title: Single cell imaging and quantification of TDP-43 and α-synuclein intercellular propagation.
abstract: The intercellular spreading of protein assemblies is a major factor in the progression of neurodegenerative disorders. The quantitative study and visualization of cell-to-cell propagation using tagged-proteins is challenging due to the steric effect of relatively large fluorescence tags and the risk of 'false positive' identification when analyzing these rare transmission events. Here, we established a cell culture model to characterize the cell-to-cell transmission of TAR DNA-binding protein and α-synuclein, involved in amyotrophic lateral sclerosis and Parkinson's disease, respectively, using the small nine amino acid influenza hemagglutinin tag. The novel use of single cell resolution imaging flow cytometry allowed the visualization and quantification of all individual transmission events. Cell-level analysis of these events indicated that the degree of transfer is lower than previously reported based on conventional flow cytometry. Furthermore, our analysis can exclude 'false positive' events of cellular overlap and extracellular debris attachment. The results were corroborated by high-resolution confocal microscopy mapping of protein localization.
authors.affiliation: Department of Molecular Microbiology and Biotechnology, Tel-Aviv University, Tel Aviv, 6997801, Israel.
authors.forename: Sivan
authors.initials: S
authors.lastname: Peled
authors.affiliation: Department of Molecular Microbiology and Biotechnology, Tel-Aviv University, Tel Aviv, 6997801, Israel.
authors.forename: Dorin
authors.initials: D
authors.lastname: Sade
authors.affiliation: Department of Molecular Microbiology and Biotechnology, Tel-Aviv University, Tel Aviv, 6997801, Israel.|Division of Gastroenterology & Hepatology, Department of Medicine, Weill Medical College of Cornell University, New York, NY, 10021, USA.
authors.forename: Yaron
authors.initials: Y
authors.lastname: Bram
authors.affiliation: Flow Cytometry Unit, Department of Biological Services, Weizmann Institute of Science, Rehovot, Israel.
authors.forename: Ziv
authors.initials: Z
authors.lastname: Porat
authors.affiliation: Department of Molecular Microbiology and Biotechnology, Tel-Aviv University, Tel Aviv, 6997801, Israel.
authors.forename: Topaz
authors.initials: T
authors.lastname: Kreiser
authors.affiliation: Department of Ophthalmology, Rambam Health Care Campus, Haifa, Israel.
authors.forename: Michael
authors.identifier: http://orcid.org/0000-0002-4661-0993
authors.initials: M
authors.lastname: Mimouni
authors.affiliation: Sackler Cellular and Molecular Imaging Center, Sackler Faculty of Medicine, Tel-Aviv University, Tel Aviv, Israel.
authors.forename: Alexandra
authors.initials: A
authors.lastname: Lichtenstein
authors.affiliation: Department of Molecular Microbiology and Biotechnology, Tel-Aviv University, Tel Aviv, 6997801, Israel.|Sagol Interdisciplinary School of Neurosciences, Tel Aviv University, Tel Aviv, 6997801, Israel.
authors.forename: Daniel
authors.initials: D
authors.lastname: Segal
authors.affiliation: Department of Molecular Microbiology and Biotechnology, Tel-Aviv University, Tel Aviv, 6997801, Israel. ehudg@post.tau.ac.il.|Sagol Interdisciplinary School of Neurosciences, Tel Aviv University, Tel Aviv, 6997801, Israel. ehudg@post.tau.ac.il.
authors.forename: Ehud
authors.initials: E
authors.lastname: Gazit
mesh_terms: D002460:Cell Line; D018920:Coculture Techniques; D004268:DNA-Binding Proteins / Q000378:metabolism*; D005434:Flow Cytometry; D005455:Fluorescent Antibody Technique; D006801:Humans; D042541:Intracellular Space; D008856:Microscopy, Fluorescence; D057054:Molecular Imaging / Q000379:methods; D021381:Protein Transport; D059010:Single-Cell Analysis; D051844:alpha-Synuclein / Q000378:metabolism*
chemical_list: D004268:DNA-Binding Proteins; C000598624:TARDBP protein, human; D051844:alpha-Synuclein