Study of the impact of Tau phosphorylation on its internalization and processing in cultured neuronal cells

Several studies have already revealed the propagation of Tau protein aggregates in
different brain regions according to a stereotypical scheme that might explained the
successive onset of symptoms in Alzheimer's disease. In comparison, the implication
of soluble Tau proteins in this transmission is poorly reported in the literature even
though it seems that they constitute the most toxic species. In this study, we
hypothesized that the phosphorylation of the protein, which is implied in its
intracellular aggregation, could also play a role in this transmission. We applied two
recombinant monomeric Tau proteins presenting a distinct phosphorylation state in
the extracellular space of neuroblastoma cells (N2A) and of primary embryonic
cortical cells. Proteins were detected by immunocytochemistry through a his-Tag
included at their C-terminal extremity. We observed that the phosphorylation
increased protein internalization by up to 10 times within both cell types. However,
cellular distribution and processing of the phosphorylated protein were different in
N2A cells and in cortical neurons. In N2A cells, proteins were found mainly around
the nucleus up to 3 days after the period of incubation. Conversely, in cortical cells,
proteins were much more distributed in soma and neurites and disappeared 24h after
the end of the incubation period. This could be explained either by a digestion or an
exocytosis of the protein. These two hypotheses will be tested by further experiments
and the formation of intra-cytoplasmic aggregates will be studied in the presence of
phosphorylated or non-phosphorylated proteins. These results could help to better
understand the role of extracellular tau in the propagation of the disease and in the
associated synaptic transmission deficits.