Molecular Mechanisms of Tau Binding to Microtubule and its Role in Microtubule Dynamics in Live Cells

Abstract

Despite extensive studies, molecular mechanisms of Tau binding to microtubule (MT) and its consequences on MT stability still remain misunderstood. It is especially true in cells where spatio-temporal distribution of Tau-MT interactions is unknown. Using Förster Resonance Energy Transfer (FRET), we showed that Tau-MT interaction was distributed along MTs in periodic hotspots of high and low FRET intensities. Fluorescence Recovery After Photobleaching (FRAP) revealed a two phase of Tau exchange with MTs as a rapid diffusion followed by a slower binding phase. A real-time FRET assay showed that high FRET occurred simultaneously with rescue and pause transitions at MT ends. To further explore the functional interaction of Tau to the MT, the binding of paclitaxel (PTX), tubulin acetylation induced by trichostatin A (TSA) and the expression of non-acetylable tubulin were used. With PTX and TSA, FRAP curves best fitted with a single phase with a long time constant, while with non-acetylable α-tubulin, curves best fitted a two phase recovery. Upon PTX and TSA incubations, quantities of high and low FRET hotspots decreased by up to 50% and no hotspot was observed during rescue and pause transitions. In the presence of non-acetylable α-tubulin, a 34% increase of low FRET hotspots was measured, and our real-time FRET assay revealed that low FRET hotspots appeared with MTs recovering growth. In conclusion, we have evidence by FRET and FRAP a discrete Tau-MT interaction where Tau could induce conformational changes of MTs, favoring recovery of MT self-assembly.