Stress-induced phospho-ubiquitin formation causes parkin degradation

Lyudmila Kovalchuke,Lloyd A Greene,Oren A Levy,Eugene V Mosharov

doi:10.1038/s41598-019-47952-5

Abstract

Mutations in the E3 ubiquitin ligase parkin are the most common known cause of autosomal recessive Parkinson’s disease (PD), and parkin depletion may play a role in sporadic PD. Here, we sought to elucidate the mechanisms by which stress decreases parkin protein levels using cultured neuronal cells and the PD-relevant stressor, L-DOPA. We find that L-DOPA causes parkin loss through both oxidative stress-independent and oxidative stress-dependent pathways. Characterization of the latter reveals that it requires both the kinase PINK1 and parkin’s interaction with phosphorylated ubiquitin (phospho-Ub) and is mediated by proteasomal degradation. Surprisingly, autoubiquitination and mitophagy do not appear to be required for such loss. In response to stress induced by hydrogen peroxide or CCCP, parkin degradation also requires its association with phospho-Ub, indicating that this mechanism is broadly generalizable. As oxidative stress, metabolic dysfunction and phospho-Ub levels are all elevated in PD, we suggest that these changes may contribute to a loss of parkin expression.

Highlights

Parkinson’s disease (PD) is a debilitating neurodegenerative disorder, affecting roughly 2% of those over the age of 801
Given the robust parkin loss we observed with 200 μM L-DOPA (68.4 ± 5.2% parkin remaining with 200 μM L-DOPA compared to 0 μM L-DOPA, p = 0.01, N = 5), we chose this dose for further experiments
We investigated the mechanism by which stressors, L-DOPA in particular, decrease cellular levels of parkin protein

Summary

Introduction

Parkinson’s disease (PD) is a debilitating neurodegenerative disorder, affecting roughly 2% of those over the age of 801. An important insight into parkin regulation comes from in vitro and in vivo observations that diverse stressors cause a decrease in parkin protein levels[7,8,28,29,30,31] These stressors include mitochondrial complex I inhibitors[8,28,29,30], oxidative agents[7,8,29,30], and a DNA-damaging agent[31]. Mitophagy results in turnover of both mitochondrial proteins and of parkin itself[36,37] It is, unclear whether parkin loss triggered by oxidative stressors utilizes such mechanisms, and, in particular, what the roles of PINK1, phospho-Ub, parkin activity, parkin autoubiquitination, and autophagy are in this process. We have explored the mechanisms of parkin loss promoted by oxidative stress For this purpose, we primarily employed L-DOPA, the precursor to dopamine (DA). L-DOPA is a standard therapy for PD, and the idea has been raised that, as well as providing symptomatic relief in PD, its prolonged use could contribute to neuronal degeneration[56,57]

Methods

Results

Conclusion