Dissecting aggregation and seeding dynamics of α-Syn polymorphs using the phasor approach to FLIM

Area of Science:

• Neurodegenerative diseases
• Protein misfolding and aggregation
• Cellular biology

Background:

• Synucleinopathies involve pathological alpha-synuclein (α-Syn) accumulation.
• The structural diversity (polymorphism) of α-Syn assemblies is linked to disease subtypes and progression.
• Mechanisms driving α-Syn polymorphism and its cellular impact remain unclear.

Purpose of the Study:

• To investigate seeded aggregation of α-Syn using fluorescence lifetime imaging microscopy (FLIM).
• To determine if distinct α-Syn polymorphs have characteristic fluorescence lifetimes.
• To analyze the differential processing of α-Syn polymorphs by cellular clearance pathways.

Main Methods:

• Utilized a biosensor cell line for seeded aggregation studies.
• Employed fluorescence lifetime imaging microscopy (FLIM) to characterize α-Syn structures.
• Monitored cellular clearance pathways' interaction with α-Syn polymorphs.

Main Results:

• Conformationally distinct α-Syn polymorphs exhibit unique fluorescence lifetimes.
• FLIM successfully differentiated between various α-Syn amyloid structures.
• α-Syn polymorphs were processed differently by cellular clearance mechanisms, generating more potent seeding species.

Conclusions:

• FLIM is a valuable tool for distinguishing amyloid structures and observing cellular remodeling of amyloids.
• Differential processing of α-Syn polymorphs can lead to highly seeding competent degradation products.
• Accumulation of these products may explain accelerated disease progression in certain synucleinopathy patients.