NR ARDT
AU Brabeck,C.; Niemann,H.; Kloz,U.; Bürkle,A.
TI Construction of transgenic mice expressing a dominant-negative PrP deletion mutant
QU TSE-Forum, 4. Kongress - Nationale TSE-Forschungsplattform, Düsseldorf 28.10.-29.10.2004, Poster GL-02
PT Konferenz-Poster
AB The prion protein (PrPc) is a GPI-anchored 35 kDa membrane glycoprotein essential for the pathogenesis of several inherited and transmissible neurodegenerative diseases such as scrapie in sheep, bovine spongiform encephalopathy (BSE) in cattle and Creutzfeldt-Jakob disease (CJD) in humans. We have focused our interest on the so-called TM1 region of PrPc (codons 110-135) being one of the most highly conserved regions of the molecule. This region comprises of an array of hydrophobic amino acids and resides in a highly flexible and unstructured portion of the protein as revealed by NMR analysis of recombinant PrP. It has been proposed that the TM1 domain plays an important role in the conversion process of PrPc into PrPsc. In previous work, we have constructed a deletion mutant lacking codons 114-121 of PrPc termed delta114-121. Using persistently infected mouse neuroblastoma cells (Sc-N2a) as a model system for prion replication, we could show that delta114-121 itself cannot be converted into a protease-resistant isoform and that it furthermore behaves as a dominant-negative version of PrPc as the accumulation of endogenous PrPsc was significantly reduced in Sc-N2a cells in the presence of delta114-121 (Hölscher et al, J Virol 1998, 72:1153-9). To further investigate the properties of delta114-121 in an in-vivo system, we now aimed to create transgenic mice expressing the deletion mutant either on a Prnp+/+ or Prnp-/- background. For this purpose, the delta114-121 ORF was subcloned into the expression plasmid MoPrP.Xho that allows expression of the construct under the control of the prion protein gene promoter (Borchelt et al, Genet Anal 1996, 13:159-63). Microinjection of MoPrPdelta114-121 into oocytes of wild-type mice gave rise to transgenic delta114-121 mice that are being backcrossed onto a 129SV wild-type genetic background. Analysis of delta114-121 mice revealed that the animals are viable and fertile and do not show any gross phenotypic changes. Transgenic delta114-121 mice expressing our PrP deletion mutant on a Prnp+/+ background will provide an excellent tool to study the dominant-negative properties of delta114-121 in the context of an in-vivo system. In addition, these mice can be useful in helping to elucidate the hitherto unknown physiological role of PrPc.
AD Christine Brabeck, Alexander Bürkle, Molecular Toxicology Group, University of Konstanz, Box X911, Konstanz, Germany; Hartmut Niemann, Alexander Bürkle, Abteilung Tumorvirologie, Deutsches Krebsforschungszentrum, Heidelberg, Germany; Ulrich Kloz, Transgenic Core Facility, Deutsches Krebsforschungszentrum, Heidelberg, Germany
SP englisch
PO Deutschland
OR Tagungsband