Phosphorylation at serine 214 correlates with tau seeding activity in an age-dependent manner in two mouse models for tauopathies and is required for tau transsynaptic propagation

Background Tau aggregation and propagation are hallmark features of Alzheimer's disease and related tauopathies. The molecular identity and post-translational modifications that contribute to tau seeding activity remain incompletely understood. Objective To characterize the temporal dynamics of tau seeding activity and identify specific phosphorylated tau species associated with tau propagation in vivo. Methods We profiled tau seeding activity using a FRET-based biosensor cell line and correlated it with the abundance of phospho- and conformational tau species in two transgenic mouse models of tauopathy (P301S-1N4R and P301S-0N4R). Immunohistochemistry and subcellular fractionation were used to examine the spatial distribution of tau species. Functional relevance of serine 214 phosphorylation was assessed via phospho-dead mutants in Drosophila and primary neuronal cultures using a microfluidic platform. Results Tau seeding activity was detected as early as 2 months and increased with age in both mouse models, preceding AT8-positive neurofibrillary tangles. Phosphorylation at serine 214 (pTau-Ser214) positively correlated with tau seeding in both models and was observed earlier and in distinct histological compartments compared to AT8. pTau-Ser214, but not AT8, localized to the synaptic compartment. Mutation of serine 214 to alanine reduced tau propagation in primary neurons and seeding activity in Drosophila , without affecting total tau levels. Conclusions pTau-Ser214 marks an early, synaptically enriched tau species that correlates with seeding activity and promotes transsynaptic propagation. These findings highlight the functional diversity of tau species and support pTau-Ser214 as a potential biomarker and therapeutic target for early-stage tauopathy.