Production of seedable Amyloid-β peptides in model of prion diseases upon PrPSc-induced PDK1 overactivation

Abstract

The presence of amyloid beta (Aβ) plaques in the brain of some individuals with Creutzfeldt-Jakob or Gertsmann-Straussler-Scheinker diseases suggests that pathogenic prions (PrP^Sc) would have stimulated the production and deposition of Aβ peptides. We here show in prion-infected neurons and mice that deregulation of the PDK1-TACE α-secretase pathway reduces the Amyloid Precursor Protein (APP) α-cleavage in favor of APP β-processing, leading to Aβ40/42 accumulation. Aβ predominates as monomers, but is also found as trimers and tetramers. Prion-induced Aβ peptides do not affect prion replication and infectivity, but display seedable properties as they can deposit in the mouse brain only when seeds of Aβ trimers are co-transmitted with PrP^Sc. Importantly, brain Aβ deposition accelerates death of prion-infected mice. Our data stress that PrP^Sc, through deregulation of the PDK1-TACE-APP pathway, provokes the accumulation of Aβ, a prerequisite for the onset of an Aβ seeds-induced Aβ pathology within a prion-infectious context.

Fig. 1

PrP^Sc deregulation of the PDK1-TACE pathway promotes the accumulation of Aβ monomers and multimers. Concentrations of Aβ40 (a), Aβ42 (b), sAPPβ (c), sAPPα (d), and multimers of Aβ40 (e) and Aβ42 (f) in the culture medium of uninfected serotonergic 1C11^5-HT and Fk-infected 1C11^5-HT neuronal cells (Fk-1C11^5-HT) treated or not with the PDK1 inhibitor BX912 (1 µM) or a combination of BX912 (1 µM) and the TACE inhibitor TAPI-2 (100 µM) for 1 h, deduced from LC-MS/MS (a–d) and ESI-IM-MS (e–f) analyses. g–h Aβ40 and Aβ42 clearance ratios measured in the culture medium of 1C11^5-HT and Fk-1C11^5-HT neuronal cells treated or not with BX912 (1 µM) for 1 h. Values are means ± sem of six independent experiments. n.d. not detected. Data in Fig. 1a–d were analyzed using the two way ANOVA test with Bonferroni post-test correction. Data in Fig. 1g–h were analyzed using the two-tail Student t-test. *denotes p < 0.05, **p < 0.01, and ***p < 0.001. Source data are provided as a Source Data file

Fig. 2

APP silencing does not affect PrP^Sc replication in 1C11 cells. a Levels of Aβ40 and Aβ42 quantified by LC-MS/MS in the culture medium and lysates of Fukuoka-infected 1C11 cells expressing (Fk-1C11) or not (APP^null-Fk-1C11) the amyloid precursor protein APP. b ESI-IM-MS detection and quantification of Aβ40 and Aβ42 trimers and tetramers in the culture medium of Fk-1C11 and APP^null-Fk-1C11 cells. c ELISA-based quantification of proteinase K-resistant PrP^Sc in Fk-1C11 and APP^null-Fk-1C11 cells. Values are means ± sem of six independent experiments. n.d. not detected. Data were analyzed using the two-tail Student t-test. ^**denotes p < 0.01. Source data are provided as a Source Data file

