The binding and phosphorylation of Thr231 is critical for Tau's hyperphosphorylation and functional regulation by glycogen synthase kinase 3beta.
Neuropathological hallmarks of Alzheimer's disease are extracellular senile plaques and intracellular neurofibrillary lesions. The neurofibrillary lesions mainly consist of the hyperphosphorylated microtubule-associated protein Tau predominantly expressed in the axon of CNS neurons. Hyperphosphorylation of Tau negatively affects its binding to tubulin and decreases the capacity to promote microtubule assembly. Among a number of proline-directed kinases capable of phosphorylating paired helical filament-Tau, glycogen synthase kinase 3beta (GSK3beta) was first identified as a Tau protein kinase I and has been demonstrated to phosphorylate Tau both in vivo and in vitro. However, the phosphorylation mechanism of Tau by GSK3beta remained unclear. In this study, we show that the T231 is the primary phosphorylation site for GSK3beta and the Tau227-237 (AVVRTPPKSPS) derived from Tau containing T231P232 motif is identified as the GSK3beta binding site with high affinity of a Kd value 0.82 +/- 0.16 mumol/L. Our results suggest that direct binding and phosphorylation of T231P232 motif by GSK3beta induces conformational change of Tau and consequentially alters the inhibitory activity of its N-terminus that allows the phosphorylation of C-terminus of Tau by GSK3beta. Furthermore, hyperphosphorylation reduces Tau's ability to promote tubulin assembly and to form bundles in N18 cells. T231A mutant completely abolishes Tau phosphorylation by GSK3beta and retains the ability to promote tubulin polymerization and bundle formation. Taken together, these results suggest that phosphorylation of T231 by GSK3beta may play an important role in Tau's hyperphosphorylation and functional regulation.
Department of Biological Sciences, National Sun Yat-Sen University, Kaohsiung, Taiwan.
Yuh-Te
YT
Lin
Jiin-Tsuey
JT
Cheng
Li-Ching
LC
Liang
Chiung-Yuan
CY
Ko
Yuk-Keung
YK
Lo
Pei-Jung
PJ
Lu
D001665:Binding Sites; D015153:Blotting, Western; D002460:Cell Line; D002942:Circular Dichroism; D005455:Fluorescent Antibody Technique; D038362:Glycogen Synthase Kinase 3 / Q000378:metabolism* / Q000502:physiology; D006801:Humans; D047468:Immunoprecipitation; D008027:Light; D016874:Neurofibrillary Tangles / Q000235:genetics / Q000473:pathology; D010766:Phosphorylation; D010957:Plasmids / Q000235:genetics; D011485:Protein Binding; D011487:Protein Conformation; D012542:Scattering, Radiation; D013912:Threonine / Q000378:metabolism*; D013917:Thrombin / Q000378:metabolism; D014162:Transfection; D014404:Tubulin / Q000096:biosynthesis; D016875:tau Proteins / Q000378:metabolism / Q000502:physiology*
D014404:Tubulin; D016875:tau Proteins; D013912:Threonine; D038362:Glycogen Synthase Kinase 3; D013917:Thrombin