Transfer of Scrapie Prion Infectivity by Cell Contact in Culture

When a cell is infected with scrapie prions, newly synthesized molecules of the prion protein PrP C are expressed at the cell surface and may subsequently be converted to the abnormal form PrP Sc. In an experimental scrapie infection of an animal, the initial innoculum of PrP Sc is cleared relatively rapidly, and the subsequent propagation of the infection depends on the ability of infected cells to convert uninfected target cells to stable production of PrP Sc. The mechanism of such cell-based infection is not understood. We have established a system in dissociated cell culture in which scrapie-infected mouse SMB cells are able to stably convert genetically marked target cells by coculture. After coculture and rigorous removal of SMB cells, the target cells express PrP Sc and also incorporate [ 35 S]methionine into PrP Sc. The extent of conversion was sensitive to the ratio of the two cell types, and conversion by live SMB required 2500-fold less PrP Sc than conversion by a cell-free prion preparation. The conversion activity of SMB cells is not detectable in conditioned medium and apparently depends on close proximity or contact, as evidenced by culturing the SMB and target cells on neighboring but separate surfaces. SMB cells were killed by fixation in aldehydes, followed by washing, and were found to retain significant activity at conversion of target cells. Cell-mediated infection of target cells in this culture system is effective and requires significantly less PrP Sc than infection by a prion preparation. Several lines of evidence indicate that it depends on cell contact, in particular, the activity of aldehyde-fixed infected cells.