14-3-3 connects glycogen synthase kinase-3 beta to tau within a brain microtubule-associated tau phosphorylation complex.

In a recent study, we reported that in bovine brain extract, glycogen synthase kinase-3beta and tau are parts of an approximately 400-500 kDa microtubule-associated tau phosphorylation complex (Sun, W., Qureshi, H. Y., Cafferty, P. W., Sobue, K., Agarwal-Mawal, A., Neufield, K. D., and Paudel, H. K. (2002) J. Biol. Chem. 277, 11933-11940). In this study, we find that when purified brain microtubules are subjected to Superose 12 gel filtration column chromatography, the dimeric scaffold protein 14-3-3 zeta co-elutes with the tau phosphorylation complex components tau and GSK3 beta. From gel filtration fractions containing the tau phosphorylation complex, 14-3-3 zeta, GSK3 beta, and tau co-immunoprecipitate with each other. From extracts of bovine brain, COS-7 cells, and HEK-293 cells transfected with GSK3 beta, 14-3-3 zeta co-precipitates with GSK3 beta, indicating that GSK3 beta binds to 14-3-3 zeta. From HEK-293 cells transfected with tau, GSK3 beta, and 14-3-3 zeta in different combinations, tau co-immunoprecipitates with GSK3 beta only in the presence of 14-3-3 zeta. In vitro, approximately 10-fold more tau binds to GSK3 beta in the presence of than in the absence of 14-3-3 zeta. In transfected HEK-293 cells, 14-3-3 zeta stimulates GSK3 beta-catalyzed tau phosphorylation in a dose-dependent manner. These data indicate that in brain, the 14-3-3 zeta dimer simultaneously binds and bridges tau and GSK3 beta and stimulates GSK3 beta-catalyzed tau phosphorylation.

Glycogen synthase kinase-3beta is complexed with tau protein in brain microtubules.

In Alzheimer's disease, microtubule-associated protein tau is hyperphosphorylated by an unknown mechanism and is aggregated into paired helical filaments. Hyperphosphorylation causes loss of tau function, microtubule instability, and neurodegeneration. Glycogen synthase kinase-3beta (GSK3beta) has been implicated in the phosphorylation of tau in normal and Alzheimer's disease brain. The molecular mechanism of GSK3beta-tau interaction has not been clarified. In this study, we find that when microtubules are disassembled, microtubule-associated GSK3beta dissociates from microtubules. From a gel filtration column, the dissociated GSK3beta elutes as an approximately 400-kDa complex. When fractions containing the approximately 400-kDa complex are chromatographed through an anti-GSK3beta immunoaffinity column, tau co-elutes with GSK3beta. From fractions containing the approximately 400-kDa complex, both tau and GSK3beta co-immunoprecipitate with each other. GSK3beta binds to nonphosphorylated tau, and the GSK3beta-binding region is located within the N-terminal projection domain of tau. In vitro, GSK3beta associates with microtubules only in the presence of tau. From brain extract, approximately 6-fold more GSK3beta co-immunoprecipitates with tau than GSK3alpha. These data indicate that, in brain, GSK3beta is bound to tau within a approximately 400-kDa microtubule-associated complex, and GSK3beta associates with microtubules via tau.