In situ characterization of prion protein structure and metal accumulation in scrapie-infected cells by synchrotron infrared and X-ray imaging

Abstract

Transmissible spongiform encephalopathies (TSEs) such as scrapie are neurodegenerative diseases that are characterized by the accumulation of a misfolded prion protein. During the pathogenesis, the normally α-helical prion protein (PrP C) refolds to the proteinase K-resistant abnormal form (PrP Sc), which is high in β-sheet content. In this study, dorsal root ganglia from the 263K hamster model of scrapie were cut into 8 μm thin sections and mounted onto an aluminium-coated glass slide. Single neuronal cells were mapped with synchrotron infrared microspectroscopy to study protein structure and X-ray fluorescence microprobe to determine metal accumulation. Both techniques provide a spatial resolution of 5–10 μm. Results showed that some IR spectra of the infected cells exhibited a significant frequency shift of the amide I band, where an increased peak intensity was observed at ∼1631 cm −1 (attributed to β-sheet) and a decreased peak intensity at ∼1658 cm −1 was also seen (attributed to α-helix). These shifts were not observed in non-infected cells. Adjacent sections were also stained with an antibody for the prion protein in order to confirm that the IR observations were associated with accumulated PrP Sc. Finally, the same cells were measured by X-ray fluorescence microprobe, which provided the first direct comparison of PrP Sc formation and metal ion accumulation. Results showed an accumulation of Fe in cells infected with PrP Sc.