Fig. 3

Aβ-free or not PrP^Sc inocula similarly affect motor activity and survival of C57Bl/6 J mice. a Survival curves of C57Bl/6 J mice inoculated with 1 × 10⁵ uninfected 1C11, Fk-1C11 or APP^null-Fk-1C11 cells via the intracerebral route (i.c.) infused or not with BX912 by intraperitoneal injection (i.p.) starting at 130 days after infection (5 mg per kg body weight per day; 0.25 µl h⁻¹; n = 10 per group). b Static rod test at 160 days after infection in mice inoculated with Fk-1C11 or APP^null-Fk-1C11 cells and treated or not with BX912 (n = 6–10 depending on the group) and in mice inoculated with uninfected 1C11 cells. c Post-mortem ELISA quantification of proteinase K-resistant PrP^Sc in brains of C57Bl/6 J mice infected with Fk-1C11 or APP^null-Fk-1C11 cells-derived inocula and treated or not with BX912. Values are means ± sem. Data in Fig. 3b were analyzed using the two way ANOVA test with Bonferroni post-test correction. Data in Fig. 3c were analyzed using the two-tail Student t-test. ***denotes p < 0.001 vs. mice inoculated with uninfected 1C11 cells, *p < 0.05 vs. untreated Fk-1C11 injected mice, and **p < 0.01 vs. untreated APP^null-Fk-1C11 injected mice. Source data are provided as a Source Data file

Fig. 4

Deregulation of the PDK1-TACE axis in prion-infected C57Bl/6 J mice causes CSF accumulation of Aβ monomers and multimers. Concentrations at 160 dpi of Aβ40 (a), Aβ42 (b), sAPPβ (c), sAPPα (d), and multimers of Aβ40 (e) and Aβ42 (f) in the CSF of C57Bl/6 J mice inoculated with uninfected 1C11, Fk-1C11 or APP^null-Fk-1C11 cells and infused or not with the PDK1 inhibitor BX912 (n = 6–10 depending on the group) deduced from LC-MS/MS (a–d) and ESI-IM-MS (e–f) analyses. g–h Aβ40 and Aβ42 clearance ratios measured in the CSF of mice inoculated with uninfected 1C11 or Fk-1C11 cells and infused or not with BX912 (n = 5). Values are means ± sem. n.d. not detected. Data in Fig. 4a–d were analyzed using the two way ANOVA test with Bonferroni post-test correction. Data in Fig. 4e–h were analyzed using the two-tail Student t test. *denotes p < 0.05, and **p < 0.01. Source data are provided as a Source Data file

Fig. 5

Addition of synthetic Aβ trimers to PrP^Sc inocula amplifies the production of Aβ multimers. a Amount of PrP^Sc in 1C11^5-HT neuronal cells exposed to 1C11, Fk-1C11, or APP^null-Fk-1C11 cells-derived homogenates supplemented or not with synthetic Aβ trimers for 10 days, as assessed by ELISA after proteinase K digestion. Concentrations of Aβ40 (b), Aβ42 (c), and multimers of Aβ40 (d) and Aβ42 (e) in the cell culture medium of 1C11^5-HT neuronal cells treated as in a, deduced from LC-MS/MS (b, c) and ESI-IM-MS (d, e) analyses. Values are means ± sem of six independent experiments. n.d. not detected. Data were analyzed using the two way ANOVA test with Bonferroni post-test correction. **denotes p < 0.01. Source data are provided as a Source Data file

Fig. 6

Aβ seeds co-transmitted with PrP^Sc provoke brain Aβ deposition and decrease the survival of APP23 mice. a Time-dependent ¹¹C-PIB uptake (% of the Injection Dose g⁻¹, upper panel) and survival curves (lower panel) of transgenic APP23 mice inoculated via the intracerebral route (i.c.) with 2.5 × 10⁵ Fk-1C11 or APP^null-Fk-1C11 cells-derived homogenates supplemented or not with synthetic Aβ trimers, and infused or not with the PDK1 inhibitor BX912 (n = 10 per group). mpi months post inoculation, dpi days post inoculation. Each number refers to one mouse in each group. b, c ELISA-based quantification of Proteinase K-resistant PrP^Sc amount (b) and PDK1 activity (c) in the brain of APP23 mice treated as in a at the end stage of prion disease or when sacrificed at 260 dpi. Values are means ± sem. Data in Fig. 6b–c were analyzed using the two way ANOVA test with Bonferroni post-test correction. *denotes p < 0.05, **p < 0.01, and ***p < 0.001 vs. APP23 mice inoculated with non-infectious 1C11 cells-derived homogenates supplemented or not with Aβ trimers. Source data are provided as a Source Data file