Characterization of Oxidized Tau Protein Aggregation and Inhibition of Tau Aggregation via MAP2

<p>Neurofibrillary tangles (NFT) composed of the microtubule stabilizing protein Tau are a pathological hallmark of Alzheimer's disease (AD) and many other dementia related diseases specified as Tauopathies. Due to alternative mRNA splicing, intrinsically disordered Tau exists as six separate isoforms in the adult human brain where they are categorized by the lack or inclusion of the second repeat in the microtubule binding domain (3R or 4R respectively). Isoforms are subcategorized by the inclusion or lack thereof, one or two N-terminal inserts. Many studies show that Tau aggregates into amyloid fibrils and these fibrils can break into small oligomeric units known as seeds which are more efficient at recruiting native monomers in a template-assisted manner similar to a prion-like mechanism.</p> <p>A vast number of researchers consider fibrils/seeds to be toxic to neurons because they interact unfavorably with cellular membranes as well as cytosolic chaperones and compete with microtubules for native monomers resulting in the loss of their vital, native function. Once recruited, monomers are locked in an ultra-stable conformation unable to return to functional monomers. Preventing incorporation of monomers to preformed aggregates has been an intense focus of our lab and other research labs. MAP2 is another intrinsically disordered protein that aids microtubule stabilization and also possesses high sequence homology with Tau. This similarity provided the motivation to explore whether MAP2 has an affinity for Tau aggregates and possibly prevents Tau monomer addition. Here we show that MAP2Ctr (truncated 3R homolog) and MAP2Dtr (truncated 4R homolog) were able to inhibit truncated Tau monomer as well as full length cellular Tau monomer incorporation.</p> <p>Literature strongly suggests post translational modifications are a modulator of Tau aggregation in vivo. Previous studies inferred that oxidation may be a post translational modification that plays a role in the amyloidogenesis of Tau. However, conflicting reports left some uncertainty, stating that intramolecular oxidized Tau monomers were incapable of forming fibrils. Our lab decided to investigate this further because cellular oxidative stress has gained momentum as a central modulator of numerous neurodegenerative diseases including AD. Here we demonstrate that oxidized full length Tau forms semi-stable fibrils that seed other oxidized monomers regardless of oxidation method (peroxide or air oxidized). Oxidized Tau exists in the form of a compact monomer as it possesses an intramolecular disulfide linkage between cysteines at positions 291 and 322. Interestingly, Cryo-EM data has shown that the fibrils these monomers aggregate into contain an intermolecular disulfide bond throughout the fibril which implies a switch from intra- to intermolecular linkage of monomers within the fibril. Experiments in HEK293 cell line overexpressing hT40P301S-EYFP, a full-length disease mutant tau construct C-terminally tagged with an Enhanced Yellow Fluorescence Protein, show air oxidized aggregates seed intracellular monomers and possess similar properties to the original in vitro aggregates. This data supports the notion that oxidized Tau may have a physiological role whether that be a preventative or accelerating aggregation mechanism.</p>

Name thesis:degree_name

Ph.D.

Level thesis:degree_level

Dissertation

Year dc:date.available

2024

Author dc:creator

• Krzesinski, Brad

Contributors dc:contributor

• Martin Margittai
• Michelle Knowles
• Todd Wells
• Allegra Aron

• Alzheimer's disease (AD)
• Neurofibrillary tangles (NFT)
• Tau protein
• Taupathies
• Cellular biology
• Biochemistry
• Biochemistry, Biophysics, and Structural Biology
• Cell and Developmental Biology
• Cell Biology
• Life Sciences

Statement dc:rights

• <p>Copyright is held by the author. User is responsible for all copyright compliance.</p>

Language dc:language

English (eng)

Repository record dc:identifier

https://digitalcommons.du.edu/etd/2390

OAI identifier oai:identifier

oai:digitalcommons.du.edu:etd-3378