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Updated: May 23, 2025
In Vitro Aggregation Assays Using Hyperphosphorylated Tau Protein
Published on: January 2, 2015
Polyamination with spermidine enhances pathogenic tau conformations while reducing filamentous aggregate formation in
Mohammed Alhadidy1,2,3, Rebecca Mueller1,2, Jared Lamp1,4
1Department of Translational Neuroscience, College of Human Medicine, Michigan State University, Grand Rapids, MI, U.S.A.
Abstract:
Tau is subject to a broad range of post-translational modifications (PTMs) that regulate its biological activity in health and disease, including microtubule (MT) dynamics, aggregation, and adoption of pathogenic conformations. The most studied PTMs of tau are phosphorylation and acetylation; however, the salience of other PTMs is not fully explored. Tissue transglutaminase (TG) is an enzyme whose activity is elevated in Alzheimer's disease (AD). TG action on tau may lead to intramolecular and intermolecular cross-linking along with the incorporation of cationic polyamines (e.g., spermidine [SPD]) onto glutamine residues (Q). Even though SPD levels are significantly elevated in AD, the effects of SPD polyamination on tau biology have yet to be examined. In this work, we describe a method to produce recombinant SPD-modified tau where SPD modifications are mainly localized to Q residues within the N-terminus. MT binding and polymerization assays showed that SPD modification does not significantly alter tau's binding to MTs but increases MT polymerization kinetics. In addition, biochemical and biophysical assays showed that SPD polyamination of tau markedly reduces tau polymerization into filamentous and β-sheet-containing aggregates. On the other hand, SPD modification promotes the formation of pathogenic conformations (e.g., oligomerization and misfolding) by tau with or without inducing tau polymerization. Taken together, these data suggest that SPD polyamination of tau enhances its ability to polymerize MTs and favors the adoption of pathogenic tau conformations but not filamentous aggregates in vitro.
Insights
Spermidine (SPD) polyamination of tau enhances microtubule polymerization but reduces filamentous aggregates. This modification promotes pathogenic tau conformations, offering new insights into Alzheimer's disease mechanisms.
Area of Science:
- Neuroscience
- Biochemistry
- Molecular Biology
Background:
- Tau protein is central to microtubule dynamics and Alzheimer's disease (AD) pathogenesis.
- Post-translational modifications (PTMs) significantly influence tau's biological activity.
- Tissue transglutaminase (TG) and spermidine (SPD) levels are elevated in AD, suggesting a role in tau pathology.
Purpose of the Study:
- To investigate the effects of spermidine (SPD) polyamination on tau protein.
- To explore how SPD modification impacts tau's interaction with microtubules (MTs) and its aggregation propensity.
- To understand the role of SPD-modified tau in adopting pathogenic conformations relevant to AD.
Main Methods:
- Production of recombinant SPD-modified tau, with modifications primarily on N-terminal glutamine residues.
- Microtubule (MT) binding and polymerization assays to assess tau-MT interactions.
- Biochemical and biophysical assays to evaluate tau aggregation and conformational changes.
Main Results:
- SPD polyamination did not significantly alter tau's binding to MTs but enhanced MT polymerization kinetics.
- SPD-modified tau showed reduced polymerization into filamentous, beta-sheet-rich aggregates.
- SPD modification promoted the formation of pathogenic tau conformations, including oligomers and misfolded species.
Conclusions:
- SPD polyamination of tau enhances microtubule polymerization.
- This modification favors pathogenic tau conformations while inhibiting the formation of stable filamentous aggregates in vitro.
- Understanding SPD's role in tau modification may reveal novel therapeutic targets for Alzheimer's disease.
