Probing Oxidant Effects on Superoxide Dismutase 1 Oligomeric States in Live Cells Using Single-Molecule Fluorescence Anisotropy
Speaker: Tai-Yen Chen, University of Houston
Host: Yanxin Liu
Abstract: The protein Cu/Zn superoxide dismutase (SOD1) is known to function as a dimer, but its concentration in cells (∼50 μM) and the dimerization constant (Kd of 500 μM) results suggest that it exists in a monomer-dimer equilibrium. It is unclear how the oligomeric state of SOD1 changes when cells are initially exposed to high levels of extracellular oxidative stress. To address this problem, we introduced the single-molecule fluorescence anisotropy (smFA) assay to explore SOD1 oligomeric states in live COS7 cells. smFA specifically probes the fluorescence polarization changes caused by molecular rotations where the fast-rotating molecules (either due to smaller hydrodynamic volume or less viscous environments) deteriorate the emission polarization and thus lower the anisotropy. After validating that smFA is effective in distinguishing monomeric and dimeric fluorescence proteins, we overexpressed SOD1 in live COS7 cells and investigated how its oligomeric state changes under basal, 2 h, and 24 h 100 μM H2O2 treatments. We found that treating cells with H2O2 promotes SOD1 dimerization and decreases cellular viscosity in 2 h. Interestingly, prolonged H2O2 treatments show similar results as the basal conditions, indicating that cells return to a steady state similar to the basal state after 24 h, despite the presence of H2O2. Our results demonstrate that SOD1 changes its oligomeric state equilibrium in response to extracellular oxidative stresses. smFA will open new opportunities to explore the relationship between the SOD1 oligomer state and its H2O2-based signaling and transcription regulation roles.
Bio: Tai-Yen earned his B.S. (2002) and M.S. (2004) in Chemistry from National Tsing Hua University and a Ph.D. (2010) from Texas A&M University before pursuing a postdoctoral associate (2011-2016) at Cornell University. Tai-Yen started his independent career as an assistant professor in the Department of Chemistry at the University of Houston in 2016. His research focuses on metal homeostasis in healthy and diseased neurons using single-molecule techniques. In particular, his group is interested in the dynamics and mechanisms of protein machinery involved in Cu trafficking within/between neurons. His research goal is to understand how metals affect neuronal signaling and cause neurodegenerative diseases. His work has been recognized with the Robert A. Welch Foundation Research Grant (2018), the NIH Maximizing Investigators' Research Award for Early Stage Investigators (2019), and the NSF CAREER Award (2023). Besides research, Tai-Yen enjoys and enthusiastically participates in multiple educational activities. In recognition of his classroom teaching and laboratory mentorship, Tai-Yen received the College's highly competitive John C. Butler Excellence in Teaching Award in 2021.
Seminars start at 4:00 pm, and refreshments will be served at 3:45 pm. All seminars are held in the Conference Room (1116) of the Institute for Physical Science and Technology (IPST) Building unless otherwise noted.