Abstract
BackgroundThe mechanisms through which aberrant α-synuclein (ASYN) leads to neuronal death in Parkinson's disease (PD) are uncertain. In isolated liver lysosomes, mutant ASYNs impair Chaperone Mediated Autophagy (CMA), a targeted lysosomal degradation pathway; however, whether this occurs in a cellular context, and whether it mediates ASYN toxicity, is unknown. We have investigated presently the effects of WT or mutant ASYN on the lysosomal pathways of CMA and macroautophagy in neuronal cells and assessed their impact on ASYN-mediated toxicity.Methods and FindingsNovel inducible SH-SY5Y and PC12 cell lines expressing human WT and A53T ASYN, as well as two mutant forms that lack the CMA-targeting motif were generated. Such forms were also expressed in primary cortical neurons, using adenoviral transduction. In each case, effects on long-lived protein degradation, LC3 II levels (as a macroautophagy index), and cell death and survival were assessed. In both PC12 and SH-SY5Y cycling cells, induction of A53T ASYN evoked a significant decrease in lysosomal degradation, largely due to CMA impairment. In neuronally differentiated SH-SH5Y cells, both WT and A53T ASYN induction resulted in gradual toxicity, which was partly dependent on CMA impairment and compensatory macroautophagy induction. In primary neurons both WT and A53T ASYN were toxic, but only in the case of A53T ASYN did CMA dysfunction and compensatory macroautophagy induction occur and participate in death.ConclusionsExpression of mutant A53T, and, in some cases, WT ASYN in neuronal cells leads to CMA dysfunction, and this in turn leads to compensatory induction of macroautophagy. Inhibition of these lysosomal effects mitigates ASYN toxicity. Therefore, CMA dysfunction mediates aberrant ASYN toxicity, and may be a target for therapeutic intervention in PD and related disorders. Furthermore, macroautophagy induction in the context of ASYN over-expression, in contrast to other settings, appears to be a detrimental response, leading to neuronal death.
Highlights
Introduction aSynuclein (ASYN) is an abundant neuronal protein closely linked to Parkinson’s Disease (PD) pathogenesis [1,2,3]
Expression of mutant A53T, and, in some cases, WT ASYN in neuronal cells leads to Chaperone Mediated Autophagy (CMA) dysfunction, and this in turn leads to compensatory induction of macroautophagy
CMA dysfunction is responsible for the decrease of total lysosomal protein degradation in rat PC12 cell lines inducibly expressing human A53T ASYN
Summary
Synuclein (ASYN) is an abundant neuronal protein closely linked to Parkinson’s Disease (PD) pathogenesis [1,2,3]. Cellular or animal models based on overexpression of ASYN demonstrate neuronal dysfunction and, occasionally, death, as well as inclusion formation and motor phenotypes [14]. These data, in conjunction, have led to the idea that ASYN is linked to PD through a toxic gain of function that is latent in the WT protein, and is manifest when levels of WT ASYN increase, when point mutations occur, or when WT ASYN is post-translationally altered, through oxidation, nitration or other modifications [15]. We have investigated presently the effects of WT or mutant ASYN on the lysosomal pathways of CMA and macroautophagy in neuronal cells and assessed their impact on ASYN-mediated toxicity
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