Dataset Information

Direct evidence of generation and accumulation of ?-sheet-rich prion protein in scrapie-infected neuroblastoma cells with human IgG1 antibody specific for ?-form prion protein.

ABSTRACT: We prepared ?-sheet-rich recombinant full-length prion protein (?-form PrP) (Jackson, G. S., Hosszu, L. L., Power, A., Hill, A. F., Kenney, J., Saibil, H., Craven, C. J., Waltho, J. P., Clarke, A. R., and Collinge, J. (1999) Science 283, 1935-1937). Using this ?-form PrP and a human single chain Fv-displaying phage library, we have established a human IgG1 antibody specific to ?-form but not ?-form PrP, PRB7 IgG. When prion-infected ScN2a cells were cultured with PRB7 IgG, they generated and accumulated PRB7-binding granules in the cytoplasm with time, consequently becoming apoptotic cells bearing very large PRB7-bound aggregates. The SAF32 antibody recognizing the N-terminal octarepeat region of full-length PrP stained distinct granules in these cells as determined by confocal laser microscopy observation. When the accumulation of proteinase K-resistant PrP was examined in prion-infected ScN2a cells cultured in the presence of PRB7 IgG or SAF32, it was strongly inhibited by SAF32 but not at all by PRB7 IgG. Thus, we demonstrated direct evidence of the generation and accumulation of ?-sheet-rich PrP in ScN2a cells de novo. These results suggest first that PRB7-bound PrP is not responsible for the accumulation of ?-form PrP aggregates, which are rather an end product resulting in the triggering of apoptotic cell death, and second that SAF32-bound PrP lacking the PRB7-recognizing ?-form may represent so-called PrP(Sc) with prion propagation activity. PRB7 is the first human antibody specific to ?-form PrP and has become a powerful tool for the characterization of the biochemical nature of prion and its pathology.

SUBMITTER: Kubota T

PROVIDER: S-EPMC3340157 | biostudies-literature | 2012 Apr

REPOSITORIES: biostudies-literature

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Direct evidence of generation and accumulation of β-sheet-rich prion protein in scrapie-infected neuroblastoma cells with human IgG1 antibody specific for β-form prion protein.

Kubota Toshiya T Hamazoe Yuta Y Hashiguchi Shuhei S Ishibashi Daisuke D Akasaka Kazuyuki K Nishida Noriyuki N Katamine Shigeru S Sakaguchi Suehiro S Kuroki Ryota R Nakashima Toshihiro T Sugimura Kazuhisa K

The Journal of biological chemistry 20120222 17

We prepared β-sheet-rich recombinant full-length prion protein (β-form PrP) (Jackson, G. S., Hosszu, L. L., Power, A., Hill, A. F., Kenney, J., Saibil, H., Craven, C. J., Waltho, J. P., Clarke, A. R., and Collinge, J. (1999) Science 283, 1935-1937). Using this β-form PrP and a human single chain Fv-displaying phage library, we have established a human IgG1 antibody specific to β-form but not α-form PrP, PRB7 IgG. When prion-infected ScN2a cells were cultured with PRB7 IgG, they generated and acc ...[more]

PMID: 22356913

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Project description:Environmental stimulation during brain development is an important risk factor for the development of neurodegenerative disease. Clinical evidence indicates that prenatal exposure to particulate air pollutants leads to diffuse damage to the neurovascular unit in the developing brain and accelerates neurodegeneration. Maternal exposure to carbon black nanoparticles (CB-NPs), used as a model for particulate air pollution, induces long-lasting diffuse perivascular abnormalities. We aimed to comprehensively characterize the perivascular abnormalities related to maternal NPs exposure using Fourier transform infrared microspectroscopy (in situ FT-IR) and classical staining analysis. Pregnant ICR mice were intranasally treated with a CB-NPs suspension (95 μg/kg at a time) on gestational days 5 and 9. Brains were collected 6 weeks after birth and sliced to prepare 10-μm-thick serial sections. Reflective spectra of in situ FT-IR were acquired using lattice measurements (x-axis: 7, y-axis: 7, 30-μm apertures) around a centered blood vessel. We also performed mapping analysis of protein secondary structures. Serial sections were stained with using periodic acid-Schiff or immunofluorescence to examine the phenotypes of the perivascular areas. Peaks of amide I bands in spectra from perivascular areas were shifted by maternal NPs exposure. However, there were two types of peak-shift in one mouse in the exposure group. Some vessels had a large peak-shift and others had a small peak-shift. In situ FT-IR combined with traditional staining revealed that the large peak-shift was induced around blood vessel adjacent to astrocytes with glial fibrillary acidic protein and aquaporin-4 over-expression and perivascular macrophages (PVMs) with enlarged lysosome granules. Furthermore, protein secondary structural analysis indicated that maternal NPs exposure led to increases in β-sheet content and decreases in α-helix content in areas that are mostly close to the centered blood vessel displaying histopathological changes. These results suggest that β-sheet-rich waste proteins, which are denatured by maternal NPs exposure, likely accumulate in the perivascular space as they are processed by the clearance systems in the brain. This may in turn lead the denaturation of PVMs and astrocyte activation. The risk of neurodegeneration may be enhanced by exposure to particulate air pollutants during brain development following the perivascular accumulation of β-sheet-rich waste proteins.

Dataset Information

Direct evidence of generation and accumulation of ?-sheet-rich prion protein in scrapie-infected neuroblastoma cells with human IgG1 antibody specific for ?-form prion protein.

Publications

Direct evidence of generation and accumulation of β-sheet-rich prion protein in scrapie-infected neuroblastoma cells with human IgG1 antibody specific for β-form prion protein.

Similar Datasets

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