City, recent benefits have shed light on their possible interactions and
City, current benefits have shed light on their probable interactions and synergistic effects during AD progression. For instance, Tau-deficient mice are less susceptible to Atoxicity than control mice (Roberson et al., 2007). Current benefits have shown that AMPK is often a potent Tau kinase (Thornton et al., 2011). As a way to reconstitute a biochemical pathway triggering AMPK activation, we expressed a GFP-tagged version of Tau and AMPK in HeLa cells, which are naturally deficient for LKB1 (Hawley et al., 2003). In this model, AMPK might be specifically activated by reintroducing its upstream activator LKB1. This experiment confirmed that AMPK phosphorylates the wellcharacterized KxGS motif on Tau Serine 262 (S262) residue (Figure 5A). When coexpressed in cell lines, both LKB1 (coexpressed with its coactivator STRAD) and CAMKK2 are potent activators of AMPK, though we observed that CAMKK2 was substantially more potent in phosphorylating AMPK on T172 than LKB1 or CAMKK1 (Figure 5B). Moreover, direct activation of AMPK using the AMP analog AICAR triggered a ALK6 custom synthesis dose-dependent enhance of Tau phosphorylation of S262 in cortical neurons (Figures 5C, 5D, and S4), a treatment that induces a dose-dependent reduction in spine density (Figures 1N and 1O). The microtubule-associated protein Tau is phosphorylated in many web-sites (Mandelkow and Mandelkow, 2012), and analysis of six well-characterized phosphorylation web pages revealed that following 24 hr treatment with AICAR, phosphorylation of Tau on S262 is drastically increased inside a dose-dependent manner but that other sites are either unchanged (for instance, the other KxGS motif on S356, as well as S396, S422) or decreased (S202T205, S404) (Figures S4A and S4B). This observation suggests that S262 is definitely an crucial target of AMPK, and phosphorylation of this web page may underlie AMPKCYP51 Storage & Stability induced spine loss. Stopping Tau Phosphorylation on S262 Protects Hippocampal Neurons in the Synaptotoxic Effects of A42 Oligomers In Vitro as well as the Dendritic Spine Loss Observed inside the APPSWE,IND Mouse Model In Vivo Previous studies in Drosophila recommended that overexpression of AMPK-related member PAR-1MARK2 induced neurotoxicity by means of phosphorylation of Tau inside the microtubulebinding domains on S262 and S356 and that phosphorylation of these sites played an initiator part inside the pathogenic phosphorylation procedure of Tau (Nishimura et al., 2004). Provided the value of phosphorylation of S262 as a “priming” web page (Biernat et al., 1993) along with the recent implication of Tau within the synaptotoxic effects of A42 oligomers (Ittner et al.,Neuron. Author manuscript; offered in PMC 2014 April ten.NIH-PA Author Manuscript NIH-PA Author Manuscript NIH-PA Author ManuscriptMairet-Coello et al.Page2010; Roberson et al., 2007), we wanted to test if expression of a type of Tau that cannot be phosphorylated on S262 could exert a protective effect within the context of A42 oligomerinduced synaptotoxicity in cultured hippocampal neurons. Expression of Tau S262A abolished the loss of spines induced by A42 oligomers (Figures 5EH), while its expression in control neurons did not have any effect on spine density. By contrast, expression of Tau WT or even a phospho-mimetic version of Tau on S262 (Tau S262E) resulted in spine loss in manage condition, plus the WT form of Tau was unable to prevent the synaptotoxic effects of A42 oligomers. Ultimately, the nonphosphorylatable type of Tau on S356 (S356A) displayed comparable protective effects as Tau S262A mutant.