Modulation of Glycosaminoglycans Affects PrP^ScMetabolism but Does Not Block PrP^ScUptake

Abstract

Mammalian prions are unconventional infectious agents composed primarily of the misfolded aggregated host prion protein PrP, termed PrP Sc . Prions propagate by the recruitment and conformational conversion of cellular prion protein into abnormal prion aggregates on the cell surface or along the endocytic pathway. Cellular glycosaminoglycans have been implicated as the first attachment sites for prions and cofactors for cellular prion replication. Glycosaminoglycan mimetics and obstruction of glycosaminoglycan sulfation affect prion replication, but the inhibitory effects on different strains and different stages of the cell infection have not been thoroughly addressed. We examined the effects of a glycosaminoglycan mimetic and undersulfation on cellular prion protein metabolism, prion uptake, and the establishment of productive infections in L929 cells by two mouseadapted prion strains. Surprisingly, both treatments reduced endogenous sulfated glycosaminoglycans but had divergent effects on cellular PrP levels. Chemical or genetic manipulation of glycosaminoglycans did not prevent PrP Sc uptake, arguing against their roles as essential prion attachment sites. However, both treatments effectively antagonized de novo prion infection independently of the prion strain and reduced PrP Sc formation in chronically infected cells. Our results demonstrate that sulfated glycosaminoglycans are dispensable for prion internalization but play a pivotal role in persistently maintained PrP Sc formation independent of the prion strain. IMPORTANCERecently, glycosaminoglycans (GAGs) became the focus of neurodegenerative disease research as general attachment sites for cell invasion by pathogenic protein aggregates. GAGs influence amyloid formation in vitro. GAGs are also found in intra-and extracellular amyloid deposits. In light of the essential role GAGs play in proteinopathies, understanding the effects of GAGs on protein aggregation and aggregate dissemination is crucial for therapeutic intervention. Here, we show that GAGs are dispensable for prion uptake but play essential roles in downstream infection processes. GAG mimetics also affect cellular GAG levels and localization and thus might affect prion propagation by depleting intracellular cofactor pools. P rion diseases are progressive, fatal neurodegenerative diseases of humans and other mammals. The formation of proteaseresistant aggregates of the host-encoded prion protein is central to pathogenesis. Growing evidence supports the hypothesis that aggregates of the misfolded host prion protein PrP C , termed PrP Sc , or folding intermediates thereof are the main, if not the sole, constituent of the infectious agent (1, 2). Prion strains from different donor species have been adapted to laboratory rodents, where they manifest themselves with different disease progressions and pathologies (3). Prion strains target different brain areas in vivo (4) and exhibit restricted cell tropism in vitro (for a review, see reference 5). A growing body of evidence argues that str...

Modulation of amyloid assembly by glycosaminoglycans: from mechanism to biological significance

“…By FRET, it was observed that when cells are D r a f t treated with heparinase, a significant decrease of propagation occurs. A similar effect of GAGmediated aggregates propagation was reported for the prion protein (Wolf et al 2015).…”

The Role of Vesicle Trafficking Defects in the Pathogenesis of Prion and Prion-Like Disorders

“…PrP c and PrP Sc interacting receptors, like glycosaminoglycans (GAGs), laminin receptor precursor (LRP/LR), and LRP1, have a prominent role in the conversion process. It has been demonstrated that cultured cells harboring reduced amounts or modified GAGs, such as heparin sulfate, are resistant to primary prion infection [ 60 , 61 ]. Interfering with lipid raft formation by drugs, such as lovastatin, filipin, and amphotericin B, was capable of reducing PrP Sc levels in neuroblastoma cell lines persistently infected with scrapie prion strains, thus emphasizing the role of lipid rafts in PrP Sc conversion [ 47 , 55 , 62 ].…”

Highly efficient intercellular spreading of protein misfolding mediated by viral ligand - receptor interactions

“…4a). Murine fibroblast cell line L929 39 and CAD5 cells 40 , two cell lines highly permissive to TSE strain 22L, were exposed to VSV-G positive EV (Fig. 4b) and subsequently cultured in the absence of EV for more than 8 passages before testing for the formation of PrP Sc (Fig.…”