Prion diseases are fatal, transmissible neurodegenerative disorders associated with conversion of the host-encoded prion protein (PrP(C)) into an abnormal pathogenic isoform (PrP(Sc)). Following exposure to the infectious agent (PrP(Sc)) in acquired disease, infection is propagated in lymphoid tissues prior to neuroinvasion and spread within the central nervous system. The mechanism of prion dissemination is perplexing due to the lack of plausible PrP(Sc)-containing mobile cells that could account for prion spread between infected and uninfected tissues. Evidence exists to demonstrate that the culture media of prion-infected neuronal cells contain PrP(Sc) and infectivity but the nature of the infectivity remains unknown. In this study we have identified PrP(C) and PrP(Sc) in association with endogenously expressing PrP neuronal cell-derived exosomes. The exosomes from our prion-infected neuronal cell line were efficient initiators of prion propagation in uninfected recipient cells and to non-neuronal cells. Moreover, our neuronal cell line was susceptible to infection by non-neuronal cell-derived exosome PrP(Sc). Importantly, these exosomes produced prion disease when inoculated into mice. Exosome-associated PrP is packaged via a novel processing pathway that involves the N-terminal modification of PrP and selection of distinct PrP glycoforms for incorporation into these vesicles. These data extend our understanding of the relationship between PrP and exosomes by showing that exosomes can establish infection in both neighbouring and distant cell types and highlight the potential contribution of differentially processed forms of PrP in disease distribution. These data suggest that exosomes represent a potent pool of prion infectivity and provide a mechanism for studying prion spread and PrP processing in cells endogenously expressing PrP.

The amyloid protein precursor (APP) of Alzheimer's disease is synthesized as an integral transmembrane protein that is released from cells in culture following proteolytic cleavage. The function of released APP is not known, although there is evidence that the protein may bind to components of the extracellular matrix (ECM). In the present study, substratum-bound APP stimulated neurite outgrowth in cultures of chick sympathetic and mouse hippocampal neurons. This effect was dependent upon the presence of substratum-bound heparan sulfate proteoglycans (HSPG). The effect of APP on neurite outgrowth was comparable to that of laminin. A 14 K N-terminal fragment of APP was found to bind heparin and a region close to the N-terminus of APP (residues 96-110) identified as a potential heparin-binding domain based on secondary structure predictions and molecular modeling. Mutagenesis of three basic residues (lysine-99, arginine-100, and arginine-102) resulted in a recombinant protein (APPhep) with decreased heparin-binding capacity. A peptide homologous to the heparin-binding domain was synthesized and found to bind strongly to heparin and to inhibit binding of 125I-labeled APP to heparin (IC50 approximately 10(-7) M). The peptide blocked the effect of APP on neurite outgrowth (IC50 approximately 10(-7) M), whereas two other peptides homologous to other domains in APP had no effect. The results indicate that the binding of APP to HSPG in the ECM may stimulate the effects of APP on neurite outgrowth.

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