Abberant α-Synuclein Confers Toxicity to Neurons in Part through Inhibition of Chaperone-Mediated Autophagy

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.

Conophylline Protects Cells in Cellular Models of Neurodegenerative Diseases by Inducing Mammalian Target of Rapamycin (mTOR)-independent Autophagy

“…In a cellular model of PD, transfection with Atg7 siRNA for 96 h did not induce cell death, even though autophagy acts cytoprotectively. We assumed that although activation of autophagy is cytoprotective when cells were damaged by aggresomes, the defect of the basal level of autophagy for a short time has no effect on the cell viability in cultured cell consistent with a previous report (32). Moreover, time course analysis showed that 12 h of treatment with CNP increased LC3-II level, and this effect did not continue until 48 h (data not shown) in HeLa cells.…”

The Solution Structure and Dynamics of Full-length Human Cerebral Dopamine Neurotrophic Factor and Its Neuroprotective Role against α-Synuclein Oligomers

“…The cellular clearance of ␣-synuclein is performed by the ubiquitin-proteasome system and chaperone-mediated autophagy (64), and interventions that stimulate chaperone-mediated autophagy can prevent ␣-synucleinopathy (65)(66)(67). Furthermore, aberrant ␣-synuclein inhibits the chaperone-mediated autophagy, which adds to the multitude of proposed mechanisms for its toxicity (68).…”

Intrabodies as Neuroprotective Therapeutics

“…Recently, Gaucher mutations were shown to lead to increased accumulation of α-Syn [69]. An α-Syn pathogenic cascade may include excess monomeric α-Syn interference at the synapse [70], and accumulation of stress-inducing phosphorylated, truncated, nitrated, or dopamine-conjugated α-Syn [71,72]. Excess α-Syn may also lead to misfolded α-Syn with increased potential to aggregate into intra-and extracellular toxic conformations.…”