Abstract
BackgroundElevated SNCA gene expression and intracellular accumulation of the encoded α-synuclein (aSyn) protein are associated with the development of Parkinson disease (PD). To date, few enzymes have been examined for their ability to degrade aSyn. Here, we explore the effects of CTSD gene expression, which encodes the lysosomal protease cathepsin D (CathD), on aSyn processing.ResultsOver-expression of human CTSD cDNA in dopaminergic MES23.5 cell cultures induced the marked proteolysis of exogenously expressed aSyn proteins in a dose-dependent manner. Unexpectedly, brain extractions, Western blotting and ELISA quantification revealed evidence for reduced levels of soluble endogenous aSyn in ctsd knock-out mice. However, these CathD-deficient mice also contained elevated levels of insoluble, oligomeric aSyn species, as detected by formic acid extraction. In accordance, immunohistochemical studies of ctsd-mutant brain from mice, sheep and humans revealed selective synucleinopathy-like changes that varied slightly among the three species. These changes included intracellular aSyn accumulation and formation of ubiquitin-positive inclusions. Furthermore, using an established Drosophila model of human synucleinopathy, we observed markedly enhanced retinal toxicity in ctsd-null flies.ConclusionWe conclude from these complementary investigations that: one, CathD can effectively degrade excess aSyn in dopaminergic cells; two, ctsd gene mutations result in a lysosomal storage disorder that includes microscopic and biochemical evidence of aSyn misprocessing; and three, CathD deficiency facilitates aSyn toxicity. We therefore postulate that CathD promotes 'synucleinase' activity, and that enhancing its function may lower aSyn concentrations in vivo.
Highlights
Elevated SNCA gene expression and intracellular accumulation of the encoded α-synuclein protein are associated with the development of Parkinson disease (PD)
A key role had been postulated for the ubiquitin proteasome pathway (UPP) in the degradation of accumulation of the encoded α-synuclein (aSyn), because mutations in two UPP-related genes, Parkin and UchL-1 have been shown to influence PD risk [15,16,17,18] and because molecular, cellular and animal studies linked these genes to UPP-dependent processing of aSyn [19,20,21]
Ectopic Cathepsin D effectively degrades wild-type and mutant α-synuclein in dopaminergic cells To investigate whether cathepsin D (CathD) has any effect on aSyn turnover, we utilized a rodent cell line of mesencephalic origin, MES23.5, which constitutively synthesizes dopamine
Summary
Elevated SNCA gene expression and intracellular accumulation of the encoded α-synuclein (aSyn) protein are associated with the development of Parkinson disease (PD). Accumulation of aggregated, insoluble aSyn is a hallmark of many other neurodegenerative diseases, including sporadic PD, dementia with Lewy bodies (DLB), multiple system atrophy (MSA), the Lewy body variant of Alzheimer's disease, and PANK2-linked neurodegeneration. These disorders are referred to as synucleinopathies [4,5,6,7]. Proteins are sequestered within lysosomes by one of three known methods, i.e., macroautophagy, microautophagy, and chaperone-mediated autophagy (CMA) (reviewed in [24]) Of these, it appears that aSyn can be a substrate for both macroautophagy and CMA [25,26,27,28]
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