Defining the Conformational Features of Anchorless, Poorly Neuroinvasive Prions
Figure 6
GPI–RML prions inoculated into WT mice (GPIRML-WT).
(A) Survival curves of WT mice inoculated with RML or GPI–RML prions (passage 2). (B) PrP and astrocyte immunostaining of brain sections at the level of cerebral cortex and hippocampus show diffuse PrPSc deposits and accompanying gliosis. (C) Lesion profile analysis. For RML- and GPIRML-infected WT mice, the severity of spongiosis, astrogliosis, and PrPSc deposition were scored for nine brain regions (see Methods) and were nearly superimposable. Each ring represents 1 point. (D) PK-digested brain samples from WT mice infected with RML or GPI–RML show no difference in their glycoform profiles. Also shown are Tg(GPI–RML) mice infected with RML, which show a shift to a lower molecular weight and only mono-glycosylated PrP. (E) Conformational stability and (F) thermal stability of RML and GPIRML-WT were nearly identical (n = 4 mice each). (G) Thermal denaturation curves of RML- and GPIRML-infected WT mice show the insoluble PrPSc remaining after heating to various temperatures as measured by ELISA. Right panel, T1/2 values were independently calculated for each mouse and plotted. For (E), plotted are the averages from n = 4 mice per prion, each run in triplicate. For (F), the middle graph shows the mean ± SE of RML and GPI–RML (n = 4 mice each). Graphs (right panels) represent the mean ± SE for all mice. Scale bar = 500 µm.