LRRK2 and oxidative stress in Parkinson’s disease

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Mutations in LRRK2 are the most common cause of autosomal dominant Parkinson's disease (PD) and it appears that all such pathogenic mutations are associated with aberrantly enhanced LRRK2 kinase activity. Independent of mutations, however, there is also evidence that increased LRRK2 kinase activity contributes to the pathogenesis of idiopathic PD (iPD). The current project is designed to elucidate how wildtype (non-mutated) LRRK2 kinase activity is stimulated, where in the cell this occurs, and what the downstream consequences are. In keeping with the well-known association of iPD with oxidative stress and mitochondrial dysfunction, we focus on these processes in relation to LRRK2. Specifically, this project examines (i) the oxidative activation of LRRK2, (ii) the translocation of LRRK2 to mitochondria under conditions of mitochondrial stress, and (iii) LRRK2 kinase activity-dependent oxidative stress. The overarching premise of this proposal is that oxidative stress and LRRK2 kinase activity are intimately and bidirectionally intertwined in PD pathogenesis. The project has 3 broad Specific Aims to address these issues: Aim 1: To elucidate the oxidative activation of LRRK2 kinase, we will (a) assess/compare WT LRRK2 activation by physiological stressors (H2O2 and DA) to the PD-associated toxicants: rotenone, paraquat (PQ) and trichloroethylene (TCE); (b) Determine whether monensin & chloroquine activate LRRK2 via oxidative mechanisms; (c) Examine the role of cysteine residues C2024/5 in oxidative LRRK2 activation. Aim 2: To examine translocation of LRRRK2 to mitochondria, we will (a) Assess mitochondrial localization of LRRK2 in response to various mitochondrial stressors; (b) See if translocation requires (i) mitochondrial ROS, (ii) cytosolic ROS, (iii) LRRK2 activity or (iv) Cys2024/5; (c) Determine whether mitochondrial translocation of LRRK2 occurs in vivo in rat models of PD; (d) Evaluate whether LRRK2 association with mitochondria is aberrantly enhanced in human iPD brain tissue. Aim 3: To elucidate the role of LRRK2 kinase in oxidative stress and its downstream consequences, we will (a) Use genetic and pharmacological approaches to see if pathogenic LRRK2 mutations cause oxidative stress and whether blocking LRRK2 kinase activity prevents oxidative stress; (b) Examine the potential role of Cys2024/5 in driving LRRK2 kinase activity-dependent oxidative stress; (c) Determine if rotenone-induced mitochondrial ROS production is reduced by LRRK2 inhibition/knockout; (d) Determine if rotenone-induced cytosolic NOX2 activation is reduced by LRRK2 inhibition/knockout; (e) Assess ROS production in patient-derived healthy control, iPD and G2019S mutant lymphoblastoid cells. Together, these experiments will begin to elucidate the intimate and bidirectional relationship between oxidative stress and LRRK2 in PD pathogenesis. By doing so, new therapeutic strategies are likely to emerge.