Lysosomal Degradation of α-Synuclein in Vivo

Abstract

Pathologic accumulation of ␣-synuclein is a feature of human parkinsonism and other neurodegenerative diseases. This accumulation may be counteracted by mechanisms of protein degradation that have been investigated in vitro but remain to be elucidated in animal models. In this study, lysosomal clearance of ␣-synuclein in vivo was indicated by the detection of ␣-synuclein in the lumen of lysosomes isolated from the mouse midbrain. When neuronal ␣-synuclein expression was enhanced as a result of toxic injury (i.e. treatment of mice with the herbicide paraquat) or transgenic protein overexpression, the intralysosomal content of ␣-synuclein was also significantly increased. This effect was paralleled by a marked elevation of the lysosome-associated membrane protein type 2A (LAMP-2A) and the lysosomal heat shock cognate protein of 70 kDa (hsc70), two essential components of chaperonemediated autophagy (CMA). Immunofluorescence microscopy revealed an increase in punctate (lysosomal) LAMP-2A staining that co-localized with ␣-synuclein within nigral dopaminergic neurons of paraquat-treated and ␣-synuclein-overexpressing animals. The data provide in vivo evidence of lysosomal degradation of ␣-synuclein under normal conditions and, quite importantly, under conditions of enhanced protein burden. In the latter, increased lysosomal clearance of ␣-synuclein was mediated, at least in part, by CMA induction. It is conceivable that these neuronal mechanisms of protein clearance play an important role in neurodegenerative processes characterized by abnormal ␣-synuclein buildup.Several lines of clinical and experimental evidence support a pathogenetic role of ␣-synuclein in Parkinson disease (PD) 2 and other neurodegenerative disorders. The precise mechanisms by which this endogenously expressed protein becomes involved in pathologic processes have yet to be fully elucidated. However, data from both clinical and laboratory studies indicate that enhanced ␣-synuclein expression is itself capable of triggering a parkinsonian syndrome in humans and PD-like pathology in animal models. Indeed, multiplication mutations of the ␣-synuclein gene that result in increased expression of the wild-type protein are causally associated with autosomal dominant parkinsonism (1, 2). From the experimental standpoint, evidence of a gain of toxic function of elevated ␣-synuclein includes the observation of neurodegeneration and neuronal inclusions in the substantia nigra of rats and monkeys that overexpress ␣-synuclein after viral-mediated neuronal transduction (3, 4).An important corollary to the concept that enhanced ␣-synuclein may have deleterious consequences is that intraneuronal mechanisms of protein homeostasis, such as degradation pathways, could well play a key role in maintaining "non-toxic" levels of ␣-synuclein. Although ␣-synuclein clearance is likely to occur through different mechanisms, recent reports have underscored the important contribution of lysosomal pathways of protein degradation and, in particular, chaperone-mediated autopha...

A Guide to Neurotoxic Animal Models of Parkinson's Disease

“…Epidemiological studies have suggested an increased risk for PD after paraquat exposure (Hertzman et al 1990;Liou et al 1997;Kamel et al 2007;Ritz et al 2009;Tanner et al 2011). Additionally, paraquat treatment increases a-synuclein aggregates reminiscent of Lewy bodies in PD Fernagut et al 2007;Mak et al 2010). Paraquat is also reported in one study to reduce noradrenergic neurons in the locus coeruleus (Fernagut et al 2007).…”

Chaperone-mediated autophagy: roles in disease and aging

“…Increased levels of LAMP-2A have been observed in the early stages of PD both in mouse models [54] and in brains of PD patients [52]. However, in advanced stages, reduced levels of LAMP-2A and hsc70 have been detected instead in the dopaminergic neurons of human brain regions [58].…”

“…Dysfunction in CMA has been described in both familial [50,52,54,55] and sporadic [51,52] PD. In the case of familial PD, sequence analysis reveals the presence of CMA-targeting motifs in the majority of PD-related proteins, supporting an important role for CMA in the control of their intracellular levels ( Figure 2A).…”

“…In the case of familial PD, sequence analysis reveals the presence of CMA-targeting motifs in the majority of PD-related proteins, supporting an important role for CMA in the control of their intracellular levels ( Figure 2A). Indeed, the two most commonly mutated proteins in patients with familial PD, α-synuclein and leucine-rich repeat kinase 2 (LRRK2), have both been demonstrated to undergo degradation in lysosomes via CMA using various experimental systems such as isolated lysosomes, primary mouse neuronal cultures, mouse models of PD and even neuronal-differentiated induced pluripotent stem cells and brains from familial [50,52,54,55] and sporadic [51,52] PD patients. In contrast, pathogenic mutant variants of α-synuclein (for example, A30P and A53T mutants) and of LRRK2 (for example, G2019S and R1441C mutants), despite being recognized by cytosolic hsc70 and successfully delivered to the lysosomal membrane, fail to reach the lysosomal lumen to be degraded by CMA [50,52] (Figure 2A).…”

Tubulin Polymerization-promoting Protein (TPPP/p25α) Promotes Unconventional Secretion of α-Synuclein through Exophagy by Impairing Autophagosome-Lysosome Fusion

“…TPPP/p25␣ Impairs the Autophagosomal Degradation Pathway-Aggregated or modified forms of ␣-synuclein are substrates of all major degradative systems in vivo and in vitro, including the ubiquitin proteasome system, chaperone-mediated autophagy, and macroautophagy (8,9,50,51). In this study we have used p25␣ expression as a tool to increase aggregation and autophagosomal uptake of ␣-synuclein.…”