NR APAM
AU Winklhofer,K.F.; Heske,J.; Heller,U.; Tatzelt,J.
TI Folding of the prion protein in the secretory pathway: implications for the formation of scrapie prions
QU International Conference - Prion diseases: from basic research to intervention concepts - TSE-Forum, 08.10.-10.10.2003, Gasteig, München - Poster session - BR-42
PT Konferenz-Poster
AB
A hallmark of mammalian prion diseases is the conversion of the cellular prion protein PrPc into an abnormally folded isoform, designated PrPsc. During transit through the secretory pathway PrP is posttranslationally modified by the attachment of two N-linked carbohydrate moieties and a glycosylphosphatidylinositol (GPI) anchor at the C-terminus. To gain insight into mechanisms implicated in the formation of PrPsc we analyzed in vivo folding pathways of PrP and the role of posttranslational modifications.
We could show that the complete N-terminus of PrP including the putative transmembrane domain and the first b-strand could be deleted without interfering with PrP maturation. Helix 1, however, turned out to be a major determinant of PrP folding. Disruption of helix 1 prevented attachment of the GPI anchor and the formation of complex N-linked glycans; instead a high mannose PrP glycoform was secreted into the cell culture supernatant. We also could show that the C-terminal GPI anchor signal sequence, independent of its role in GPI anchor attachment, mediates core glycosylation of nascent PrP. Interestingly, conversion of high mannose glycans to complex type glycans only occurred when PrP was membrane-anchored (1). Furthermore, we demostrated that geldanamycin, an inhibitor of Hsp90, interferes with the formation of complex glycosylated PrP. Similarly to inhibitors of alpha-mannosidases, geldanamycin stabilized a high mannose glycoform and prevented the subsequent processing into complex structures. Moreover, a PrP/Grp94 complex could be isolated from geldanamycin-treated cells. Inhibition of complex glycosylation did not interfere with the GPI anchor attachment and cellular trafficking of PrPc. In scrapie-infected cells, however, high mannose PrPc glycoforms were preferred substrates for the formation of PrPsc(2).
1. Winklhofer et al. (2003), JBC 278, pp. 14961;
2. Winklhofer et al. (2003), Traffic 4, pp. 313
AD Konstanze F. Winklhofer, Johanna Heske, Ulrich Heller, Jörg Tatzelt, Max-Planck-Institute for Biochemistry, Germany
SP englisch
PO Deutschland