Journal Article

DZNE-2020-06536

http://join2-wiki.gsi.de/foswiki/pub/Main/Artwork/join2_logo100x88.png A Bioluminescent Cell Assay to Quantify Prion Protein Dimerization.

Wüsten, A. (First author)DZNE* ; Pasham Parameshwar, R.DZNE* ; Smiyakin, A.DZNE* ; Bernis, M. E.DZNE* ; Tamgüney, G. (Last author)DZNE*

2018
Macmillan Publishers Limited, part of Springer Nature [London]

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Please use a persistent id in citations: doi:10.1038/s41598-018-32581-1

Abstract: The prion protein (PrP) is a cell surface protein that in disease misfolds and becomes infectious causing Creutzfeldt-Jakob disease in humans, scrapie in sheep, and chronic wasting disease in deer and elk. Little is known regarding the dimerization of PrP and its role in disease. We developed a bioluminescent prion assay (BPA) to quantify PrP dimerization by bimolecular complementation of split Gaussia luciferase (GLuc) halves that are each fused to PrP. Fusion constructs between PrP and N- and C-terminal GLuc halves were expressed on the surface of RK13 cells (RK13-DC cells) and dimerized to yield a bioluminescent signal that was decreased in the presence of eight different antibodies to PrP. Dimerization of PrP was independent of divalent cations and was induced under stress. Challenge of RK13-DC cells with seven different prion strains did not lead to detectable infection but was measurable by bioluminescence. Finally, we used BPA to screen a compound library for compounds inhibiting PrP dimerization. One of the most potent compounds to inhibit PrP dimerization was JTC-801, which also inhibited prion replication in RML-infected ScN2a and SMB cells with an EC50 of 370 nM and 220 nM, respectively. We show here that BPA is a versatile tool to study prion biology and to identify anti-prion compounds.

Keyword(s): Animals (MeSH) ; Biological Assay: methods (MeSH) ; Cations, Divalent: metabolism (MeSH) ; Cell Line (MeSH) ; Cell Line, Tumor (MeSH) ; Creutzfeldt-Jakob Syndrome: metabolism (MeSH) ; Deer (MeSH) ; Dimerization (MeSH) ; Humans (MeSH) ; Luminescent Measurements: methods (MeSH) ; Mice (MeSH) ; Prion Proteins: metabolism (MeSH) ; Protein Folding (MeSH) ; Rabbits (MeSH) ; Scrapie: metabolism (MeSH) ; Sheep (MeSH) ; Wasting Disease, Chronic: metabolism (MeSH) ; Cations, Divalent ; Prion Proteins

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Contributing Institute(s):

1. Prion and prion-like diseases (AG Tamgüney 2)

Research Program(s):

1. 342 - Disease Mechanisms and Model Systems (POF3-342) (POF3-342)

Appears in the scientific report 2018

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