Abstract: Tau, a natively unstructured protein, interacts with the microtubules (MTs) via its microtubule-binding (MTB) domain, located in the C-terminal half of the protein. The protein, when aggregated in Paired Helical Filaments (PHF) in Alzheimer's disease (AD)-affected neurons, is invariably hyperphosphorylated and potentially phosphorylated on sites unique to the disease. Nuclear magnetic Resonance (NMR) Spectroscopy was used to explore the different aspects of the Tau normal and pathological functions but proved challenging, because the protein contains 441 amino acids and has poor signal dispersion. The interaction of Tau with taxol-stabilized microtubules was first studied using NMR experiments to obtain a per residue information. Tau can be forced to form AD like aggregates through the addition of poly-anions such as heparin. High Resolution Magic Angle Spinning NMR and solution NMR were used to further characterize these fibers. We next ask how the heparin can promote the fiber formation. The interaction with Tau was studied by using small heparin fragments of well-defined size, at 20 ⁰C, in conditions where no aggregation occur. Still, the relevance of heparin as the intracellular aggregating agent is far from clear. We have set out to dissect the phosphorylation pattern of Tau in order to understand better its role in the aggregation process and the microtubule binding. Our current knowledge on the functional consequences of specific phosphorylation patterns is still limited, mainly because producing and assessing quantitatively phosphorylated Tau samples is far from straightforward. We have applied NMR spectroscopy to identify and quantify in a single experiment the phosphorylation pattern of Tau, after in vitro phosphorylation, for example with the PKA kinase. Once the pattern of phosphorylation is characterized by NMR, the phosphorylated Tau can be use for functional assays like aggregation or microtubules interaction.We present here our results on Tau samples that have been in vitro phosphorylated by the Cyclin dependent kinase 2 (CDK2)/Cyclin A3 (CycA3) kinase complex. The impact of these phosphorylations on the local structuration of Tau was also analyzed. Finally, NMR was use to explore other aspects of the regulation of Tau function by phosphorylation. This includes characterisation of its interaction with phospho-dependent protein partners, such as the prolyl cis/trans isomerase PIN1 and dephosphorylation by the PP2A phosphatase.

Brief Biography of the Speaker:
As a PhD student, I spent 2 years in the European Molecular Biology Laboratory (EMBL), Grenoble Outstation in France, learning about Protein Biochemistry. I received my PhD in Agronomy and Biological Engineering from the Agronomical Faculty of Gembloux (FUSAGx) in Belgium in 1997. As Belgium National Foundation Fellow, I next joined the Department of Plant System Biology in Gent University/ Vlaams Institute for Biotechnology in Belgium, in the lab of Prof D. Inze and Prof. M. Van Montagu. During this time, I studied cell cycle regulatory proteins from Arabidopsis thaliana. I did functional characterizations but also worked on their structural characterization using NMR spectroscopy. In 2001, I joined the French National Science Center (CNRS) and started to work in the NMR group directed by Prof. G. Lippens in Lille, France. I obtained my habilitation in 2003 from the Science and Technology University in Lille, France. My main interest is ever since the regulation of Tau function by phosphorylation and the recognition of these phosphorylated sites by protein partners and various enzymes.