Modifications post-traductionnelles et leur rôle dans l'agrégation de Tau dans la maladie d'Alzheimer

Alzheimer's disease (AD) is a neurodegenerative disorder characterized by the accumulation of amyloid plaques and neurofibrillary tangles composed of hyperphosphorylated Tau protein. Tau aggregation plays a central role in the progression of tauopathies, a group of age-related neurodegenerative disorders, by disrupting microtubule stability and promoting the spread of Tau pathology through seeding mechanisms.Glycogen synthase kinase 3 beta (GSK3β) is one of the key kinases implicated in Tau hyperphosphorylation, and its dysregulation is closely associated with the pathogenesis of AD. This study explores the complex regulation of GSK3β activity through post-translational modifications, including phosphorylation and O-GlcNAcylation, and their impact on Tau function and aggregation. We found that Akt-mediated phosphorylation at Ser9 inhibits GSK3β activity on unprimed Tau substrates, while selectively restoring its activity on primed Tau, highlighting a differential regulatory mechanism. GSK3β is also extensively O-GlcNAcylated at multiple sites, including Ser9, which does not significantly alter kinase activity but reduces Ser9 phosphorylation by Akt, indicating a crosstalk between these modifications that may influence GSK3β function.Additionally, we investigated the functional and seeding properties of Tau fragments, R2Ct and R3Ct, which encompass key aggregation-prone regions and the PHF-1 epitope (S396, S400, S404). The R3Ct fragment demonstrated a greater loss of microtubule-stabilizing function and an increased propensity for toxic seeding compared to R2Ct, with these effects being exacerbated by GSK3β phosphorylation. To further understand the impact of specific phosphorylation events on Tau behavior, we developed a semi-synthetic strategy to produce homogeneous, site-specifically phosphorylated fulllength 2N4R Tau, including PHF1-3P Tau with three phosphorylation sites (pS396, pS400, pS404).Our results indicate that PHF1-3P Tau significantly impairs microtubule assembly and, while reducing heparin-induced aggregation, promotes the formation of long PHF-like fibrils via cross-seeding.Furthermore, we explored the use of nanobodies as potential therapeutic agents for inhibiting Tau aggregation. Nanobodies, which are single-domain antibodies derived from camelid species, offer unique advantages due to their small size, high stability, and ability to bind to specific epitopes with high affinity. By targeting key regions within Tau's intrinsically disordered domains, nanobodies can effectively inhibit aggregation and prevent the formation of neurofibrillary tangles. In our studies, we identified nanobodies that specifically bind to the PHF6 and PHF6* motifs within Tau, critical sites for fibril formation, and demonstrated that these nanobodies can reduce Tau aggregation in vitro.This research provides critical insights into the regulation of GSK3β by phosphorylation and OGlcNAcylation, the differential effects of Tau phosphorylation on seeding and aggregation, and the use of semi-synthetic approaches to study complex PTM interactions on Tau. Additionally, the identification of nanobodies as inhibitors of Tau aggregation opens new avenues for therapeutic intervention in AD.These findings enhance our understanding of the molecular mechanisms driving Tau pathology in AD and highlight potential therapeutic targets for mitigating Tau aggregation and progression in tauopathies.