Abstract
The degeneration of dopaminergic neurons during Parkinson’s disease (PD) is intimately linked to malfunction of α-synuclein (αSyn), the main component of the proteinaceous intracellular inclusions characteristic for this pathology. The cytotoxicity of αSyn has been attributed to disturbances in several biological processes conserved from yeast to humans, including Ca2+ homeostasis, general lysosomal function and autophagy. However, the precise sequence of events that eventually results in cell death remains unclear. Here, we establish a connection between the major lysosomal protease cathepsin D (CatD) and the Ca2+/calmodulin-dependent phosphatase calcineurin. In a yeast model for PD, high levels of human αSyn triggered cytosolic acidification and reduced vacuolar hydrolytic capacity, finally leading to cell death. This could be counteracted by overexpression of yeast CatD (Pep4), which re-installed pH homeostasis and vacuolar proteolytic function, decreased αSyn oligomers and aggregates, and provided cytoprotection. Interestingly, these beneficial effects of Pep4 were independent of autophagy. Instead, they required functional calcineurin signaling, since deletion of calcineurin strongly reduced both the proteolytic activity of endogenous Pep4 and the cytoprotective capacity of overexpressed Pep4. Calcineurin contributed to proper endosomal targeting of Pep4 to the vacuole and the recycling of the Pep4 sorting receptor Pep1 from prevacuolar compartments back to the trans-Golgi network. Altogether, we demonstrate that stimulation of this novel calcineurin-Pep4 axis reduces αSyn cytotoxicity.
Highlights
Parkinson’s disease (PD) is an age-associated neurodegenerative disorder characterized by the progressive loss of dopaminergic neurons in the substantia nigra pars compacta
The larger fraction of Pep4-GFP was still targeted to the vacuole, visualized via CellTracker Blue CMAC, a dye that is rapidly sequestered into the vacuolar lumen
Expression of αSyn led to the accumulation of Pep4 in prevacuolar compartments in some cells, suggesting that αSyn interferes with the trafficking of this protease from the trans-Golgi network to the vacuole, which might in part account for the observed reduction of Pep4 activity
Summary
Parkinson’s disease (PD) is an age-associated neurodegenerative disorder characterized by the progressive loss of dopaminergic neurons in the substantia nigra pars compacta. Even though CatD seems to act as the major protease involved in the degradation of αSyn and high CatD levels mostly provided neuroprotection (Qiao et al, 2008; Sevlever et al, 2008; Matrone et al, 2016), pharmacological inhibition of this lysosomal aspartyl protease has been demonstrated to decrease αSyn aggregation propensity and toxicity (Takahashi et al, 2007) While both calcineurin and CatD have been connected to neurotoxic events in general and to αSyn-associated cellular demise in particular, an interplay between these two proteins has never been shown.
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