Mechanisms of Prion Spread and Neuronal Toxicity

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Synaptic dysfunction and neuritic dystrophy are prominent pathologic features of the prion- and Alzheimer’s disease-affected brain. Ubiquitinated protein inclusions are also commonly observed, providing strong evidence of impaired proteostatic pathways. Ubiquitination of cell membrane proteins and clearance through the ESCRT pathway (endosomal sorting complex required for transport) is critical to maintaining synaptic homeostasis. Here we will deeply investigate the ESCRT pathway contributions to disrupted synaptic homeostasis in prion disease. In prion-infected mice, we have found markedly reduced ESCRT-0 (an Hrs and STAM1 protein complex) and an enrichment of ubiquitinated proteins in synaptosomes. Strikingly, depleting neuronal Hrs in prion-infected mice shortened survival time and accelerated the degeneration of synapses, biochemically and structurally. Additionally, in a longitudinal study of the prion-infected hippocampus, we found an upregulation in the synaptic activity response gene, Arc/Arg3.1, and a chronic elevation in phosphorylated CaMKII and phosphorylated AMPA receptors, suggestive of enhanced and altered synapse function beginning in early disease. Our long-term goal is to decipher how prion and amyloid-β oligomers disrupt signaling pathways linked to the cellular prion protein, inducing proteostatic dysfunction and synaptic degeneration. Using electrophysiology, correlative light-electron microscopy, and proteomics on uninfected and prion-infected cultured neurons, we will first determine how Hrs expression impacts synapses, assessing activity, pre-and post-synaptic proteins, structure, and signaling. We will then test how distinct prion conformers impact the ESCRT pathway and neuronal signaling at glutamatergic synapses. Finally, we will investigate the contribution of glutamate receptor activity to prion spread and neurodegeneration. We will directly test how the findings in these genetically manipulated models compare to human prion-affected brain. These studies are the first to test how neuronal activity impacts prion dissemination, synaptic degeneration, and disease progression, and outcomes are expected to provide key insights into the deregulated synaptic signaling that drives neuron loss, thus revealing new therapeutic targets.