Macquarie University
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Investigating protein kinases, protein interactions and mechanisms of multi-site phosphorylation of the microtubule-associated protein tau

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posted on 2023-08-24, 03:49 authored by Kristie Stefanoska

Tauopathies refer to a diverse group of progressive neurodegenerative diseases characterized by the intracellular accumulation of hyperphosphorylated tau. The most common and extensively studied tauopathy is Alzheimer's disease (AD). At present there is no effective treatment for AD or related tauopathies and the disease process starts decades before clinical presentation. Thus, for future therapy it is important to identify the events that occur early during neurodegeneration. Tau is considered an intrinsically disordered protein, which is known to have interactions with other binding partners, in addition to microtubules. These interactions are critical for mediating tau's physiological, as well as pathological processes.

Humans are particularly susceptible to tau-mediated neurodegeneration as compared with other mammals. In paper I we report a primate-specific 11-amino acid motif in the N-terminal region (residues 18-28), which is not present in non-primate tau. This sequence differentially mediates interactions with neuronal proteins. Specifically, the binding of vesicle associated machinery, synaptic transmission, signaling and actin-binding proteins. We identified a novel interaction of tau with Annexin AS that is directly linked to this 11-amino acid motif. Thus, human tau has evolved to undergo species-specific interactions that depend on protein features inherent to primate tau.

Post-translational modifications, such as phosphorylation regulate tau complex formation and drive diverse cellular functions. Although hyperphosphorylation of tau is associated with disease, site-specific phosphorylation of tau has previously been shown to mediate protective effects. In paper II, we focus on proline-directed phosphorylation - the most abundant form and demonstrate tau phosphorylation at different sites is modulated by a complex interdependence hierarchy. Using experimental data on site interdependence, we identified distinct sites as phosphorylation 'Master' sites within N-terminal (TSO, T69, Tl ll) and Proline-rich regions (TlS3, T181, T20S). 'Master' sites determine the propagation of tau hyperphosphorylation at other sites. Changes to Master site phosphorylation could alter tau phosphorylation status. Furthermore, we define p38a MAP kinase with the strongest impact on Master site-dependent effects. Most notably, prior phosphorylation at 'Master' sites could alter p38a MAP kinase activity towards tau, resulting in extensive modification of multiple epitopes, while deletion of p38a in mice could reduce tau phosphorylation, suggesting p38a is a critical tau kinase.

p38 MAP kinases are involved in diverse signaling processes and their activity is regulated by upstream factors, feedback and crosstalk within signaling networks. In paper III, we addressed a role of neuronal p38a within the CNS in the modulation of anxiety-related behavioral responses in mice. Neuron-specific p38a knockout mice present with aberrant anxiety-related behavioural responses and increased activity of jun N-terminal kinase in the brain. Inhibition and CRISPR-mediated targeting of p38a in cells, results in low levels of active p38a and concomitant dysregulation of JNK activation. Furthermore, treatment with a JNK-specific inhibitor reverses abnormally high levels of JNK activity and subsequent anxiety-related responses in p38a knockout mice. Thus, we report that neuronal p38a negatively regulates JNK activity that is required for specific modulation of anxiety-related behaviour.


Table of Contents

Chapter 1: Literature Review -- Chapter 2: An N-terminal motif unique to primate tau enables differential protein-protein interactions -- Chapter 3: Systematic analysis of phosphorylation interdependence establishes 'Master' sites and kinases involved in tau hyperphosphorylation -- Chapter 4: Neuronal MAP kinase p38a inhibits c-Jun N-terminal kinase to modulate anxiety-related behaviour -- Chapter 5: General Discussion, Limitations, Future Directions and Conclusion -- 6. References -- 7. Appendix - Ethics Approval

Awarding Institution

Macquarie University

Degree Type

Thesis PhD

Department, Centre or School

Department of Biomedical Sciences

Year of Award


Principal Supervisor

Arne Ittner


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258 pages