• mex3a
• neural aging
• TAU
• tauopathies
• xenopus
• zebrafish
• znf367

Neural aging is a universal inevitable gradual process characterized by cells and synapses loss, stem cell exhaustion, deregulation in protein-control pathways, and mitochondrial dysfunction. These alterations lead to cognitive decline, increased susceptibility to pathologies, impairment of adult neurogenesis. Insights on new genetic and molecular mechanisms involved in neural aging are required for determining prognostic and therapeutic actions against neurodegeneration.
To generate new tools to investigate the mechanisms underlying neural aging, I exploited Xenopus laevis and Danio rerio (zebrafish) as model systems. Hence, my PhD project focused on two main goals: i) the functional study of the age-related genes mex3a and znf367; ii) the generation of a new in vivo assay for studying human TAU behaviour associated with neurodegeneration.
First goal: I investigated the role of mex3a in embryonic neurogenesis performing gene gain and loss of function experiments in Xenopus embryos. In vertebrates, the expression of mex3a declines with age; my work indicates that mex3a has a key role in the maintenance of neural precursors proliferation during embryonic neurogenesis. These results pave the way for future studies aimed to clarify mex3a involvement in adult neurogenesis and neural aging.
I addressed the study of the znf367 gene using zebrafish as a model system. At first, I analysed the spatial and temporal expression pattern of znf367, which showed its localization in the embryonic nervous system and in neurogenic niches of adult fish suggesting an uninterrupted role through the lifespan for this gene. To study znf367 function, I initially performed a transient knock-down approach. Then, I generated a knock-out line by means of the CRISPR/CAS9 methodology to achieve a better understanding of znf367 function in embryonic and adult neurogenesis.
Second goal: to deepen our knowledge about molecular mechanisms related to pathological neural aging, I developed a new in vivo model to study the onset of tauopathies. Tauopathies are a heterogeneous group of neurodegenerative disorders characterized by the deposition of hyperphosphorylated TAU proteins with the formation of Neuro-Fibrillary Tangles (NFTs). To our knowledge, the mechanisms driving the pathological changes of TAU are not well elucidated and tauopathies, such as Alzheimer's disease, are diagnosed in advanced stages. Recent evidence has indicated that the increased fraction of soluble hyperphosphorylated TAU, compared to that bound to microtubules, triggers the early steps of aggregation. To date, no assays allow the detection in vivo of this imbalance hampering the identification of drugs blocking the earliest events in the pathological progression. In order to generate and validate a new functional assay for studying in vivo human TAU behaviour, I developed a stable transgenic zebrafish line expressing human normal TAU fused to a Conformational Sensitive TAU sensor (CST) in post-mitotic neurons. CST sensor, already described in vitro by our collaborators Dr. Cristina Di Primio and colleagues, is the first TAU-based sensor allowing the study of TAU conformational shift from physiological to pathological. Hence, I exploited tol2-transposon mediated system to generate a novel transgenic zebrafish line for in vivo high-throughput screenings of potential therapeutic compounds.
Understanding the function of the age-related genes during neural aging and revealing the mechanisms by which TAU detaches from the microtubules and aggregates should contribute to the identification of new interventions to restore efficient brain function both in physiological and pathological conditions.