Citation:

Masood, S., E. Pennington, P. Bromberg, A. Gold, Z. Zhang, R. Rice, AND J. Samet. Detecting Protein Sulfenylation in Human Airway Epithelial Cells (HAEC) Exposed to Environmental Peroxides. Society of Redox Biology and Medicine 29th Annual Conference, Orlando, FL, November 16 - 19, 2022.

Impact/Purpose:

The goal of this study is to investigate the mechanisms toxicity, specifically the mechanism of oxidative stress imposed by environmetal hydroperoxides, specifically ISOPOOH. We have previousely demostrated that ISOPOOH is capable of induced glutathione oxidation, however recently I have been able to demostrate that ISOPOOH can induce GAPDH sulfenylation. This has major biological revelance as GAPDH is involved in cellular bioenergetics. Thus if ISOPOOH can lead to GAPDH sulfenylation it will potentially lead to major alterations in cellular bioenergetics. The goal for this presenation is to interact with experts regarding cellular oxidative stress and cellular bioenergetic mechanisms to have gain better insight in how ISOPOOH can lead to these allerations in cellular oxidative respones. 

Description:

Exposure to airborne fine particulate matter (PM2.5) is a leading cause of morbidity and mortality worldwide. A major progenitor of PM2.5 is derived from the atmospheric oxidation of isoprene by hydroxy radical to form isoprene hydroxy hydroperoxide (ISOPOOH). ISOPOOH formation is well understood however relatively little is known about the adverse health effects of ambient ISOPOOH exposure. We previously showed that ISOPOOH exposure induces oxidative stress by inducing glutathione oxidation independently of the generation of intracellular H2O2 in HAEC. Knock down of GPx4 expression ablates the ISOPOOH induced glutathione oxidation, implicating GPx4 involvement specifically. It also been demonstrated that ISOPOOH exposure of cellular and acellular membranes induces lipid peroxidation which can be modulated by supplementation with poly-unsaturated fatty acids. Our current hypothesis is that in addition to ISOPOOH induced glutathione oxidation, ISOPOOH exposure of HAEC leads to protein sulfenylation specifically GAPDH sulfenylation which, in turn, leads to shunting of glucose to the pentose phosphate pathway. Our experimental approach primarily uses dimedone-based reagents and copper catalyzed azo-alkynyl cycloaddition reactions to tag intracellular protein sulfenic acids. Low micromolar exposure of H2O2 and ISOPOOH induces dose-dependent protein sulfenylation of GAPDH. Exposure of GAPDH to H2O2 or ISOPOOH in the presence of glutathione leads to glutathionylation of GAPDH. Another low molecular weight hydroperoxide, tert-butyl hydroperoxide, also induces protein sulfenylation and glutathionylation of GAPDH at low micromolar concentrations. Fluorescent images of intracellular protein sulfenylation were detected in HAEC exposed to ISOPOOH. These findings demonstrate that ISOPOOH is potent environmental hydroperoxides capable of inducing oxidative stress through mechanisms involving glutathione oxidation and sulfenylation regulatory proteins in HAEC. 

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