Pseudophosphorylation of Tau at distinct epitopes or the presence of the P301L mutation targets the microtubule-associated protein Tau to dendritic spines

Abstract

Alzheimer's disease is characterized by the accumulation of amyloid-β (Aβ) and Tau in the brain. In mature neurons, Tau is concentrated in the axon and found at lower levels in the dendrite where it is required for targeting Fyn to the spines. Here Fyn mediates Aβ toxicity, which is vastly abrogated when Tau is either deleted or a truncated form of Tau (Tau(1-255)) is co-expressed. Interestingly, MAP2, a microtubule-binding protein with mainly dendritic localization that shares Fyn-binding motifs with Tau, does not mediate Aβ's synaptic toxicity in the absence of Tau. Here we show in hippocampal neurons that endogenous Tau enters the entire spine, albeit at low levels, whereas MAP2 only enters its neck or is restricted to the dendritic shaft. Based on an extensive mutagenesis study, we also reveal that the spine localization of Tau is facilitated by deletion of the microtubule-binding repeat domain. When distinct phosphorylation sites (AT180-T231/S235, 12E8-S262/S356, PHF1-S396/S404) were pseudophosphorylated (with glutamic acid, using alanine replacements as controls), Tau targeting to spines was markedly increased, whereas the pseudophosphorylation of the late phospho-epitope S422 had no effect. In determining the role physiological Fyn has in the spine localization of Tau, we found that neither were endogenous Tau levels reduced in Fyn knockout compared with wild-type synaptosomal brain fractions nor was the spine localization of over-expressed pseudophosphorylated or P301L Tau. This demonstrates that although Fyn targeting to the spine is Tau dependent, elevated levels of phosphorylated Tau or P301L Tau can enter the spine in a Fyn-independent manner.

Can Molecular Gradients Wire the Brain?

“…Seven µm-thick brain sections were cut with a microtome in the frontal or sagittal plane and mounted on silane-coated slides. Brains of 7-8, 11-12, and 16 month-old Tau58-2/B mice were analyzed for the presence of pathological tau species (Xia et al, 2015) (Fig 1). The following anti-tau antibodies were used: pSer235…”

The molecular mechanism of the Tau-Fyn interaction and dendritic targeting in neurons

“…On the other hand, P301L Tau has been shown to accumulate in an Aβ-dependent manner in spines in vivo [17]. Similarly, over-expression of P301L Tau was also found to cause increased spine localization [115]. Together, these findings demonstrate that Tau localization to spines is tightly regulated.…”

“…This PhD thesis resulted in one review [28] and three original research articles [93,115,130]. The first publication analysed a transgenic mouse line generated by me as part of this thesis that expresses a constitutively active form of Fyn that resulted in premature lethality, hyperactivity and an aberrant phosphorylation of the Fyn substrate Tau [93].…”

“…However, the two studies differ in several aspects: they transfected 0N4R and not 2N4R Tau, and they transfected at DIV7-10 and analysed the neurons at DIV21 [115,127], whereas we transfected the neurons only at DIV18 and analysed them at DIV19/20. Together, our findings reveal that even under physiological conditions, Tau is targeted to dendritic spines, and that over-expression of P301L mutant Tau causes a decrease in spine density.…”

“…Tau localization in the dendrite has previously been found to extend to spines; however, levels of endogenous Tau were reported to be low, unless Tau was over-expressed, causing it to enter spines in a phosphorylation-dependent manner [115,127]. To assess spine localization of Tau, we co- expressed EGFP-tagged Lifeact, a versatile marker to visualize F-actin [128], as a spine marker, due to its co-localization with RFP-tagged PSD95 in DIV18 WT neurons (Fig.…”