Dataset Information

Thiol Redox Regulation of Plant ?-Carbonic Anhydrase.

ABSTRACT: ?-carbonic anhydrases (?CA) accelerate the equilibrium formation between CO2 and carbonate. Two plant ?CA isoforms are targeted to the chloroplast and represent abundant proteins in the range of >1% of chloroplast protein. While their function in gas exchange and photosynthesis is well-characterized in carbon concentrating mechanisms of cyanobacteria and plants with C4-photosynthesis, their function in plants with C3-photosynthesis is less clear. The presence of conserved and surface-exposed cysteinyl residues in the ?CA-structure urged to the question whether ?CA is subject to redox regulation. Activity measurements revealed reductive activation of ?CA1, whereas oxidized ?CA1 was inactive. Mutation of cysteinyl residues decreased ?CA1 activity, in particular C280S, C167S, C230S, and C257S. High concentrations of dithiothreitol or low amounts of reduced thioredoxins (TRXs) activated oxidized ?CA1. TRX-y1 and TRX-y2 most efficiently activated ?CA1, followed by TRX-f1 and f2 and NADPH-dependent TRX reductase C (NTRC). High light irradiation did not enhance ?CA activity in wildtype Arabidopsis, but surprisingly in ?ca1 knockout plants, indicating light-dependent regulation. The results assign a role of ?CA within the thiol redox regulatory network of the chloroplast.

SUBMITTER: Dreyer A

PROVIDER: S-EPMC7463553 | biostudies-literature | 2020 Jul

REPOSITORIES: biostudies-literature

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Thiol Redox Regulation of Plant β-Carbonic Anhydrase.

Dreyer Anna A Schackmann Alexander A Kriznik Alexandre A Chibani Kamel K Wesemann Corinna C Vogelsang Lara L Beyer André A Dietz Karl-Josef KJ

Biomolecules 20200730 8

β-carbonic anhydrases (βCA) accelerate the equilibrium formation between CO<sub>2</sub> and carbonate. Two plant βCA isoforms are targeted to the chloroplast and represent abundant proteins in the range of >1% of chloroplast protein. While their function in gas exchange and photosynthesis is well-characterized in carbon concentrating mechanisms of cyanobacteria and plants with C4-photosynthesis, their function in plants with C3-photosynthesis is less clear. The presence of conserved and surface- ...[more]

PMID: 32751472

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Project description:The Rac1 GTPase is an essential and ubiquitous protein that signals through numerous pathways to control critical cellular processes, including cell growth, morphology, and motility. Rac1 deletion is embryonic lethal, and its dysregulation or mutation can promote cancer, arthritis, cardiovascular disease, and neurological disorders. Rac1 activity is highly regulated by modulatory proteins and posttranslational modifications. Whereas much attention has been devoted to guanine nucleotide exchange factors that act on Rac1 to promote GTP loading and Rac1 activation, cellular oxidants may also regulate Rac1 activation by promoting guanine nucleotide exchange. Herein, we show that Rac1 contains a redox-sensitive cysteine (Cys(18)) that can be selectively oxidized at physiological pH because of its lowered pKa. Consistent with these observations, we show that Rac1 is glutathiolated in primary chondrocytes. Oxidation of Cys(18) by glutathione greatly perturbs Rac1 guanine nucleotide binding and promotes nucleotide exchange. As aspartate substitutions have been previously used to mimic cysteine oxidation, we characterized the biochemical properties of Rac1(C18D). We also evaluated Rac1(C18S) as a redox-insensitive variant and found that it retains structural and biochemical properties similar to those of Rac1(WT) but is resistant to thiol oxidation. In addition, Rac1(C18D), but not Rac1(C18S), shows greatly enhanced nucleotide exchange, similar to that observed for Rac1 oxidation by glutathione. We employed Rac1(C18D) in cell-based studies to assess whether this fast-cycling variant, which mimics Rac1 oxidation by glutathione, affects Rac1 activity and function. Expression of Rac1(C18D) in Swiss 3T3 cells showed greatly enhanced GTP-bound Rac1 relative to Rac1(WT) and the redox-insensitive Rac1(C18S) variant. Moreover, expression of Rac1(C18D) in HEK-293T cells greatly promoted lamellipodia formation. Our results suggest that Rac1 oxidation at Cys(18) is a novel posttranslational modification that upregulates Rac1 activity.

Dataset Information

Thiol Redox Regulation of Plant ?-Carbonic Anhydrase.

Publications

Thiol Redox Regulation of Plant β-Carbonic Anhydrase.

Similar Datasets

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