α-synuclein buildup is alleviated via ESCRT-dependent endosomal degradation brought about by p38MAPK inhibition in cells expressing p25α.

ABSTRACT

α-synucleinopathy is driven by an imbalance of synthesis and degradation of α-synuclein (αSyn), causing a build up of αSyn aggregates and post-translationally modified species, which not only interfere with normal cellular metabolism but also by their secretion propagates the disease. Therefore, a better understanding of αSyn degradation pathways is needed to address α-synucleinopathy. Here, we used the nerve growth factor-differentiated catecholaminergic PC12 neuronal cell line, which was conferred α-synucleinopathy by inducible expression of αSyn and tubulin polymerization-promoting protein p25α. p25α aggregates αSyn, and imposes a partial autophagosome-lysosome block to mimic aspects of lysosomal deficiency common in neurodegenerative disease. Under basal conditions, αSyn was degraded by multiple pathways but most prominently by macroautophagy and Nedd4/Ndfip1-mediated degradation. We found that expression of p25α induced strong p38MAPK activity. Remarkably, when opposed by inhibitor SB203580 or p38MAPK shRNA knockdown, endolysosomal localization and degradation of αSyn increased, and αSyn secretion and cytotoxicity decreased. This effect was specifically dependent on Hsc70 and the endosomal sorting complex required for transport machinery, but different from classical microautophagy, as the αSyn Hsc70 binding motif was unnecessary. Furthermore, in a primary neuronal (h)-αSyn seeding model, p38MAPK inhibition decreased pathological accumulation of phosphorylated serine-129-αSyn and cytotoxicity. In conclusion, p38MAPK inhibition shifts αSyn degradation from various forms of autophagy to an endosomal sorting complex required for transport-dependent uptake mechanism, resulting in increased αSyn turnover and cell viability in p25α-expressing cells. More generally, our results suggest that under conditions of autophagolysosomal malfunction, the uninterrupted endosomal pathway offers a possibility to achieve disease-associated protein degradation.