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Proc Natl Acad Sci U S A. 2015 Sep 1;112(35):E4949-58. doi: 10.1073/pnas.1513426112. Epub 2015 Aug 18.

Propagation of prions causing synucleinopathies in cultured cells.

Author information

1
Institute for Neurodegenerative Diseases, University of California, San Francisco, CA 94143;
2
Institute for Neurodegenerative Diseases, University of California, San Francisco, CA 94143; Department of Neurology, University of California, San Francisco, CA 94143;
3
Institute for Neurodegenerative Diseases, University of California, San Francisco, CA 94143; Daiichi Sankyo Company, Limited, Tokyo 140-8710, Japan;
4
Daiichi Sankyo Company, Limited, Tokyo 140-8710, Japan;
5
Department of Pathology, University of California, San Francisco, CA 94143;
6
Center for Alzheimer's and Neurodegenerative Diseases, University of Texas Southwestern Medical Center, Dallas, TX 75390;
7
Department of Neurology, University of California, San Francisco, CA 94143; Department of Pathology, University of California, San Francisco, CA 94143;
8
Ageing Research Unit, School of Public Health, Imperial College London, London SW7 2AZ, United Kingdom;
9
Centre for Neuroinflammation and Neurodegeneration, Department of Medicine, Imperial College London, London SW7 2AZ, United Kingdom;
10
C. S. Kubik Laboratory for Neuropathology, Department of Pathology, Massachusetts General Hospital, Boston, MA 02114;
11
Department of Molecular and Cellular Physiology and Howard Hughes Medical Institute, Stanford University School of Medicine, Stanford, CA 94305;
12
Institute for Neurodegenerative Diseases, University of California, San Francisco, CA 94143; Department of Neurology, University of California, San Francisco, CA 94143; Department of Biochemistry and Biophysics, University of California, San Francisco, CA 94143 stanley@ind.ucsf.edu.

Abstract

Increasingly, evidence argues that many neurodegenerative diseases, including progressive supranuclear palsy (PSP), are caused by prions, which are alternatively folded proteins undergoing self-propagation. In earlier studies, PSP prions were detected by infecting human embryonic kidney (HEK) cells expressing a tau fragment [TauRD(LM)] fused to yellow fluorescent protein (YFP). Here, we report on an improved bioassay using selective precipitation of tau prions from human PSP brain homogenates before infection of the HEK cells. Tau prions were measured by counting the number of cells with TauRD(LM)-YFP aggregates using confocal fluorescence microscopy. In parallel studies, we fused α-synuclein to YFP to bioassay α-synuclein prions in the brains of patients who died of multiple system atrophy (MSA). Previously, MSA prion detection required ∼120 d for transmission into transgenic mice, whereas our cultured cell assay needed only 4 d. Variation in MSA prion levels in four different brain regions from three patients provided evidence for three different MSA prion strains. Attempts to demonstrate α-synuclein prions in brain homogenates from Parkinson's disease patients were unsuccessful, identifying an important biological difference between the two synucleinopathies. Partial purification of tau and α-synuclein prions facilitated measuring the levels of these protein pathogens in human brains. Our studies should facilitate investigations of the pathogenesis of both tau and α-synuclein prion disorders as well as help decipher the basic biology of those prions that attack the CNS.

KEYWORDS:

Parkinson’s disease; multiple system atrophy; neurodegeneration; strains; α-synuclein

PMID:
26286986
PMCID:
PMC4568231
DOI:
10.1073/pnas.1513426112
[Indexed for MEDLINE]
Free PMC Article

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