Abstract
Prions replicate in the host cell by the self-propagating refolding of the normal cell surface protein, PrP(C), into a beta-sheet-rich conformer, PrP(Sc). Exposure of cells to prion-infected material and subsequent endocytosis can sometimes result in the establishment of an infected culture. However, the relevant cell surface receptors have remained unknown. We have previously shown that cellular heparan sulfates (HS) are involved in the ongoing formation of scrapie prion protein (PrP(Sc)) in chronically infected cells. Here we studied the initial steps in the internalization of prions and in the infection of cells. Purified prion "rods" are arguably the purest prion preparation available. The only proteinaceous component of rods is PrP(Sc). Mouse neuroblastoma N2a, hypothalamus GT1-1, and Chinese hamster ovary cells efficiently bound both hamster and mouse prion rods (at 4 degrees C) and internalized them (at 37 degrees C). Treating cells with bacterial heparinase III or chlorate (a general inhibitor of sulfation) strongly reduced both binding and uptake of rods, whereas chondroitinase ABC was inactive. These results suggested that the cell surface receptor of prion rods involves sulfated HS chains. Sulfated glycans inhibited both binding and uptake of rods, probably by competing with the binding of rods to cellular HS. Treatments that prevented endocytosis of rods also prevented the de novo infection of GT1-1 cells when applied during their initial exposure to prions. These results indicate that HS are an essential part of the cellular receptor used both for prion uptake and for cell infection. Cellular HS thus play a dual role in prion propagation, both as a cofactor for PrP(Sc) synthesis and as a receptor for productive prion uptake.
Highlights
The transmissible spongiform encephalopathies (TSE), which comprise infectious, familial, and sporadic neurodegenerations such as Creutzfeldt-Jakob disease (CJD) of humans [1], scrapie of sheep and bovine spongiform encephalopathy (BSE) [2], are caused by prions [3]
Infected ScN2a-M cells were treated for 5 days with these compounds, and protease-resistant PrP Sc was analyzed by Western blots (WB) developed with 3F4
The ability of these compounds to reduce rods internalization correlated well with their anti-PrPSc potency (compare panels A and B, and Fig. 6; we have previously reported that heparan mimetics (HMs) are not general endocytosis inhibitors [32])
Summary
The transmissible spongiform encephalopathies (TSE), which comprise infectious, familial, and sporadic neurodegenerations such as Creutzfeldt-Jakob disease (CJD) of humans [1], scrapie of sheep and bovine spongiform encephalopathy (BSE) [2], are caused by prions [3] These proteinaceous agents are thought to propagate by refolding a normal cell surface glycoprotein of the host, the cellular prion protein PrPC, into an abnorma l ß-sheet rich [4,5] conformation (reviewed in 6). Prion “rods” are arguably the purest form of prions known [20,21] These infectious, unbranched amyloidic structures are prepared from prioninfected tissues by the combined action of detergents and proteases [22] (often supplemented by nucleases) and their only proteinaceous component is PrP27-30 [23], the protease-resistant core of PrP Sc. The size of rods is very heterogeneous, and they may contain up to several thousands PrP molecules [20]. We set out to characterize receptors for rods in two infectible mouse cell lines, the neurobla stoma N2a [24] and the hypothalamic cell line GT1-1 [25,26,27], and Chinese hamster ovary (CHO) cells, which seem to be refractive to prion infection
Sign-in/Register to view highlights & summary Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
