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Updated: Mar 28, 2026
In Vitro Assay for Studying the Aggregation of Tau Protein and Drug Screening
Published on: November 20, 2018
Cysteines are critical determinants of spontaneous and seeded tau aggregation in cells
Parvathy Jayan1,2, Simran Rastogi1,3,2, Vaibhav Bommareddy1
1Center for Alzheimer's and Neurodegenerative Diseases, Peter O'Donnell Jr. Brain Institute, University of Texas Southwestern Medical Center, Dallas, TX 75390, USA.
Abstract:
The frontotemporal dementia-linked S320F mutation in the microtubule-associated protein tau promotes spontaneous aggregation, yet the structural basis of its amyloidogenesis remains unclear. Using cryo-electron microscopy, we determined the structure of an S320F295-330 tau fibril composed of parallel chains stabilized by the 306VQIVYK311 amyloid motif, with S320F buried in the fibril core and a C322-C322 disulfide linking two protofilaments. Although cysteines are dispensable for fibril formation by isolated peptide fragments in vitro, tau repeat domain constructs containing both C291 and C322 generate more potent seeds in cellular assays. In contrast, the C322S mutation suppresses spontaneous aggregation of S320F tau in cells, and combined C291S and C322S mutations inhibit seeded aggregation in both wild-type and S320F contexts. Systematic alanine mutagenesis coupled with seeding by tauopathy-derived material identifies cysteine residues as critical determinants of tau seeding, comparable in importance to core amyloid motifs. Together, these findings establish cysteines as central chemical regulators of tau aggregation and propagation.
Insights
The frontotemporal dementia-linked S320F tau mutation promotes aggregation. Cysteine residues, particularly C322, are critical chemical regulators of tau aggregation and seeding, impacting disease progression.
Area of Science:
- Neuroscience
- Structural Biology
- Biochemistry
Background:
- Frontotemporal dementia (FTD) is linked to mutations in the microtubule-associated protein tau.
- The S320F mutation in tau promotes spontaneous aggregation, but its structural basis is unclear.
- Understanding tau aggregation is crucial for FTD pathogenesis.
Purpose of the Study:
- To elucidate the structural basis of S320F tau aggregation and amyloidogenesis.
- To investigate the role of cysteine residues in tau aggregation and seeding.
Main Methods:
- Cryo-electron microscopy (cryo-EM) to determine fibril structure.
- Site-directed mutagenesis (S320F, C291S, C322S) to assess functional impact.
- In vitro and cellular assays to evaluate aggregation and seeding propensity.
Main Results:
- Determined the structure of S320F tau fibrils, revealing a core amyloid motif (VQIVYK) and a C322-C322 disulfide bond.
- Cysteine residues are critical for tau seeding, comparable to core amyloid motifs.
- C322S mutation suppresses S320F tau aggregation; combined C291S/C322S mutations inhibit aggregation in wild-type and S320F tau.
Conclusions:
- Cysteine residues are central chemical regulators of tau aggregation and propagation.
- Disulfide bonds involving cysteines play a significant role in tau fibril formation and seeding.
- Targeting cysteine-mediated regulation may offer therapeutic strategies for tauopathies.
