Faculty Mentor(s)
Dr. Leah Chase, Biology and Chemisty
Document Type
Poster
Event Date
4-14-2023
Abstract
System xc- is a membrane transport system that plays a critical role in mitigating oxidative stress. As such, its regulation is critical for proper brain functioning. Recent work in our lab has shown that System xc- activity increases immediately during an oxidative insult by undergoing a change in localization to the plasma membrane, but we have yet to identify the specific mechanism for the redistribution of the transporter. Previous studies have demonstrated that post-translational modifications of proteins can lead to differential protein distribution within cells. Therefore, in this study, we sought to determine if post-translational modification (PTM) of the transporter regulates its trafficking. First, we identified four conserved lysines (K37, K422, K472, K473) which exhibit decreased activity upon mutation to arginine, suggesting that PTM of these sites increases activity. We used biotinylation to examine the effects of the mutations on transporter localization in the cells, and we evaluated the effects these mutations had on the tendency for these transporters to undergo PTM. As such, this approach allowed us to directly relate changes in PTM status at these select lysines with changes in transporter localization. Our biotinylation results demonstrate K472R and K473R do not appear to shift to the membrane following peroxide treatment, and follow up immunocytochemistry analysis suggests they may be stuck in the endoplasmic reticulum. We also observed that K473R exhibits a 5-10 kD decrease in the molecular weight, indicating that K473 is modified under basal conditions. However, neither mutation impacted the ubiquitination status of xCT. Therefore, we are currently working to identify the PTM that occurs at these lysines, and our preliminary data suggests that K473R may exhibit changes in its glycosylation relative to wild-type and the other mutants. Collectively, these data suggest that PTM of K472 and K473 support xCT delivery to the membrane under basal conditions.
Recommended Citation
Repository citation: Koppin, Anna; Buck, Claire; and Gibson, Amanda, "Characterization of Select Lysine Mutations of the Cystine/Glutamate Transporter, System xc-" (2023). 22nd Annual Celebration of Undergraduate Research and Creative Activity (2023). Paper 20.
https://digitalcommons.hope.edu/curca_22/20
April 14, 2023. Copyright © 2023 Hope College, Holland, Michigan.
Comments
This research was supported by the A. Paul and Carol Schaap Undergraduate Research Fund and the De Vries Summer Research Fund.