Sarkosyl differentially solubilizes patient-derived alpha-synuclein fibril strains

Area of Science:

• Neuroscience
• Biochemistry
• Pathology

Background:

• Insoluble alpha-synuclein (αSyn) filaments are key pathological hallmarks in Parkinson's disease (PD) and Multiple System Atrophy (MSA).
• Sarkosyl extraction is a standard method for isolating αSyn filaments for structural analysis, but its impact on different patient-derived strains is unclear.

Purpose of the Study:

• To investigate whether αSyn filament strains from PD and MSA patients exhibit differential stability when extracted with sarkosyl.
• To determine if differential stability confounds the interpretation of structural variations in patient-derived αSyn filaments.

Main Methods:

• Compared sarkosyl extraction stability of cerebrospinal fluid-derived αSyn filaments from four PD and four MSA patients using sedimentation and immunoassays.
• Assessed seeding competence and strain characteristics of sarkosyl-soluble fractions via seed amplification assay (SAA) and Thioflavin T (ThT) fluorescence.

Main Results:

• PD-derived αSyn filaments demonstrated lower resistance to sarkosyl extraction compared to MSA-derived filaments after freezing and sonication.
• A greater release of monomers from PD filaments was observed, but the soluble fractions from both PD and MSA contained seeding-competent aggregates with preserved disease-specific characteristics.

Conclusions:

• Sarkosyl extraction differentially destabilizes patient-derived αSyn filament strains, potentially limiting the comprehensive understanding of αSyn filament structures obtained through cryo-electron microscopy (cryo-EM).
• Development of gentler extraction protocols is recommended to better preserve the integrity of diverse αSyn filament strains for accurate structural and pathological studies.