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Iodoacetyl tandem mass tag (iodoTMT)-based redox proteomics of Arabidopsis thaliana

ABSTRACT: Fluctuating light affects photosynthesis in chloroplasts, resulting changes of a series of redox reactions and accumulation of reactive oxygen species (ROS). Chloroplast redox regulation have been considered as “a fine regulation of protein function which is crucial for efficient photosynthesis”. However, the role of redox regulation in acclimation to fluctuating light for plants is still unclear. In this study, we performed global quantitative mapping of the Arabidopsis thaliana (wild-type and pgr5 mutant) cysteine thiol switching using the latest iodoTMT-based redox proteomics technology, systematically revealing a high-quality landscape of fluctuating light-responsive redox-modified proteins for the first time. Notably, photosynthesis-related pathway, especially PSI-related proteins, are operational thiol-switching hotspots.

ORGANISM(S): Arabidopsis Thaliana

SUBMITTER: Hongbin Wang

PROVIDER: PXD029772 | iProX | Wed Nov 17 00:00:00 GMT 2021

REPOSITORIES: iProX

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Project description:In bacteria, the biosynthesis of cysteine is accomplished by two enzymes that are encoged by the cysK and cysM genes. CysM is also able to incorporate thiosulfate to produce S-sulfocysteine. In plant cells, the biosynthesis of cysteine occurs in the cytosol, mitochondria and chloroplasts. Chloroplasts contain two O-acetylserine(thiol)lyase homologs, which are encoded by the OAS-B and CS26 genes. An in vitro enzymatic analysis of the recombinant CS26 protein demonstrated that this isoform possesses S-sulfocysteine synthase activity and lacks O-acetylserine(thiol)lyase activity. In vivo functional analysis of this enzyme in knockout mutants demonstrated that mutation of cs26 suppressed the S-sulfocysteine synthase activity that was detected in wild type; furthermore, the mutants exhibited a growth phenotype, but penetrance depended on the light regime. The cs26 mutant plants also had reductions in chlorophyll content and photosynthetic activity (neither of which were observed in oas-b mutants), as well as elevated glutathione levels. However, cs26 leaves were not able to properly detoxify ROS, which accumulated to high levels under long-day growth conditions. The transcriptional profile of the cs26 mutant revealed that the mutation had a pleiotropic effect on many cellular and metabolic processes. Our finding reveals that S-sulfocysteine and the activity of S-sulfocysteine synthase play an important role in chloroplast function and are essential for light-dependent redox regulation within the chloroplast. Using the Affymetrix ATH1 GeneChips, we performed a comparative transcriptomic analysis on leaves of the cs26 and wild type plants under two different photoperiod conditions. Wild type and cs26 mutant plants were grown on soil under a long-day photoperiod (LD) or under a short-day photoperiod (SD). Total RNA was extracted from the leaves of 3-week-old plants grown under identical LD conditions, and from the leaves of 5-week-old plants grown under identical SD conditions. Three biological replicates were performed for each sample and hybridized to the chips. We made two different comparisons to classify the differently expressed genes in the mutant plant: cs26 leaves under LD versus wild-type leaves under LD and cs26 leaves under SD versus wild-type leaves under SD.

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Iodoacetyl tandem mass tag (iodoTMT)-based redox proteomics of Arabidopsis thaliana

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