Microtubules

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Marcel Knossow - One of the best experts on this subject based on the ideXlab platform.

  • systematic identification of tubulin interacting fragments of the microtubule associated protein tau leads to a highly efficient promoter of microtubule assembly
    Journal of Biological Chemistry, 2011
    Co-Authors: Caroline Fauquant, Dries Verdegem, Virginie Redeker, Benoit Gigant, Isabelle Landrieu, Jean-michel Wieruszeski, Guy Lippens, Olivier Laprévote, Marcel Knossow
    Abstract:

    Tau is a microtubule-associated protein that stabilizes Microtubules and stimulates their assembly. Current descriptions of the tubulin-interacting regions of Tau involve Microtubules as the target and result mainly from deletions of Tau domains based on sequence analysis and from NMR spectroscopy experiments. Here, instead of Microtubules, we use the complex of two tubulin heterodimers with the stathmin-like domain of the RB3 protein (T2R) to identify interacting Tau fragments generated by limited proteolysis. We show that fragments in the proline-rich region and in the microtubule-binding repeats domain each interact on their own not only with T2R but also with Microtubules, albeit with moderate affinity. NMR analysis of the interaction with T2R of constructs in these two regions leads to a fragment, composed of adjacent parts of the microtubule-binding repeat domain and of the proline-rich region, that binds tightly to stabilized Microtubules. This demonstrates the synergy of the two Tau regions we identified in the Tau-microtubule interaction. Moreover, we show that this fragment, which binds to two tubulin heterodimers, stimulates efficiently microtubule assembly.

  • Systematic identification of tubulin interacting fragments of the microtubule-associated protein TAU leads to a highly efficient promoter of microtubule assembly.
    Journal of Biological Chemistry, 2011
    Co-Authors: Caroline Fauquant, Dries Verdegem, Virginie Redeker, Benoit Gigant, Isabelle Landrieu, Guy Lippens, Olivier Laprévote, Jean-michel Wieruzseski, Marcel Knossow
    Abstract:

    Tau is a microtubule-associated protein which stabilizes Microtubules and stimulates their assembly. Current descriptions of the tubulin-interacting regions of Tau involve Microtubules as the target and result mainly from deletions of Tau domains based on sequence analysis and from NMR spectroscopy experiments. Here, instead of Microtubules, we use the complex of two tubulin heterodimers with the stathmin-like domain of the RB3 protein (T2R) to identify interacting Tau fragments generated by limited proteolysis. We show that fragments in the proline rich region and in the microtubule binding repeats domain each interact on their own not only with T2R but also with Microtubules, albeit with moderate affinity. NMR analysis of the interaction with T2R of constructs in these two regions leads to a fragment composed of adjacent parts of the microtubule binding repeat domain and of the proline rich region that binds tightly to stabilized Microtubules. This demonstrates the synergy of the two Tau regions we identified in the Tau-microtubule interaction. Moreover, we show that this fragment binds two tubulin heterodimers in a complex that incorporates in Microtubules more readily than a single heterodimer and in this way stimulates microtubule assembly.

Caroline Fauquant - One of the best experts on this subject based on the ideXlab platform.

  • systematic identification of tubulin interacting fragments of the microtubule associated protein tau leads to a highly efficient promoter of microtubule assembly
    Journal of Biological Chemistry, 2011
    Co-Authors: Caroline Fauquant, Dries Verdegem, Virginie Redeker, Benoit Gigant, Isabelle Landrieu, Jean-michel Wieruszeski, Guy Lippens, Olivier Laprévote, Marcel Knossow
    Abstract:

    Tau is a microtubule-associated protein that stabilizes Microtubules and stimulates their assembly. Current descriptions of the tubulin-interacting regions of Tau involve Microtubules as the target and result mainly from deletions of Tau domains based on sequence analysis and from NMR spectroscopy experiments. Here, instead of Microtubules, we use the complex of two tubulin heterodimers with the stathmin-like domain of the RB3 protein (T2R) to identify interacting Tau fragments generated by limited proteolysis. We show that fragments in the proline-rich region and in the microtubule-binding repeats domain each interact on their own not only with T2R but also with Microtubules, albeit with moderate affinity. NMR analysis of the interaction with T2R of constructs in these two regions leads to a fragment, composed of adjacent parts of the microtubule-binding repeat domain and of the proline-rich region, that binds tightly to stabilized Microtubules. This demonstrates the synergy of the two Tau regions we identified in the Tau-microtubule interaction. Moreover, we show that this fragment, which binds to two tubulin heterodimers, stimulates efficiently microtubule assembly.

