A Bioluminescent Cell Assay to Quantify Prion Protein Dimerization

• Published in: Scientific reports Vol. 8; no. 1; pp. 14178 - 14
• Main Authors: Wüsten, Katharina Annick, Reddy, Pasham Parameshwar, Smiyakin, Andrej, Bernis, Maria Eugenia, Tamgüney, Gültekin
• Format: Journal Article
• Language: English
• Published: London Nature Publishing Group UK 21.09.2018; Nature Publishing Group
• Subjects: 13; 13/1; 13/31; 631/154/1435/2163; 692/617/375/1937; 82; 96; Bioluminescence; Cations; Cell surface; Chronic wasting disease; Creutzfeldt-Jakob disease; Deer; Dimerization; Humanities and Social Sciences; multidisciplinary; Prion protein; Protein folding; Proteins; Science; Science (multidisciplinary); Scrapie; Transmissible spongiform encephalopathy; RK13 Cells; Prion Protein (PrP); Gaussia Luciferase; Bioluminescence; Prion Strains
• ISSN: 2045-2322; 2045-2322
• DOI: 10.1038/s41598-018-32581-1

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 b ioluminescent p rion a ssay (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 EC 50 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.