NR ATLM
AU Heinig,L.; Stuke,A.W.
TI Regulated prion protein expression for prion conversion monitoring
QU International Conference - Prion 2005: Between fundamentals and society's needs - 19.10.-21.10.2005, Congress Center Düsseldorf - Poster Session: Diagnosis DIA-50
PT Konferenz-Poster
AB
The prion protein (PrP) is known as the infectious agent for different transmissible spongiform encephalopathies (TSE) disease in human, sheep or cow. During pathogenesis the ubiquitous cellular prion protein (PrPc) converts into the infective isoform (PrPsc).
We have created a vector system which expresses the prnp gene of different species after transfection into eukaryotic cells. Prnp wild-type genes were isolated from different mammals such as human, non-human primates, human primates, mouse, cow, sheep and great tumbler. These genes can be regulated by a tetracyline (Tc) responding element in PrP wild-type expressing and prnp knock-out cell lines. The stable expression is controlled by different Tc or doxycycline concentrations to adjust the optimal PrPc expression level for the conversion reaction. Endogenous PrPc is incubated with exogenous PrPsc to monitore the conversion into endogenous PrPsc. The infectious material is homo- and heterologous PrPsc brain homogenate (bovie, mice, non-human primates and human). We used BSE infected stem brain for pilot experiments. Endogenous PrPsc amplification can be observed by proteinase K digestion and Western blotting after cell expansion and passaging. Furthermore, spectroscopy can be used to monitore the ratio of the different isoforms in the infected cell line.
Our system will be used for the detection and characterization of prion diseases on a cellular level. In contrast to animal models, the stably manipulated cell linies shall have a shorter incubation time after prion infection. This may also allow us to investigate the molecular mechanism of the species barrier and PrP conversion.
AD Lars Heinig, Andreas W. Stuke, German Primate Centre, Department of Virology and Immunology, Kellnerweg 4, 37077 Göttingen, Germany
SP englisch
PO Deutschland