  • Systematic identification of tubulin interacting fragments of the microtubule-associated protein TAU leads to a highly efficient promoter of microtubule assembly.
    Journal of Biological Chemistry, 2011
    Co-Authors: Caroline Fauquant, Dries Verdegem, Virginie Redeker, Benoit Gigant, Isabelle Landrieu, Guy Lippens, Olivier Laprévote, Jean-michel Wieruzseski, Marcel Knossow
    Abstract:

    Tau is a microtubule-associated protein which stabilizes Microtubules and stimulates their assembly. Current descriptions of the tubulin-interacting regions of Tau involve Microtubules as the target and result mainly from deletions of Tau domains based on sequence analysis and from NMR spectroscopy experiments. Here, instead of Microtubules, we use the complex of two tubulin heterodimers with the stathmin-like domain of the RB3 protein (T2R) to identify interacting Tau fragments generated by limited proteolysis. We show that fragments in the proline rich region and in the microtubule binding repeats domain each interact on their own not only with T2R but also with Microtubules, albeit with moderate affinity. NMR analysis of the interaction with T2R of constructs in these two regions leads to a fragment composed of adjacent parts of the microtubule binding repeat domain and of the proline rich region that binds tightly to stabilized Microtubules. This demonstrates the synergy of the two Tau regions we identified in the Tau-microtubule interaction. Moreover, we show that this fragment binds two tubulin heterodimers in a complex that incorporates in Microtubules more readily than a single heterodimer and in this way stimulates microtubule assembly.

Anthony A Hyman - One of the best experts on this subject based on the ideXlab platform.

  • kinetically distinct phases of tau on Microtubules regulate kinesin motors and severing enzymes
    Nature Cell Biology, 2019
    Co-Authors: Valerie Siahaan, Amayra Hernandezvega, Stefan Diez, Anthony A Hyman, Jochen Krattenmacher, Zdenek Lansky, Marcus Braun
    Abstract:

    Tau is an intrinsically disordered protein, which diffuses on Microtubules1. In neurodegenerative diseases, collectively termed tauopathies, malfunction of tau and its detachment from axonal Microtubules are correlated with axonal degeneration2. Tau can protect Microtubules from microtubule-degrading enzymes such as katanin3. However, how tau carries out this regulatory function is still unclear. Here, using in vitro reconstitution, we show that tau molecules on Microtubules cooperatively form cohesive islands that are kinetically distinct from tau molecules that individually diffuse on Microtubules. Dependent on the tau concentration in solution, the islands reversibly grow or shrink by addition or release of tau molecules at their boundaries. Shielding Microtubules from kinesin-1 motors and katanin, the islands exhibit regulatory qualities distinct from a comparably dense layer of diffusible tau. Superprocessive kinesin-8 motors penetrate the islands and cause their disassembly. Our results reveal a microtubule-dependent phase of tau that constitutes an adaptable protective layer on the microtubule surface. We anticipate that other intrinsically disordered axonal proteins display a similar cooperative behaviour and potentially compete with tau in regulating access to the microtubule surface.

  • local nucleation of microtubule bundles through tubulin concentration into a condensed tau phase
    bioRxiv, 2017
    Co-Authors: Amayra Hernandezvega, Marcus Braun, Lara Scharrel, Marcus Jahnel, Susanne Wegmann, Simon Alberti, Stefan Diez, Bradley T Hyman, Anthony A Hyman
    Abstract:

    Non-centrosomal microtubule bundles play important roles in cellular organization and function. Although many diverse proteins are known that can bundle Microtubules, biochemical mechanisms by which cells could locally control the nucleation and formation of microtubule bundles are understudied. Here, we demonstrate that concentration of tubulin into a condensed, liquid-like compartment composed of the unstructured neuronal protein tau is sufficient to nucleate microtubule bundles. We show that under conditions of macro-molecular crowding, tau forms liquid drops. Tubulin partitions into these drops, efficiently increasing tubulin concentration and driving the nucleation of Microtubules. These growing Microtubules form bundles enclosed in a liquid sheath of tau. Our data suggest that condensed compartments of microtubule bundling proteins could promote the local formation of microtubule bundles in neurons by acting as non-centrosomal microtubule nucleation centers, and that liquid-like tau encapsulation could provide both stability and plasticity to long axonal microtubule bundles.

  • Preparation of marked Microtubules for the assay of the polarity of microtubule-based motors by fluorescence microscopy.
    Methods in Cell Biology, 2008
    Co-Authors: Jonathon Howard, Anthony A Hyman
    Abstract:

    Publisher Summary This chapter describes the preparation of marked Microtubules for the assay of the polarity of microtubule-based motors by fluorescence microscopy. It explores how to assemble Microtubules with obvious polarity when viewed by fluorescence microscopy; the direction of movement of a motor protein with respect to such polarity-marked Microtubules indicates the motor's polarity. Microtubules are polymers of the heterodimeric protein tubulin. The inherent polarity of the microtubule polymer has two major functional implications—namely, (1) the growth rates at the different ends of the Microtubules are different and (2) different microtubule-based motor proteins move in opposite directions along the microtubule. Thus, the cylindrical surface of the microtubule has a different texture in the two directions, a texture that the motor proteins can detect. The knowledge of the polarity of movement of a motor protein is essential for interpreting its biological function. Microtubule nucleation will not occur below a certain critical concentration, but new microtubule growth will occur from the pre-existing ends of Microtubules down to tubulin concentrations below that critical concentration.

