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Proteome-wide light/dark modulation of thiol oxidation in cyanobacteria revealed by quantitative site-specific redox proteomics.

ABSTRACT: Reversible protein thiol oxidation is an essential regulatory mechanism of photosynthesis, metabolism, and gene expression in photosynthetic organisms. Herein, we present proteome-wide quantitative and site-specific profiling of in vivo thiol oxidation modulated by light/dark in the cyanobacterium Synechocystis sp. PCC 6803, an oxygenic photosynthetic prokaryote, using a resin-assisted thiol enrichment approach. Our proteomic approach integrates resin-assisted enrichment with isobaric tandem mass tag labeling to enable site-specific and quantitative measurements of reversibly oxidized thiols. The redox dynamics of ?2,100 Cys-sites from 1,060 proteins under light, dark, and 3-(3,4-dichlorophenyl)-1,1-dimethylurea (a photosystem II inhibitor) conditions were quantified. In addition to relative quantification, the stoichiometry or percentage of oxidation (reversibly oxidized/total thiols) for ?1,350 Cys-sites was also quantified. The overall results revealed broad changes in thiol oxidation in many key biological processes, including photosynthetic electron transport, carbon fixation, and glycolysis. Moreover, the redox sensitivity along with the stoichiometric data enabled prediction of potential functional Cys-sites for proteins of interest. The functional significance of redox-sensitive Cys-sites in NADP-dependent glyceraldehyde-3-phosphate dehydrogenase, peroxiredoxin (AhpC/TSA family protein Sll1621), and glucose 6-phosphate dehydrogenase was further confirmed with site-specific mutagenesis and biochemical studies. Together, our findings provide significant insights into the broad redox regulation of photosynthetic organisms.

SUBMITTER: Guo J

PROVIDER: S-EPMC4256482 | biostudies-literature | 2014 Dec

REPOSITORIES: biostudies-literature

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Proteome-wide light/dark modulation of thiol oxidation in cyanobacteria revealed by quantitative site-specific redox proteomics.

Guo Jia J Nguyen Amelia Y AY Dai Ziyu Z Su Dian D Gaffrey Matthew J MJ Moore Ronald J RJ Jacobs Jon M JM Monroe Matthew E ME Smith Richard D RD Koppenaal David W DW Pakrasi Himadri B HB Qian Wei-Jun WJ

Molecular & cellular proteomics : MCP 20140812 12

Reversible protein thiol oxidation is an essential regulatory mechanism of photosynthesis, metabolism, and gene expression in photosynthetic organisms. Herein, we present proteome-wide quantitative and site-specific profiling of in vivo thiol oxidation modulated by light/dark in the cyanobacterium Synechocystis sp. PCC 6803, an oxygenic photosynthetic prokaryote, using a resin-assisted thiol enrichment approach. Our proteomic approach integrates resin-assisted enrichment with isobaric tandem mas ...[more]

PMID: 25118246

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Project description:The excessive light illumination of mammalian retina is known to induce oxidative stress and photoreceptor cell death linked to progression of age-related macular degeneration. The photochemical damage of photoreceptors is suggested to occur via two apoptotic pathways that involve either excessive rhodopsin activation or constitutive phototransduction, depending on the light intensity. Both pathways are dramatically activated in the absence of rhodopsin desensitization by GRK1. Previously, we have shown that moderate illumination (halogen lamp, 1,500 lx, 1-5 h) of mammalian eyes provokes disulfide dimerization of recoverin, a calcium-dependent regulator of GRK1. Here, we demonstrate under in vivo conditions that both moderate long-term (metal halide lamp, 2,500 lx, 14 h, rat model) and intense short-term (halogen lamp, 30,000 lx for 3 h, rabbit model) illumination of the mammalian retina are accompanied by accumulation of disulfide dimer of recoverin. Furthermore, in the second case we reveal alternatively oxidized derivatives of the protein, apparently including its monomer with sulfinic group. Histological data indicate that thiol oxidation of recoverin precedes apoptosis of photoreceptors. Both disulfide dimer and oxidized monomer (or oxidation mimicking C39D mutant) of recoverin exhibit lowered α-helical content and thermal stability of their apo-forms, as well as increased Ca2+ affinity. Meanwhile, the oxidized monomer and C39D mutant of recoverin demonstrate impaired ability to bind photoreceptor membranes and regulate GRK1, whereas disulfide dimer exhibits notably improved membrane binding and GRK1 inhibition in absence of Ca2+. The latter effect is expected to slow down rhodopsin desensitization in the light, thereby favoring support of the light-induced oxidative stress, ultimately leading to photoreceptor apoptosis. Overall, the intensity and duration of illumination of the retina affect thiol oxidation of recoverin likely contributing to propagation of the oxidative stress and photoreceptor damage.

Dataset Information

Proteome-wide light/dark modulation of thiol oxidation in cyanobacteria revealed by quantitative site-specific redox proteomics.

Publications

Proteome-wide light/dark modulation of thiol oxidation in cyanobacteria revealed by quantitative site-specific redox proteomics.

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