Index | Round
Tables | Selected
Abstracts | News
by Luc Buee
20 July 1998. Three abstracts presented during this session, as well as those by Vincent et al. (abstract 594) and Arendt et al. (abstract 595), indicate that cell cycle and mitotic mechanisms may be a key factor in the understanding of tau phosphorylation. Illenberger et al. (abstract 912) analyzed tau phosphorylation in LAN-5 neuroblastoma cells and CHO cells stably transfected with tau protein. In both cells non-synchronized, the phosphorylation pattern using 32P and 2D phosphopeptide mapping and sequencing was similar: phosphorylation of Ser-Pro motifs and Ser214 and 262. CHO cells were also blocked in anaphase with nocodazole. In this situation, tau proteins are free and do not bind to microtubules. They are highly phosphorylated, especially at Ser214. Finally, Ser214 phosphorylation by PKA prevents microtubules nucleation. However, AT100 is not found in mitotic cells (see abstract 914).
Stem cells are potential precursor cells for both neurons and glial cells. Tatebabayashi et al. (abstract 913) developed this approach. First, they characterized the cells: they are MAP2 negative (except for MAP2c), NF negative, GFAP negative, NSE positive and also tau-positive (except for Tau-1 epitope). Tau expression and phosphorylation increases in a dose-dependent manner with FGF2.
Zheng-Fishhöfer et al. (abstract 914) demonstrated that in vitro AT100 epitope generation needs the sequential phosphorylation of GSK3ß and PKA. In fact, they used two approaches, one way using tau constructs, kinase activity from rat brain and specific kinase inhibitors and the second way using activated kinases and tau constructs. They nicely showed that first a PHF-like conformation is required (that can be mimicked by heparin or RNA), second GSK3ß phosphorylation at AT8 sites and Thr212 and finally Ser214 pKA phosphorylation.
Mutations on the tau gene suggest that when tau proteins do not bind to microtubules they are free to aggregate (see news summary of Hereditary Fronto-Temporal Dementia and Pick's disease). However, it does not explain why they are hyperphosphorylated when aggregated into filaments in all neurodegenerative disorders (abstract 591). After tau release from microtubules, cell cycle mechanisms may be reactivated and lead to tau phosphorylation. It is part of the new challenge to understand neurofibrillary degeneration: What is the role of phosphorylation?
The two following abstracts may be useful for illuminating this question. One indicates that tau phosphorylation is also linked to the cholinergic system, and the other demonstrates that non-phosphorylated tau proteins may aggregate in to PHF.
Forlenza et al. (abstract 915) confirmed the study by Sadot et al. in Journal of Neurochemistry showing that muscarinic agonists reduce tau phosphorylation through PKC activation of m1 and m3 receptors. They demonstrated that this decrease in tau phosphorylation is mediated through GSK3 inhibition. In conclusion, 100 µM carbachol induces a decrease in tau phosphorylation and an increase in the formation of microtubules bundles.
Mandelkow et al. (abstract 916) reviewed their in vitro experiments on tau aggregation. One of the first step is the dimerization of tau isoforms through a disulfide bridge (Cys322). Presence of polyanions including RNA and glycosaminoglycans enhances tau assembly into PHF. Effects of glycosaminoglycans on tau aggregation into filaments were also described by Avila et al (abstract 619) and Goedert et al. (abstract 926). Moreover, N- and C-terminal truncation of tau isoforms further increase the rate of in vitro PHF formation. The role of truncation was also emphasized by the two next speakers, Novak (abstract 917) and Cattaneo (abstract 918).
Mitro et al. used the monoclonal antibody MN423 that recognizes tau isoforms cleaved at Glu 391 (-DHGAE) to demonstrate that truncated tau proteins are found within neurofibrillary tangles (abstract 917). Tau truncation seems to be an early event in tau pathology. Furthermore, a significant population of MN423-positive neurons was apoptotic. To further address this problem of apoptosis, Fasulo et al., from the same group, (abstract 918) used cell transfection to demonstrate that truncated tau isoform (residues 151-391) induce apoptosis in about 50% of COS cells. Interesting preliminary results seemed to indicate that induction of apoptosis may also lead to truncated tau proteins (abstracts 1120 and 1121).
Arawaka et al. (abstract 919) described new monoclonal antibodies that recognizes sequence boundaries of regions encoded by alternatively spliced exons. They developed specific antibodies against sequences encoded by exons 2, 3 and 10 but also against sequences at the boundaries between exons 1 and 4, 2 and 4, 9 and 11. All antibodies labeled NFT by immunohistochemistry. They are highly specific when tested on recombinant tau proteins. However, there was no data on phosphorylated tau proteins or on total brain homogenates. If these antibodies are still specific for hyperphosphorylated tau proteins, they may be useful tools for studying neurofibrillary degeneration.