Copper drives prion protein phase separation and modulates aggregation

Abstract

Prion diseases are characterized by prion protein (PrP) transmissible aggregation and toxicity in the brain. The physiological function of PrP seems related to sequestering and internalization of redox-active Cu2+. It is unclear whether Cu2+contributes to PrP aggregation, recently shown to be mediated by PrP condensation. We investigated the role of Cu2+and oxidation in PrP condensation and aggregation using multiple biophysical and biochemical methods. We find that Cu2+promotes PrP condensation at the cell surface andin vitrothrough co-partitioning. Molecularly, Cu2+inhibited PrP β-structure and hydrophobic residues exposure. Oxidation, induced by H2O2, triggered liquid-to-solid transition of PrP:Cu2+condensates and promoted amyloid-like PrP aggregation. In cells, overexpression of PrPCinitially protected against Cu2+cytotoxicity but led to PrPCaggregation upon extended copper exposure. Our data suggest that PrP condensates function as a buffer for copper that prevent copper toxicity but can transition into PrP aggregation at prolonged oxidative stress.

The potential role of liquid-liquid phase separation in the cellular fate of the compartments for unconventional protein secretion

“…Given that Grh1 liquid condensation is significantly enhanced in response to acidification and most unconventional secretion events are not constitutive but stress-induced [2], we proposed that the CUPS could represent a phase-separated structure formed by a Grh1-rich phase capable of recruiting UPS cargo. Accordingly, the detected amyloid signature previously observed in cell [26] might be attributable to droplet ageing, a property observed previously in other well-known amyloidogenic proteins [46–48]. We showed here that Grh1 LLPS is fully reversible, enhanced under starvation conditions, and notably diminished under normal conditions, which are observations aligning with the CUPS dynamics observed in vivo [24].…”

The potential role of liquid–liquid phase separation in the cellular fate of the compartments for unconventional protein secretion

“…Given that Grh1 condensation is significantly enhanced in response to acidification and most unconventional secretion events are not constitutive but stress-induced (Rabouille, 2017), we proposed that the CUPS could represent a phase-separated structure formed by a Grh1-rich phase capable of recruiting UPS cargo. Accordingly, the detected amyloid signature previously observed in cell (Fontana et al, 2021) might be attributable to a liquidto-solid transition, a property observed previously in other well-known amyloidogenic proteins (Amaral et al, 2023;Carey & Guo, 2022;Ray et al, 2020). We showed here that Grh1 coacervates were fully reversible, enhanced under starvation conditions, and notably diminished under normal conditions, which are observations aligning with the CUPS dynamics observed in vivo (Bruns et al, 2011).…”

Intermolecular Energy Migration via HomoFRET Captures the Modulation in the Material Property of Phase-Separated Biomolecular Condensates

“…Previous studies have utilized homoFRET to identify and characterize membrane-induced clustering and oligomerization in vitro as well as in membrane-anchored proteins in situ 65 . HomoFRET imaging can be employed to investigate membrane-associated phase separation of membrane-bound proteins 66,67 . Recent studies have shown the tuning of FUS condensates and their material properties by the protein quality control (PQC) machinery, including small heat shock proteins and chaperones 68,69 .…”