NR ATHW
AU Mehlhase,J.; Löwer,J.; Montrasio,F.
TI Cytosolic Prion Protein (PrP23-231) is Non Toxic in Neuro-2a Cells
QU International Conference - Prion 2005: Between fundamentals and society's needs - 19.10.-21.10.2005, Congress Center Düsseldorf - Poster Session: Cell Biology of PrPc and PrPsc CELL-28
PT Konferenz-Poster
AB
Transmissible spongiform encephalopathies are lethal transmissible neurodegenerative disorders caused by prions, infectious agents that are devoid of informational nucleic acids and consist of an "infectious" protein (termed PrPsc) capable of converting a normal host protein called PrPc into a likeness of themselve. However, the processes leading to neuronal degeneration in the central nervous system upon prion infections are still not identified. In the recent past, it has been proposed that neuronal death might be triggered by accumulation of PrP in the cytosol due to impairment of proteasomal degradation of misfolded PrP molecules retranslocated from the endoplasmatic reticulum.
In order to elucidate the molecular mechanisms leading to cytotoxic effects of cytosolic PrP, we used transiently transfected Neuro-2a cell lines conditionally expressing either wild-type (wtPrP)- or cytosolic-PrP (Cy-PrP(23-231)). Here we show, that upon ecdysone-induction, Cy-PrP(23-231) was expressed at levels that were detectable by immunoblot technique. Furthermore, specific inhibition of the proteasomal degradation system led to an accumulation of Cy-PrP(23-231). Cellular localisation analysis of Cy-PrP(23-231) by immunofluorescence and subcellular fractionation surprisingly showed that Cy-PrP(23-231) was mainly associated with endoplasmatic reticulum (ER) structures.
We then analyzed neurotoxic properties of Cy-PrP(23-231) by the quantification of Neuro-2a cells viability and apoptosis via MTT, LDH and caspase 3 activation assays. In contrast to data generated by others, we did not observe any neurotoxic effects of Cy-PrP(23-231) both in the absence or presence of proteasome inhibitors. Therefore, our work fails to confirm a potential role of cytosolic localised PrP molecules in prion neurotoxicity.
AD J.Mehlhase, J.Löwer, F.Montrasio, Prion Research Group, Paul-Ehrlich-Institut, Langen, Germany
SP englisch
PO Deutschland