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@PHDTHESIS{Wsten:144823,
author = {Wüsten, Annick},
title = {{E}in zellbasierter {B}iolumineszenzassay zur
{U}ntersuchung der {D}imerisierung und {A}ggregation des
{P}rionproteins},
school = {Rheinische Friedrich-Wilhelms-Universität Bonn},
type = {Dissertation},
address = {Bonn},
reportid = {DZNE-2020-00265},
pages = {92 pages : illustrations},
year = {2017},
note = {Dissertation, Rheinische Friedrich-Wilhelms-Universität
Bonn, 2017},
abstract = {Prion diseases like Creutzfeldt-Jakob disease (CJD) are
fatal neurodegenerative disorders that are triggered by
misfolding of the cellular prion protein (PrPC) to an
infectious isoform rich in β-sheet structure termed PrPSc.
There are evidence that PrPC possibly forms dimers
indicating that dimerization of PrPC may be an essential
intermediate step for the formation of PrPSc aggregates upon
infection.To study PrP dimerization and PrP aggregation upon
infection, a bimolecular complementation assay was developed
using RK13 cells that lack endogenous PrP expression and
stably expressed two fusion constructs between PrP and each
half of split Gaussia luciferase. In these cells
bioluminescence only occured when PrP dimerized or
aggregated and the two split luciferase halves complemented
each other.Using a panel of eight antibodies directed
against different PrP domains, the central domain of PrPC
was identified to be critical for dimerization.Unlike normal
brain homogenate infection of RK13 cells with brain
homogenates from diseased animals containing six different
mouse-adapted prion strains, RML, 22L, ME7, 87V, 79A, and
22A, lead to an increase in bioluminescence. In contrast,
treatment of chronically infected but not uninfected RK13
cells with the anti-prion compound quinacrine reduced
bioluminescence.Finally, a compound-screening with a
compound library including 1650 bioactive compounds based on
the bimolecular complementation assay identified JTC-801 as
one compound that inhibited PrPC-dimerization. Treatment of
ScN2a cells with JTC-801 reduced PrPSc levels in these cells
at nanomolar concentrations.Overall, these results showed
that PrPC clearly dimerizes in cells and that prion
infection led to increased bioluminescence in the newly
developed bimolecular complementation assay. The
identification of JTC-801 as a bioactive compounds that
inhibited PrPC-dimerization and reduced PrPSc-levels in
ScN2a cells at nanomolar concentrations suggested that PrPC
dimerization may be a critical step in the conversion
process of PrPC to PrPSc.},
cin = {AG Tamgüney},
cid = {I:(DE-2719)1013022},
pnm = {342 - Disease Mechanisms and Model Systems (POF3-342)},
pid = {G:(DE-HGF)POF3-342},
typ = {PUB:(DE-HGF)11},
urn = {urn:nbn:de:hbz:5n-48485},
url = {https://pub.dzne.de/record/144823},
}