738B Poster - 12. Physiology, metabolism and aging
Friday April 08, 2:00 PM - 4:00 PM

Author:
Sarah Stanhope

Affiliation: Purdue University

Keywords:
a. stress responses; j. epigenetics

Reactive oxygen species (ROS) serve as intracellular signaling molecules; however, in excess ROS molecules are damaging to biomacromolecules such as DNA, lipids, and proteins. The excess accumulation of ROS leads to oxidative stress, disrupting redox homeostasis in the cell and has been linked with aging and neurodegenerative disease. The eye is particularly vulnerable to oxidative stress due to the tissue’s high energy demand and generation of ROS by products. In proteins, the thiol group on cysteine residues is susceptible to oxidation. Cysteine residues have multiple oxidation states that can be classified into two categories—reversible and irreversible. Irreversible oxidation states include sulfinic and sulfonic acids, which may alter or impair the activity of the protein depending on the position of the cysteine residue. Understanding how the proteome is affected by increasing ROS levels may provide insights into proteins that are potential targets for oxidation. Here, we sought to evaluate how increasing amounts of oxidative stress altered the redox proteome landscape in Drosophila. To characterize the redox proteome, we developed an iodoTMT-multiplex isolation, labeling, and enrichment method to identify cysteine availability and potential oxidation in both S2 cells and w¹¹¹⁸ fly eyes. To do this we exposed S2 cells to 20mM H₂O₂ and w¹¹¹⁸ flies to prolonged blue light, followed by redox proteome profiling with iododTMT-sixplex reagents. S2 cell studies revealed 127 significantly oxidized proteins including multiple well-known oxidative stress proteins such as Glutathione S-transferase D1 (GstD1) and Superoxide dismutase [Cu-Zn] (Sod1). GO term analysis elucidated proteins involved in cellular metabolic processes, sulfur compound processes, and cofactor metabolic processes. Further studies in blue light exposed w¹¹¹⁸ eyes revealed 42 significantly oxidized proteins involved in nucleoside metabolic processes, regulation of cytosolic calcium ion concentration, and glycosyl compound metabolic processes. Interestingly, the overlap of the identified proteins from the S2 cell and w¹¹¹⁸ studies revealed oxidation of methionine metabolism enzymes suggesting that oxidative stress may perturb methionine metabolism affecting metabolic processes within the cell. As methionine metabolism is critical for lifespan across various organisms understanding how it can be affected by increasing oxidative stress and its role in the eye is critical.