Objective: In the present study, we aim to counteract regulatory and transcriptional modifiers of autophagy to enhance neuroprotective proteostasis and rescue alpha-synuclein (asyn) toxicity in multiple novel cell models of Parkinson disease (PD).

Background: Parkinson disease (PD) is a neurodegenerative movement disorder marked by progressive motor and non-motor symptoms that lead to profound disability. Neurodegeneration in PD relates to toxic aggregation of alpha-synuclein (asyn), and mounting evidence shows that asyn can be degraded through the conserved pathway of autophagy. However, available methods to modulate autophagy fail to confer clinical benefits due to intrinsic resistance of neurons to these methods. This resistance stems in part from enrichment of the autophagy suppressor, MTMR5, in neurons. Our preliminary work has also uncovered additional genetic and regulatory inhibitory factors in genetic/familial forms of PD that exacerbate autophagy impairment.

Method: To study asyn proteostasis in human neurons, we stably integrated Tet-ON rapid differentiation cassettes into the genomes of human induced pluripotent stem cells (iPSCs) to enable efficient (>99%) differentiation into mature neurons (iNeurons) within 14 days. To model PD, we used iPSCs expressing WT asyn, a homozygous mutation in SNCA(c.209G>A, p.A53T), or a heterozygous GBA1 mutation (c.1226A>G, p.N370S). To potentiate the autophagy, we used lentiviral delivery of shRNA or CRISRPa to modify genetic targets of interest, then assessed effects on asyn, autophagy substrates, and cell viability.

Results: In WT and PD iNeurons, knockdown of MTMR5 lead to marginal decrements in levels of asyn and autophagy substrates LC3-II and p62, but greater reductions with Torin1 treatment. Knockdown of transcriptional inhibitors or overexpression of transcriptional activators of the autophagy-lysosome network also reduced asyn, p62, and LC3-II, and increased survival in PD iNeurons. These effects were abrogated by bafilomycin, but not bortezomib.

Conclusion: Reversing negative regulatory mechanisms of autophagy significantly enhanced autophagic proteostasis of asyn and rescued neurodegeneration in multiple genetic contexts of PD. These molecular mechanisms provide discrete targets of high translational potential for developing innovative disease-modifying therapies for PD and related forms of neurodegeneration.

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MDS Abstracts - https://www.mdsabstracts.org/abstract/rescuing-alpha-synuclein-toxicity-through-neuron-specific-enhancement-of-autophagy/