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Redox regulation of NADP-malate dehydrogenase is vital for land plants under fluctuating light environment.

ABSTRACT: Many enzymes involved in photosynthesis possess highly conserved cysteine residues that serve as redox switches in chloroplasts. These redox switches function to activate or deactivate enzymes during light-dark transitions and have the function of fine-tuning their activities according to the intensity of light. Accordingly, many studies on chloroplast redox regulation have been conducted under the hypothesis that "fine regulation of the activities of these enzymes is crucial for efficient photosynthesis." However, the impact of the regulatory system on plant metabolism is still unclear. To test this hypothesis, we here studied the impact of the ablation of a redox switch in chloroplast NADP-malate dehydrogenase (MDH). By genome editing, we generated a mutant plant whose MDH lacks one of its redox switches and is active even in dark conditions. Although NADPH consumption by MDH in the dark is expected to be harmful to plant growth, the mutant line did not show any phenotypic differences under standard long-day conditions. In contrast, the mutant line showed severe growth retardation under short-day or fluctuating light conditions. These results indicate that thiol-switch redox regulation of MDH activity is crucial for maintaining NADPH homeostasis in chloroplasts under these conditions.

SUBMITTER: Yokochi Y

PROVIDER: S-EPMC8017969 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:Background and aimsPlastid NADP-dependent malate dehydrogenase (MDH) catalyses the conversion of oxaloacetate to malate. In C4 plants, it is involved in photosynthetic carbon assimilation. In Poaceae, one NADP-MDH gene has been identified in rice (C3; Erhartoideae) and maize (C4; Panicoideae), whereas two tandemly repeated genes have been identified in Sorghum (C4; Panicoideae). In the present study, the molecular evolution of the NADP-MDH multigene family was investigated in order to analyse how the C4 isoform has evolved over a broader range of panicoid grasses.MethodsPolymerase chain reaction (PCR)-based cloning was used to isolate cDNAs encoding NADP-MDHs from 15 species of Panicoideae. A gene phylogeny was reconstructed based on cDNA sequences using distance and maximum parsimony methods. Episodic selection along some branches of the phylogenetic tree was tested by analysing non-synonymous and synonymous rate ratios. Transcription of NADP-MDH genes was compared in green leaves of five accessions of Saccharum, Sorghum and Vetiveria using a semi-quantitative PCR approach.Key resultsPhylogenetic analyses of these data support the existence of two NADP-MDH gene lineages (NMDH-I and NMDH-II) in several Andropogoneae (i.e. Saccharum, Sorghum and Vetiveria). Episodic positive selection was shown along the basal branch of the NMDH-II clade. Three amino acid modifications allow the two gene lineages to be distinguished, suggesting a positive selection at these sites. In green leaves, we showed that the transcript accumulation was higher for NMDH-I than for NMDH-II.ConclusionsIt is hypothesized that the maintenance of both NADP-MDH genes in some Andropogoneae is due to a partition of the original functions across both copies. NMDH-I probably corresponds to the C4 isoform as previously suggested. Nevertheless, some C4 species (e.g. maize) only have one gene which should be selected for its high expression level in leaves. This study confirms that gene duplicates have been recruited for C4 photosynthesis but are not required in every case.

Project description:Purpose: Overexpression of VDE, ZEP and PsbS (VPZ) in plants confers enhanced dynamics of non-photochemical quenching of NPQ under FL light. VPZ overexpression in tobacco has been reported to increase plant biomass under fluctuating light. However, our work demonstrates that such strategy had no advantageous effect when asssayed in Arabidopsis thaliana. To investigate whether this observation is due to impairements in signaling pathways due to the expression of VPZ in Arabidopsis thaliana, transcriptome profiles of VPZ plants exposed to fluctuating light or high light were conducted. Methods: mRNA-Seq libraries were prepared from 21-d-old Col0 plants and VPZ lines #2 and #4 cultivated under standard control conditions and then exposed to fluctuating light or high light for 6h and 30 h. Standard Illumina protocols were used for mRNA-Seq sequencing. Triplicates for 3 independent plants were used. Adaptor sequences were removed with Trimmomatic and resulting reads mapped to the Arabidopsis genome (Araport11) with Tophat 2.1.1. Normalised counts reads were quantified with featureCounts to provide processed data files and differential expression analysis were conducted with the DEseq2 pipeline. Results: mRNA-Seq libraries contained app. 9 milion reads each. Transcript analysis for differential gene expression was conducted according to the Tophat/featureCounts/DESeq2 pipeline. To declare significant differences, a cut-off for absolute log2(FC) ≥ 1 compared to the initial timepoint (0h) and FDR ≤ 0.05 was applied. Significant transcripts exclusively found for Col0 and both VPZ lines in each time-point were considered for subsequent analysis. Accordingly, 528 and 415 transcripts (6h and 30h) were found for Col0, and 326 and 896 (6h and 30h) for the VPZ under control conditions. For FL, 490 and 591 transcripts qualified as specific (6h and 30h) for Col0 and 938 and 401 (6h and 30h) for VPZ. Strikingly, the amount of transcripts raised to 2847 and 2902 DEGs (6h and 30h) for Col0, whereas only 546 and 304 (6h and 30h) were detected for VPZ under HL. transcripts). Conclusions: the overexpression of VPZ in Arabidopsis leads to particular transcriptome response under fluctuating light related to changes in photosynthesis and response to abscisic acid, among all. However, Col0 displayed wider transcriptome responses than VPZ after exposure to high light .

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Redox regulation of NADP-malate dehydrogenase is vital for land plants under fluctuating light environment.

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