Figure 9. Sho protein in chronically infected SMB cells.

a) Prion-infected SMB cells (SMB Sc) or pentosan sulfate cured SMB cells (SMB-PS) were acutely transfected with a control cDNA expression vector (empty vector, e.v.) or the same plasmid vector bearing a cDNA insert encoding wt mouse Sho (Sho). Upper panel: Cell lysates were prepared from cells 24 hours after transfection and were immunoblotted with a C-terminal anti-Sho antibody 06SH3a. Lower panel: A similar transfection efficiency into healthy and infected cells was established by probing for expression of neomycin phosphotransferase encoded within both varieties of plasmid vector (“NPT II”). C1 is a C-terminal fragment of Sho. b) The presence of PrP^Sc in infected SMB cultures was verified by protease digestion. Two PK resistant fragments co-migrated with fragments present in undigested lysates of infected cells, but not those of uninfected cells (open arrowheads; see also Figure S1). c) Conditioned medium was harvested from the transfected cells, acetone precipitated and then western blotted for the presence of the Sho glycoprotein with 06SH3a antibody.