September 2, 2020
Journal Article

Stochiometric Quantification of the Thiol Redox Proteome of Macrophages Reveals Subcellular Compartmentalization and Susceptibility to Oxidative Perturbations

Jicheng Duan
Xiaolu Li
Ming Xian
Weijun Qian

Abstract

Posttranslational modifications of protein cysteine thiols play a significant role in redox regulation and the pathogenesis of human diseases. However, the cellular redox landscape in terms of quantitative, site-specific occupancies of thiol modifications at the proteome level, especially under physiological conditions, is still largely uncharacterized. Herein, we report the site occupancies of both S-glutathionylation (SSG) and total reversible thiol oxidation (total oxidation) in RAW 264.7 macrophage cells under basal conditions. The occupancies of thiol modifications for ~4,000 cysteine sites were quantified, which revealed a mean site occupancy of 4.0% for SSG and 11.9% for total oxidation, respectively. Proteome-wide site occupancy analysis revealed a strong subcellular compartmentalization in thiol redox status, where the average occupancies of SSG and total oxidation in distinct compartments correlate well with the redox potentials of respective organelles. The lowest site occupancies were observed in more reducing compartments such as mitochondria and nucleus, while the highest site occupancies were found in more oxidizing organelles such as endoplasmic reticulum (ER) and lysosome. Correlations between site occupancies and structural features such as pKa, relative residue surface accessibility, and hydrophobicity were also observed. Furthermore, under oxidative stress induced by exposure to engineered metal oxide nanoparticles, we observed that mitochondria and ER are less susceptible to low oxidative stress perturbations than nucleus and cytoplasm, presumably due to the differences in basal redox states and antioxidant capacity in different compartments.

Revised: September 2, 2020 | Published: September 1, 2020

Citation

Duan J., T. Zhang, M.J. Gaffrey, K.K. Weitz, R.J. Moore, X. Li, and M. Xian, et al. 2020. "Stochiometric Quantification of the Thiol Redox Proteome of Macrophages Reveals Subcellular Compartmentalization and Susceptibility to Oxidative Perturbations." Redox Biology 36. PNNL-SA-153520. doi:10.1016/j.redox.2020.101649