Starkie, Dale Owen;
(2021)
Isoform-specific tau antibodies and degrabodies to study 4R tau function in MAPT mutant iPSC-derived neurons.
Doctoral thesis (Ph.D), UCL (University College London).
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Abstract
Hyperphosphorylated, insoluble aggregates of the microtubule-associated protein tau are a pathological hallmark of a group of clinically diverse diseases termed the tauopathies. There are six protein isoforms of tau protein separated into two groups; 3-repeat (3R) and 4-repeat (4R) tau, based on the number of repeats within the microtubule-binding region. Within neurons under normal healthy conditions, the ratio of expression of 3R and 4R tau is equal. However, in several tauopathies there is an imbalance of 3R and 4R tau, suggesting a disruption in tau splicing may be associated with disease. This is observed in Frontotemporal dementia and parkinsonism linked to chromosome-17, where intronic and splice-site mutations in MAPT increase the relative amount of 4R tau and cause early-onset neurodegeneration. This altered tau splicing has been recapitulated in induced pluripotent stem cell (iPSC) derived neurons, which recapitulate increased 4R tau levels along with other phenotypes including hyperpolarised mitochondrial membranes and an associated increase in reactive oxygen species (ROS) production. This highly relevant to neurodegeneration as ROS-induced cell death is a key feature of many neurodegenerative diseases. The mechanisms by which increased 4R results in neuronal dysfunction and neurodegeneration are not fully understood, and progress has been limited in part by a lack of suitable tools to investigate tau isoform imbalance. In this project, I generated and characterised novel 3R and 4R tau specific antibodies. I then used these antibodies to develop a multiplexed flow cytometry assay to show a correlation between 4R tau levels and mitochondrial membrane polarisation in iPSC-derived neurons containing the 10+16 MAPT splice mutation. Finally, I generated a 4R tau specific degrabody (target degrading intracellular antibody). I demonstrated this causes the specific degradation of 4R tau to achieve phenotypic restoration of non-mutant levels of mitochondrial membrane polarisation within the mutant neurons.
| Type: | Thesis (Doctoral) |
|---|---|
| Qualification: | Ph.D |
| Title: | Isoform-specific tau antibodies and degrabodies to study 4R tau function in MAPT mutant iPSC-derived neurons |
| Event: | UCL |
| Open access status: | An open access version is available from UCL Discovery |
| Language: | English |
| Additional information: | Copyright © The Author [year]. Original content in this thesis is licensed under the terms of the Creative Commons Attribution-NonCommercial 4.0 International (CC BY-NC 4.0) Licence (https://creativecommons.org/licenses/by-nc/4.0/). Any third-party copyright material present remains the property of its respective owner(s) and is licensed under its existing terms. Access may initially be restricted at the author’s request. |
| UCL classification: | UCL UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Brain Sciences UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Brain Sciences > UCL Queen Square Institute of Neurology |
| URI: | https://discovery.ucl.ac.uk/id/eprint/10132210 |
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