Oxidative Modifications of Parkin Underlie its Selective Neuroprotection in Adult Human Brain

Abstract

The mechanisms by which Parkinson disease-linked parkin confers neuroprotection of human dopamine cells remain elusive. We hypothesized that its cysteines mediate multiple anti-oxidant effects in the midbrain. By studying >60 control specimens, we found that in adult human brainbut not in skeletal muscle-parkin is mostly aggregated and insoluble due to oxidative modifications, such as at C253. In vitro, parkin's oxidation directly reduces hydrogen peroxide (H2O2) to water. In parkin-deficient human brain, H2O2 concentrations are elevated. In dopamine toxicity studies, wild-type parkin -but not disease-associated mutants-prevents neural death by lowering H2O2 and sequestering radicals within insoluble aggregates. Parkin conjugates dopamine metabolites at the human-specific residue C95 and augments melanin formation in vitro. Using epitope-mapped antibodies, we found that in adult Substantia nigra neurons parkin localizes to neuromelanin within LAMP-3/CD63-positive lysosomes. We conclude that parkin's own oxidation, previously considered a loss-of-function event, underlies three neuroprotective effects in adult midbrain: its cysteines participate in H2O2 reduction, dopamine radical conjugation and the formation of neuromelanin.Parkinson disease (PD) remains an incurable disorder of the human brain. Bi-allelic mutations in the PRKN gene, which encodes parkin, lead to young-onset, autosomal-recessive PD (ARPD) 1 . Clinicopathological studies of parkin-deficient brains have demonstrated restricted cell loss in the S. nigra and L. coeruleus, two nuclei synthesizing dopamine (DA). The field has not yet explained the selective neurodegeneration of PRKN-linked ARPD vs. other forms of PD.Parkin, a principally cytosolic protein, has been associated with diverse cellular functions, foremost related to ubiquitin ligase activity and mitochondrial integrity 2 . However, none of these roles has explained its neuroprotective selectivity, and animal models of genomic prkn deletion have failed to reproduce DA cell loss and parkinsonism. This observation could be due to compensatory mechanisms, a shorter life span, or the uniqueness of human DA metabolism. The latter is exemplified by the generation of neuromelanin in dopaminergic cells beginning in late childhood 3,4 . Nevertheless, select prkn-null models have revealed changes in high energy-producing cells of flies 5 and murine brain indicative of augmented oxidative stress [6][7][8] . We postulated that wild-type parkin promotes redox homeostasis in vivo.Redox equilibrium frequently involves cysteine-based chemistry; there, thiols are subject to oxidative modifications by reactive oxygen-, reactive nitrogen-and reactive electrophilic species (ROS, RNS, RES) 9 , some of which are reversible. Proteins irreversibly conjugated by RES, including by select DA radicals, are degraded, secreted or sequestered within inclusions. It is thought that this process underlies neuromelanin formation 10 .Human parkin contains 35 cysteines 1 . Several reports previously demonstrated its sensitivit...

Autophagy and Redox Homeostasis in Parkinson’s: A Crucial Balancing Act

“…In addition, Meng et al described that Parkin can be sulfonated in an in vitro Parkinson's model, leading to protein aggregation, and possibly contributing to the formation of Lewy's bodies in Parkinson's [ 298 ]. However, in a recent article (at preprint stage at the point of writing this review), Tokarew et al highlighted the importance of Parkin's own oxidation in neuroprotection [ 299 ]. Previously, Vandiver et al showed that Parkin can also undergo sulfhydration, enhancing its catalytic activity and its protective function [ 300 ].…”