NR AOXR
AU Paquet,S.; Chapuis,J.; Laude,H.; Vilette,D.
TI From the entry of prion to the initiation of cellular infection
QU International Conference - Prion diseases: from basic research to intervention concepts - TSE-Forum, 08.10.-10.10.2003, Gasteig, München - Poster session - PG-28
PT Konferenz-Poster
AB
During prion infections, the glycoprotein PrPc is converted into a conformational isoform, PrPsc. This abnormal protein, partially resistant to degradation by proteases, is proposed to be the infectious agent. Although PrPsc has been observed in different cellular compartments (plasma membrane, endosomes and lysozomes), many questions are still raised concerning the molecular and cellular mechanisms of prion entry into cells, and the subcellular localization of the sites where PrPc is converted into PrPsc.
Our study uses rabbit epithelial Rov cells, permissive to ovine prion replication. Under doxycyclin treatment, these cells stably express a transfected ovine PrP and after contact with infectious sheep prion, they efficiently replicate the ovine agent (Vilette et al., 2001).
Using confocal microscopy and cell surface immunoprecipitation, we show that PrPc expression is restricted to the apical side of the Rov cell membrane. We also show that the infection of polarized cells is obtained exclusively through the apical side of the cells. We then analysed the cellular entry of the PrPsc present in the infectious inoculum. Rov cells, in which PrPc expression has or has not been induced, were exposed to an infectious inoculum, and the association of prion infectivity and PrPsc to the cells was analysed by bioassay or biochemically. Our results show that PrPsc from the inoculum becomes cell-associated with an efficiency that doesn't depend on the presence of cellular PrP. The associated PrPsc persists several days in the cells. However, the PrPsc associated by cells expressing no PrPc is unable to promote infection once PrP expression has been induced in these cells.
Overall, our data suggest that the conversion of PrPc into PrPsc is restricted to specific subcellular sites located in the plasma membrane or in a yet unknown compartment permissive to the conversion, access to which is controlled by PrPc expression.
AD Sophie Paquet, Jérôme Chapuis, Hubert Laude, Didier Vilette, INRA, France
SP englisch
PO Deutschland