title: Phosphorylation of tau protein in tau-transfected 3T3 cells.
abstract: The tau protein of Alzheimer paired helical filaments (PHFs) is aberrantly phosphorylated, as evidenced by its reactivity with several phosphate-dependent antibodies. We sought to identify whether this unusual phosphorylation state exists in tau expressed by transfected NIH 3T3 fibroblasts. Immunoblot analysis of cell clones transfected with constructs for either the 3-repeat or 4-repeat isoforms of tau revealed two tau bands, with the lower band migrating with unmodified tau in each case. Antibodies T3P and tau-1 were used to probe these bands, as they also react with PHF-tau in a phosphate-dependent manner. The epitopes for both antibodies were phosphorylated in both tau isoforms. Only the upper band was phosphorylated at the T3P site whereas phosphorylation at the tau-1 site was not always associated with a shift of tau mobility on gels. Tau in both bands was soluble, in contrast to PHF-tau, and was competent to bind to exogenously added bovine microtubules. Colchicine treatment of the cells resulted in an inhibition of phosphorylation at both sites, through an unknown mechanism. In conclusion human tau expressed in 3T3 cells was phosphorylated at the T3P and tau-1 sites as is PHF-tau, although no PHFs formed and the phosphorylated tau was competent to bind to microtubules.
authors.affiliation: ICI Pharmaceuticals Group, ICI Americas, Wilmington, DE 19897.
authors.forename: L A
authors.initials: LA
authors.lastname: Sygowski
authors.forename: A W
authors.initials: AW
authors.lastname: Fieles
authors.forename: M M
authors.initials: MM
authors.lastname: Lo
authors.forename: C W
authors.initials: CW
authors.lastname: Scott
authors.forename: C B
authors.initials: CB
authors.lastname: Caputo
mesh_terms: D016475:3T3 Cells / Q000378:metabolism*; D000818:Animals; D003078:Colchicine / Q000494:pharmacology; D051379:Mice; D008870:Microtubules / Q000187:drug effects / Q000378:metabolism; D010766:Phosphorylation; D011485:Protein Binding; D011499:Protein Processing, Post-Translational; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D014162:Transfection; D014404:Tubulin / Q000378:metabolism; D016875:tau Proteins / Q000378:metabolism*
chemical_list: D011993:Recombinant Fusion Proteins; D014404:Tubulin; D016875:tau Proteins; D003078:Colchicine