Amyloid conformation-dependent disaggregation revealed by a reconstituted yeast prion system

Abstract

Disaggregation of amyloid fibrils is a fundamental biological process required for amyloid propagation. However, due to the lack of experimental systems, the molecular mechanism of how amyloid is disaggregated by cellular factors remains poorly understood. Here, we established a robust, in vitro reconstituted system of yeast prion propagation and found that Hsp104, Ssa1, and Sis1 chaperones are essential for efficient disaggregation of Sup35 amyloid. Real-time imaging of single-molecule fluorescence coupled with the reconstitution system revealed that amyloid disaggregation is achieved by ordered, timely binding of the chaperones to the amyloid. Remarkably, we uncovered two distinct, prion strain conformation-dependent modes of disaggregation, fragmentation and dissolution. We characterized distinct chaperon dynamics in each mode and found that transient, repeated binding of Hsp104 to the same site of amyloid results in fragmentation. These findings provide a physical foundation for otherwise puzzling in vivo observations and for therapeutic development for amyloid-associated neurodegenerative diseases.