PP69 - High-glucose-induced impairment of the proteasomal system in pancreatic Min6 cells

Abstract

Protein degradation by the 26S proteasome is a fundamental process involved in a broad range of cellular activities, yet how proteasome activity is regulated remains poorly understood. In particular, the mechanisms of redox-dependent regulation of the ubiquitin-proteasomal system (UPS) are not clear. High glucose has been proposed to promote oxidative stress in different cellular models. Min6 b-cells grown under high glucose conditions showed a decreased 20S (chemotrypsin) activity compared to control cells, while 20S core subunits were increased as determined by western blot. Peroxiredoxin 5 levels were increased under these conditions, suggesting a response to chronic oxidative stress conditions. On the other hand, short-term exposure of the cells to high glucose concentrations caused a decrease in 26S activity and an increase in 20S activity. Similar results were observed when cells were incubated with the mitochondrial uncoupler antimycin A or hydrogen peroxide. Glucose-induced increase of 20S activity was time-dependent and this effect was abolished when the 20S activity was measured in the presence of the thiol reducing agent dithiotreitol. Dimerization of mitochondrial and cytosolic peroxiredoxins evidenced an increase in the steady-state concentration of peroxides under these conditions. Moreover, non-denaturing PAGE followed by in-gel proteasome activity evidenced the formation of aggregates and changes in proteasome structure which were partially reversed by thiol reduction. Our data suggest that high glucose promotes oxidative stress in Min6 b-cells with a concomitant UPS impairment. Further studies will focus on the thiol modifications of the UPS induced under this conditions and its reversibility.