Stress-induced phospho-ubiquitin formation causes parkin degradation

Lyudmila Kovalchuke; Eugene V. Mosharov; Oren A. Levy; Lloyd A. Greene

In this study, we investigated the mechanism by which stressors, L-DOPA in particular, decrease cellular levels of parkin protein. We found that L-DOPA causes parkin loss via two distinct pathways: an oxidative stress-dependent pathway driven by L-DOPA autoxidation and an oxidative stress-independent pathway, each of which is responsible for about half of parkin loss (Fig. 11 ). We conclude that oxidative stress plays a role in only one of these pathways because parkin mutants deficient in binding phospho-Ub were only partly protected from L-DOPA-induced loss, whereas they were fully protected from loss induced by the oxidative stressor hydrogen peroxide. Furthermore, exposure to the antioxidant glutathione, which is critical for neuronal defense against ROS [105] , only attenuated L-DOPA-induced parkin loss by half. Although glutathione lacks membrane permeability [106] , [107] and does not react efficiently with all kinds of ROS [108] , glutathione treatment has been previously shown to be effective at completely preventing the autoxidation of both L-DOPA and the dopamine analog 6hydroxydopamine (6-OHDA) to their respective quinones [109] , [110] . Additionally, glutathione treatment has been found to completely prevent cell death induced by L-DOPA [109] , 6-OHDA [110] , dopamine [111] [112] [113] , hydrogen peroxide [110] , and the oxidant nitric oxide [114] . In support of glutathione's capacity to prevent L-DOPA-induced oxidative stress in our hands, we observed an almost complete abrogation of L-DOPA-induced phospho-poly-Ub formation upon glutathione treatment. Nevertheless, it's possible that not all aspects of L-DOPA-induced oxidative stress were fully neutralized by glutathione in our model, which would have led us to underestimate the contribution of oxidative stress to parkin loss. The copyright holder for this preprint (which was not peer-reviewed) is the author/funder. It . https://doi.org/10.1101/484857 doi: bioRxiv preprint