as apoforms which display deglutathionylation activity or as holoforms which bind Fe-S clusters. Using biochemical, spectroscopic and structural approaches, we have characterized four plastidial Grxs, showing that three of these can bind an Fe-S cluster. Site-directed mutagenesis experiments and resolution of the crystal structure of class I Grxs (GrxC5 and S12) revealed the critical role of some active site residues for cluster formation and for protein activity. The deglutathionylation activity of these two Grxs proceeds through a monothiol mechanism, which is important for the regeneration of the thiol-peroxidase and methionine sulfoxide reductase families. The two plastidial class II Grxs (GrxS14 and S16) can bind more labile Fe-S clusters that can serve for the maturation of other proteins as demonstrated by in vitro Fe-S cluster transfer experiments and by the complementation of a yeast strain deleted for the mitochondrial Grx5. Using yeast two hybrid and bimolecular fluorescence complementation, we have shown that class II but not class I Grxs interact with plastidial BolA proteins, including SufE1, a sulfurtransferase required for the maturation of plastidial Fe-S proteins.Hence, we propose (i) that, based on its peculiar catalytic and thermodynamic properties, GrxS12 and potentially GrxC5 could act as redox sensors, and (ii) that GrxS14 and GrxS16 could participate to the maturation of iron-sulfur proteins either by being directly involved in Fe-S cluster trafficking or by having regulatory roles in particular via their interaction with BolA proteins

Plastidial glutaredoxins: glutathione-dependent enzymes involved in detoxification, redox signalling and iron-sulfur cluster biogenesis.

International audienceGlutaredoxins (Grxs) are oxidoreductases structurally related to thioredoxins (Trxs). They have two major proposed biochemical roles, the reduction of disulfide bonds and the binding of iron-sulfur (Fe-S) clusters, both of which usually involve glutathione. In photosynthetic organisms, Grxs are distributed into six classes. Several Grxs from classes I and II can indeed exist either as apoforms which display deglutathionylation activity or as holoforms which bind Fe-S clusters. Using biochemical, spectroscopic and structural approaches, we have characterized four plastidial Grxs, showing that three of these can bind an Fe-S cluster. Site-directed mutagenesis experiments and resolution of the crystal structure of class I Grxs (GrxC5 and S12) revealed the critical role of some active site residues for cluster formation and for protein activity. The deglutathionylation activity of these two Grxs proceeds through a monothiol mechanism, which is important for the regeneration of the thiol-peroxidase and methionine sulfoxide reductase families. The two plastidial class II Grxs (GrxS14 and S16) can bind more labile Fe-S clusters that can serve for the maturation of other proteins as demonstrated by in vitro Fe-S cluster transfer experiments and by the complementation of a yeast strain deleted for the mitochondrial Grx5. Using yeast two hybrid and bimolecular fluorescence complementation, we have shown that class II but not class I Grxs interact with plastidial BolA proteins, including SufE1, a sulfurtransferase required for the maturation of plastidial Fe-S proteins.Hence, we propose (i) that, based on its peculiar catalytic and thermodynamic properties, GrxS12 and potentially GrxC5 could act as redox sensors, and (ii) that GrxS14 and GrxS16 could participate to the maturation of iron-sulfur proteins either by being directly involved in Fe-S cluster trafficking or by having regulatory roles in particular via their interaction with BolA proteins

The multiple facets of glutaredoxins: one fold, several partners and functions

International audienceGlutaredoxins (Grxs) are oxidoreductases structurally related to thioredoxins (Trxs). They have two major proposed biochemical roles, the reduction of disulfide bonds and the binding of iron-sulfur (Fe-S) clusters, both of which usually involve glutathione. In photosynthetic organisms, Grxs are distributed into six classes. Several Grxs from classes I and II can indeed exist either as apoforms which display deglutathionylation activity or as holoforms which bind Fe-S clusters. Using biochemical, spectroscopic and structural approaches, we have characterized four plastidial Grxs, showing that three of these can bind an Fe-S cluster. Site-directed mutagenesis experiments and resolution of the crystal structure of class I Grxs (GrxC5 and S12) revealed the critical role of some active site residues for cluster formation and for protein activity. The deglutathionylation activity of these two Grxs proceeds through a monothiol mechanism, which is important for the regeneration of the thiol-peroxidase and methionine sulfoxide reductase families. The two plastidial class II Grxs (GrxS14 and S16) can bind more labile Fe-S clusters that can serve for the maturation of other proteins as demonstrated by in vitro Fe-S cluster transfer experiments and by the complementation of a yeast strain deleted for the mitochondrial Grx5. Using binary yeast two hybrid and bimolecular fluorescence complementation experiments, we have shown that class II but not class I Grxs interact with plastidial BolA proteins, including SufE1, a sulfurtransferase required for the maturation of plastidial Fe-S proteins. Finally, using spectroscopic and structural approaches, we have determined that the Grx-BolA couples form two types of complexes involving distinct regions of both partners. Hence, we propose (i) that, based on its peculiar catalytic and thermodynamic properties, GrxS12 and potentially GrxC5 could rather act as oxidoreductases and/or redox sensors, and (ii) that GrxS14 and GrxS16 could participate to the maturation of iron-sulfur proteins either by being directly involved in Fe-S cluster trafficking or by having regulatory roles in particular via their interaction with BolA proteins

The multiple facets of glutaredoxins: one fold, several partners and functions

International audienceGlutaredoxins (Grxs) are oxidoreductases structurally related to thioredoxins (Trxs). They have two major proposed biochemical roles, the reduction of disulfide bonds and the binding of iron-sulfur (Fe-S) clusters, both of which usually involve glutathione. In photosynthetic organisms, Grxs are distributed into six classes. Several Grxs from classes I and II can indeed exist either as apoforms which display deglutathionylation activity or as holoforms which bind Fe-S clusters. Using biochemical, spectroscopic and structural approaches, we have characterized four plastidial Grxs, showing that three of these can bind an Fe-S cluster. Site-directed mutagenesis experiments and resolution of the crystal structure of class I Grxs (GrxC5 and S12) revealed the critical role of some active site residues for cluster formation and for protein activity. The deglutathionylation activity of these two Grxs proceeds through a monothiol mechanism, which is important for the regeneration of the thiol-peroxidase and methionine sulfoxide reductase families. The two plastidial class II Grxs (GrxS14 and S16) can bind more labile Fe-S clusters that can serve for the maturation of other proteins as demonstrated by in vitro Fe-S cluster transfer experiments and by the complementation of a yeast strain deleted for the mitochondrial Grx5. Using binary yeast two hybrid and bimolecular fluorescence complementation experiments, we have shown that class II but not class I Grxs interact with plastidial BolA proteins, including SufE1, a sulfurtransferase required for the maturation of plastidial Fe-S proteins. Finally, using spectroscopic and structural approaches, we have determined that the Grx-BolA couples form two types of complexes involving distinct regions of both partners. Hence, we propose (i) that, based on its peculiar catalytic and thermodynamic properties, GrxS12 and potentially GrxC5 could rather act as oxidoreductases and/or redox sensors, and (ii) that GrxS14 and GrxS16 could participate to the maturation of iron-sulfur proteins either by being directly involved in Fe-S cluster trafficking or by having regulatory roles in particular via their interaction with BolA proteins