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Comparison of abnormal isoform of prion protein in prion-infected cell lines and primary-cultured neurons by PrPSc-specific immunostaining

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J.Gen.Virol.v.97p.2030-2042(2016) .pdf10.07 MBPDFView/Open
Please use this identifier to cite or link to this item:http://hdl.handle.net/2115/66951

Title: Comparison of abnormal isoform of prion protein in prion-infected cell lines and primary-cultured neurons by PrPSc-specific immunostaining
Authors: Tanaka, Misaki Browse this author
Fujiwara, Ai Browse this author
Suzuki, Akio Browse this author
Yamasaki, Takeshi Browse this author
Hasebe, Rie Browse this author →KAKEN DB
Masujin, Kentaro Browse this author
Horiuchi, Motohiro Browse this author →KAKEN DB
Keywords: prions
Immunofluorescence assay
primary neurons
guanidine isothiocyanate
monoclonal antibody
Issue Date: Aug-2016
Publisher: Society for General Microbiology
Journal Title: Journal of General Virology
Volume: 97
Start Page: 2030
End Page: 2042
Publisher DOI: 10.1099/jgv.0.000514
Abstract: We established abnormal isoform of prion protein (PrPSc)-specific double immunostaining using mAb 132, which recognizes aa 119-127 of the PrP molecule, and novel PrPSc-specific mAb 8D5, which recognizes the N-terminal region of the PrP molecule. Using the PrPSc-specific double immunostaining, we analysed PrPSc in immortalized neuronal cell lines and primary cerebral-neuronal cultures infected with prions. The PrPSc-specific double immunostaining showed the existence of PrPSc positive for both mAbs 132 and 8D5, as well as those positive only for either mAb 132 or mAb 8D5. This indicated that double immunostaining detects a greater number of PrPSc species than single immunostaining. Double immunostaining revealed cell-type-dependent differences in PrPSc staining patterns. In the 22 L prion strain-infected Neuro2a (N2a)-3 cells, a subclone of N2a neuroblastoma cell line, or GT1-7, a subclone of the GT1 hypothalamic neuronal cell line, granular PrPSc stains were observed at the perinuclear regions and cytoplasm, whereas unique string-like PrPSc stains were predominantly observed on the surface of the 22 L strain-infected primary cerebral neurons. Only 14% of PrPSc in the 22 L strain-infected N2a-3 cells were positive for mAb 8D5, indicating that most of the PrPSc in N2a-3 lack the N-terminal portion. In contrast, nearly half PrPSc detected in the 22 L strain-infected primary cerebral neurons were positive for mAb 8D5, suggesting the abundance of full-length PrPSc that possesses the N-terminal portion of PrP. Further analysis of prion-infected primary neurons using PrPSc-specific immunostaining will reveal the neuron-specific mechanism for prion propagation.
Type: article (author version)
URI: http://hdl.handle.net/2115/66951
Appears in Collections:獣医学院・獣医学研究院 (Graduate School of Veterinary Medicine / Faculty of Veterinary Medicine) > 雑誌発表論文等 (Peer-reviewed Journal Articles, etc)

Submitter: 堀内 基広

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