NR AKQK
AU Scott,M.R.D.; Kohler,R.; Foster,D.B.; Prusiner,S.B.
TI Chimeric prion protein expression in cultured cells and transgenic mice
QU Protein Science 1992 Aug; 1(8): 986-97
IA http://www.proteinscience.org/cgi/reprint/1/8/986
PT journal article
AB The efficient expression of exogenous prion protein (PrP) molecules in mouse neuroblastoma cells that are chronically infected with murine scrapie prions (ScN2a cells; Butler, D.A., et al., 1988, J. Virol. 62, 1558-1564) and in transgenic mice is described. This technology allows investigation of the PrP molecule for structural regions involved in determining species specificity, as well as ablation experiments designed to address the functionality of particular regions of the PrP molecule. Previous reports demonstrated that the PrP gene specifies the host range for susceptibility of transgenic animals to prions (Scott, M., et al., 1989, Cell 59, 847-857; Prusiner, S.B., et al., 1990, Cell 63, 673-686). Consistent with these results, we showed that Syrian hamster (SHa) PrP is ineligible for efficient conversion to PrPsc in ScN2a cells. By constructing a series of chimeric mouse (Mo)/SHaPrP genes, we developed an epitopically tagged functional variant of the MoPrP gene, which can efficiently form protease-resistant PrP molecules upon expression in ScN2a cells. The presence of a defined epitope for an SHa-specific monoclonal antibody allows the products of this chimeric gene to be discriminated from endogenous MoPrP and creates a useful reagent for exploring structure/function relationships via targeted mutagenesis. In addition, we developed a transgenic mouse expression vector by manipulation of an SHaPrP cosmid clone. This vector permits the efficient expression of foreign PrP genes in the brains of transgenic animals, enabling pathological consequences of in vitro mutagenesis to be studied.
MH Animal; Base Sequence; Brain/*metabolism; Chimeric Proteins/*biosynthesis/isolation & purification; Cosmids; Genetic Vectors; Hamsters; Kanamycin Kinase; Mesocricetus; Mice; Mice, Transgenic; Molecular Sequence Data; Oligodeoxyribonucleotides; Phosphotransferases/biosynthesis/genetics; Plasmids; Prions/*biosynthesis/genetics; Recombinant Proteins/biosynthesis/isolation & purification; Restriction Mapping; Support, Non-U.S. Gov't; Support, U.S. Gov't, P.H.S.; Transfection; Tumor Cells, Cultured
AD Department of Neurology, University of California, San Francisco 94143.
SP englisch
PO USA