Yaqoob, Hamid (2004) STUDIES ON MECHANISM OF TAU PROTEIN HYPERPHOSPHORYLATION IN ALZHEIMER'S DISEASE. PhD thesis, University of Agriculture, Faisalabad.
In Alzheimers disease (AD) microtubule-associated protein tau becomes hyperphosphorylated and aggregates into paired helical filaments. Abnormal phosphorylation has been suggested to cause the loss of tau function, microtubule instability, and neurodegeneration in AD brain. As the GSK3 and 14-3-3c are associated with tau in brain and are considered to be involved in tau hyperphophorylation, so it is necessary to understand their iteraction in brain. To investigate the biochemical nature of microtubule associated GSK3,14-3-3c and to study its interaction with tau, the microtubules was dissociated by cold incubationand removed tubulin from the MAP fraction containing GSK3 and 14-3-3c by phosphocellulose chromatography. When the MAP fraction was subjected to an FPLC gel filtration analysis, GSK3 and 14-3-3c eluted as a ~400 kDa complex. 14-3-3c and the tau phosphorylation complex in the microtubule fraction cannot be separated from each other by Phosphocellulose, gel filtration, and Mono S chromatographies. In vitro, 14-3-3c binds to tau and GSK3. When the ~400-kDa comlex was chromatographed through an anti- GSK3 immunoaffinity column, tau co-eluted with GSK3. Similarly, taud3 co-immunoprecipitated with each other from fractions containing the ~400-kDa complex In vitro, GSK3 bound to R-tau. These data indicate that GSK3, 14-3-3c and tau ae the components of a ~400-kDa microtubule-associated complex. In this study it is demonstrated that GSK3 binds to microtubules through tau and also determined that GSK3 binds to the N-terminal projection domain of tau. Since the microtubule-binding region of tau does not overlap with the N-terminatlprojection domain, it is likely that tau can simultaneously bind to microtubules and GSK3 in vitro and perhaps in vivo. It was found that tau within brain extract and 400-kDa complex is heterogeneous by using antibodies Tau-1, 12E8, PHF-1, and AD-2 but only Tau-1 cross reacts with the tau that elutes from the anti- GSK3 immunoaffinity column. These data indicate that tau isoforms within GSK3 tau complex are not phosphorylated on Ser198, Ser199, Ser202, Ser298, Ser400, and Ser404 and suggest that GSK3 bound tau isoforms may be in nonphosphorylated states. However in vivo, tau is also phosphorylated on several other sites for which antibodies are not available. Thus, it concludes that within the GSK3 tau complex, tau isoforms are not phosphorylated on above indicated sites. The presence of tau not phosphorylated on Ser198, Ser199, Ser202, Ser235, Ser398, and Ser404 within the tau. GSK3 complex suggests that GSK3 tau interaction may be regulated by phosphorylation of tau on these sites. Since Cdk5 phosphorylates Ser199, Ser202, Ser235, Ser398, and Ser404 in vitro. GSK3 binds to R-tau but not Cdk5-phosphorylated R-tau. Previous studies have shown that Cdk5 is also a component of neuronal microtubules, binds to tau, and associates with microtubules via tau in a manner similar to GSK3. Taken together, these observations suggest that GSK3 tau binding may be regulated by phosphorylation of tau by Cdk5. This in turn may suggest that Cdk5 may not only prime tau for GSK3 action but may also regulate tau- GSK3 binding in vitro and possibly in vivio. GSK3-tau and tau-microtubule interactions may therefore be regulated by phosphorylation on the same tau sites. It is possible that, in the brain, nonphosphorylated tau simultaneously binds to GSK3 and microtubules, bridges GSK3 tomicrotubules, and stabilizes microtubule structure. Upon phosphorylation, tau dissociates from both microtubules and GSK3.
|Item Type:||Thesis (PhD)|
|Uncontrolled Keywords:||gsk3, microtubule, dna, protein, peptide concentrations, phosphorylation|
|Subjects:||Physical Sciences (f) > Chemistry(f2)|
|Deposited By:||Mr. Muhammad Asif|
|Deposited On:||11 Aug 2006|
|Last Modified:||04 Oct 2007 21:01|
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