Abnormal hyperphosphorylation of tau appears to be crucial in neurofibrillary degeneration in Alzheimer's disease (AD). Previous studies suggest that a down-regulation of protein phosphatase 2A (PP2A), the major tau phosphatase in human brain, contributes to tau hyperphosphorylation in AD. However, the effects of PP2A down-regulation on site-specific tau hyperphosphorylation is not well understood. In the present study, we showed that PP2A dephosphorylated tau at several phosphorylation sites with different efficiencies. Among the sites studied, Thr205, Thr212, Ser214, and Ser262 were the most favorable sites, and Ser199 and Ser404 were the least favorable sites for PP2A in vitro. Inhibition of PP2A with okadaic acid in metabolically active rat brain slices caused inhibition of glycogen synthase kinase-3beta (GSK-3beta) via an increase in its phosphorylation at Ser9. GSK-3beta phosphorylated tau at many sites, with Ser199, Thr205, and Ser396 being the most favorable sites in cells. The overall alterations in tau phosphorylation induced by PP2A inhibition were the result of the combined effects of both reduced tau dephosphorylation due to PP2A inhibition directly and reduced phosphorylation by GSK-3beta due to its inhibition. Because the impacts of tau phosphorylation on its biological activity and on neurofibrillary degeneration are site-specific, this study provides a new insight into the role of PP2A down-regulation in neurofibrillary degeneration in AD.

Abnormal hyperphosphorylation of tau is pivotally involved in the pathogenesis of Alzheimer's disease (AD) and related tauopathies. Glycogen synthase kinase 3β (GSK-3β) is a primary tau kinase that is most implicated in tau pathology in AD. However, the exact molecular nature of GSK-3β involved in AD is unclear. In the present study, we found that GSK-3β was truncated at C-terminus and correlated with over-activation of calpain I in AD brain. Truncation of GSK-3β was positively correlated with tau hyperphosphorylation, tangles score and Braak stage in human brain. Calpain I proteolyzed GSK-3β in vitro at C-terminus, leading to an increase of its kinase activity, but keeping its characteristic to preferentially phosphorylate the protein kinase A-primed tau. Excitotoxicity induced by kainic acid (KA) caused GSK-3β truncation at C-terminus and hyperphosphorylation of tau in mouse brain. Inhibition of calpain prevented the KA-induced changes. These findings suggest that truncation of GSK-3β by Ca2+/calpain I markedly increases its activity and involvement of this mechanism probably is responsible for up-regulation of GSK-3β and consequent abnormal hyperphosphorylation of tau and neurofibrillary degeneration in AD.

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Hyperphosphorylation and deposition of tau into neurofibrillary tangles is a hallmark of Alzheimer disease (AD).Alternative splicing of tau exon 10 generates tau isoforms containing three or four microtubule binding repeats (3R-tau and 4R-tau), which are equally expressed in adult human brain. Dysregulation of exon 10 causes neurofibrillary degeneration. Here, we report that cyclic AMP-dependent protein kinase, PKA, phosphorylates splicing factor SRSF1, modulates its binding to tau pre-mRNA, and promotes tau exon 10 inclusion in cultured cells and in vivo in rat brain. PKA-C␣, but not PKA-C␤, interacts with SRSF1 and elevates SRSF1-mediated tau exon 10 inclusion. In AD brain, the decreased level of PKA-C␣ correlates with the increased level of 3R-tau. These findings suggest that a downregulation of PKA dysregulates the alternative splicing of tau exon 10 and contributes to neurofibrillary degeneration in AD by causing an imbalance in 3R-tau and 4R-tau expression.Tau is a neuronal microtubule-associated protein, the function of which is to stimulate microtubule assembly and stabilize microtubules. Hyperphosphorylation of tau leads to its aggregation into neurofibrillary tangles, a hallmark of Alzheimer disease (AD) 2 and related neurodegenerative diseases called tauopathies (1-4). Adult human brain expresses six different tau isoforms from a single gene by alternative splicing of exons 2, 3, and 10 of its pre-mRNA (5). The exon 10 encodes the second microtubule binding repeat (6). Alternative splicing of exon 10 generates tau with three or four microtubule binding repeats (3R-tau or 4R-tau), which is under developmental and cell type-specific regulation. Only 3R-tau is expressed during embryogenesis, whereas 3R-tau and 4R-tau are expressed in approximately equal amounts in adult human brain (6, 7). Several mutations in tau gene result in either an increase or a decrease in 4R-tau expression and cause frontotemporal dementia with parkinsonism linked to chromosome 17 (FTDP-17), one of the tauopathies (8). Thus, alteration in the 3R-tau/ 4R-tau ratio is sufficient to trigger neurodegeneration in frontotemporal dementia and might also play a role in other neurodegenerative disorders such as Pick's disease, progressive nuclear palsy, or corticobasal degeneration in which the 3R-tau/4R-tau ratio is markedly altered (9 -12). Thus, the regulation of alternative splicing of human tau exon 10 has been of critical interest. However, results of studies of the alternative splicing of tau exon 10 in AD brain have been contradictory (13)(14)(15). Recent studies have shown that aggregation and deposition of 3R-tau may be associated with more advanced stages (16,17).Alternative splicing of tau exon 10 is regulated by several trans-acting factors, including serine-and arginine-rich (SR) proteins, and their phosphorylation (18 -24). Splicing factor 2/alternative splicing factor (ASF/SF2), now named SRSF1 (serine/arginine-rich splicing factor 1) (25), is a prototypical SR protein that participates in both constitutive and alternativ...