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Extracellular plaques of amyloid-? (A?) are the main histopathological signatures of Alzheimer's disease (AD). In vitro and in vivo analysis of amyloid deposits in AD revealed various N- and C-terminal variants, in addition biochemical studies showed that A? peptides isolated from AD brains were post-translationally modified. N-terminal deletions and pyroglutamylation enhance aggregation of A? into neurotoxic species and such peptides may initiate or nucleate the pathological deposition of A? into plaques. Among these, A?pE3-42 strongly affects cultured neuron and astrocyte survival. In the last years, a new molecular actor has appeared in AD pathophysiology: the cellular prion protein (PrPC). The role of this protein is still not completely disclosed, but its involvement in the complex pathway connecting A? to Fyn/Tau neuronal toxicity is now recognized. In particular, the trigger of the molecular cascade leading to synapses impairment and cellular death seems to be the direct interaction of A? oligomers (A?o) with PrPC. In this work we exploit the interactions between A? peptides and N-terminal truncated PrPC. In particular we study the effect of PrPc (90-231) on aggregation of A?1-42, A?pE3-42 and of their mixtures. To reach this goal we investigated the different interactions by NMR and we characterized the intermediate species by turbidimetry, circular dichroism, transmission electron microscopy, ThT and ANS fluorescence. To determine if PrPC is a mediator of A?-induced neurotoxicity, we studied the physiological response of cells to A?o, and the dysregulation of calcium homeostasis, both on glioblastoma cells where PrPC expression was inhibited or not

According to the “gain-of-toxicity mechanism”, neuronal loss during cerebral proteinopathies is caused by accumulation of aggregation-prone conformers of misfolded cellular proteins, although it is still debated which aggregation state actually corresponds to the neurotoxic entity. Autophagy, originally described as a variant of programmed cell death, is now emerging as a crucial mechanism for cell survival in response to a variety of cell stressors, including nutrient deprivation, damage of cytoplasmic organelles, or accumulation of misfolded proteins. Impairment of autophagic flux in neurons often associates with neurodegeneration during cerebral amyloidosis, suggesting a role in clearing neurons from aggregation-prone misfolded proteins. Thus, autophagy may represent a target for innovative therapies. In this work, we show that alterations of autophagy progression occur in neurons following in vitro exposure to the amyloidogenic and neurotoxic prion protein-derived peptide PrP90-231. We report that the increase of autophagic flux represents a strategy adopted by neurons to survive the intracellular accumulation of misfolded PrP90-231. In particular, PrP90-231 internalization in A1 murine mesencephalic neurons occurs in acidic structures, showing electron microscopy hallmarks of autophagosomes and autophagolysosomes. However, these structures do not undergo resolution and accumulate in cytosol, suggesting that, in the presence of PrP90-231, autophagy is activated but its progression is impaired; the inability to clear PrP90-231 via autophagy induces cytotoxicity, causing impairment of lysosomal integrity and cytosolic diffusion of hydrolytic enzymes. Conversely, the induction of autophagy by pharmacological blockade of mTOR kinase or trophic factor deprivation restored autophagy resolution, reducing intracellular PrP90-231 accumulation and neuronal death. Taken together, these data indicate that PrP90-231 internalization induces an autophagic defensive response in A1 neurons, although incomplete and insufficient to grant survival; the pharmacological enhancement of this process exerts neuroprotection favoring the clearing of the internalized peptide and could represents a promising neuroprotective tool for neurodegenerative proteinopathies.