  • binding of the adenomatous polyposis coli protein to Microtubules increases microtubule stability and is regulated by gsk3β phosphorylation
    Current Biology, 2001
    Co-Authors: Jurg Zumbrunn, Anthony A Hyman, Kazuhisa Kinoshita, Inke S Nathke
    Abstract:

    Abstract Truncation mutations in the adenomatous polyposis coli protein (APC) are responsible for familial polyposis, a form of inherited colon cancer. In addition to its role in mediating β-catenin degradation in the Wnt signaling pathway, APC plays a role in regulating Microtubules. This was suggested by its localization to the end of dynamic Microtubules in actively migrating areas of cells and by the apparent correlation between the dissociation of APC from polymerizing Microtubules and their subsequent depolymerization [1, 2]. The microtubule binding domain is deleted in the transforming mutations of APC [3, 4]; however, the direct effect of APC protein on Microtubules has never been examined. Here we show that binding of APC to Microtubules increases microtubule stability in vivo and in vitro. Deleting the previously identified microtubule binding site from the C-terminal domain of APC does not eliminate its binding to Microtubules but decreases the ability of APC to stabilize them significantly. The interaction of APC with Microtubules is decreased by phosphorylation of APC by GSK3β. These data confirm the hypothesis that APC is involved in stabilizing microtubule ends. They also suggest that binding of APC to Microtubules is mediated by at least two distinct sites and is regulated by phosphorylation.

Dries Verdegem - One of the best experts on this subject based on the ideXlab platform.

  • systematic identification of tubulin interacting fragments of the microtubule associated protein tau leads to a highly efficient promoter of microtubule assembly
    Journal of Biological Chemistry, 2011
    Co-Authors: Caroline Fauquant, Dries Verdegem, Virginie Redeker, Benoit Gigant, Isabelle Landrieu, Jean-michel Wieruszeski, Guy Lippens, Olivier Laprévote, Marcel Knossow
    Abstract:

    Tau is a microtubule-associated protein that stabilizes Microtubules and stimulates their assembly. Current descriptions of the tubulin-interacting regions of Tau involve Microtubules as the target and result mainly from deletions of Tau domains based on sequence analysis and from NMR spectroscopy experiments. Here, instead of Microtubules, we use the complex of two tubulin heterodimers with the stathmin-like domain of the RB3 protein (T2R) to identify interacting Tau fragments generated by limited proteolysis. We show that fragments in the proline-rich region and in the microtubule-binding repeats domain each interact on their own not only with T2R but also with Microtubules, albeit with moderate affinity. NMR analysis of the interaction with T2R of constructs in these two regions leads to a fragment, composed of adjacent parts of the microtubule-binding repeat domain and of the proline-rich region, that binds tightly to stabilized Microtubules. This demonstrates the synergy of the two Tau regions we identified in the Tau-microtubule interaction. Moreover, we show that this fragment, which binds to two tubulin heterodimers, stimulates efficiently microtubule assembly.

  • Systematic identification of tubulin interacting fragments of the microtubule-associated protein TAU leads to a highly efficient promoter of microtubule assembly.
    Journal of Biological Chemistry, 2011
    Co-Authors: Caroline Fauquant, Dries Verdegem, Virginie Redeker, Benoit Gigant, Isabelle Landrieu, Guy Lippens, Olivier Laprévote, Jean-michel Wieruzseski, Marcel Knossow
    Abstract:

    Tau is a microtubule-associated protein which stabilizes Microtubules and stimulates their assembly. Current descriptions of the tubulin-interacting regions of Tau involve Microtubules as the target and result mainly from deletions of Tau domains based on sequence analysis and from NMR spectroscopy experiments. Here, instead of Microtubules, we use the complex of two tubulin heterodimers with the stathmin-like domain of the RB3 protein (T2R) to identify interacting Tau fragments generated by limited proteolysis. We show that fragments in the proline rich region and in the microtubule binding repeats domain each interact on their own not only with T2R but also with Microtubules, albeit with moderate affinity. NMR analysis of the interaction with T2R of constructs in these two regions leads to a fragment composed of adjacent parts of the microtubule binding repeat domain and of the proline rich region that binds tightly to stabilized Microtubules. This demonstrates the synergy of the two Tau regions we identified in the Tau-microtubule interaction. Moreover, we show that this fragment binds two tubulin heterodimers in a complex that incorporates in Microtubules more readily than a single heterodimer and in this way stimulates microtubule assembly.

Virginie Redeker - One of the best experts on this subject based on the ideXlab platform.

  • systematic identification of tubulin interacting fragments of the microtubule associated protein tau leads to a highly efficient promoter of microtubule assembly
    Journal of Biological Chemistry, 2011
    Co-Authors: Caroline Fauquant, Dries Verdegem, Virginie Redeker, Benoit Gigant, Isabelle Landrieu, Jean-michel Wieruszeski, Guy Lippens, Olivier Laprévote, Marcel Knossow
    Abstract:

    Tau is a microtubule-associated protein that stabilizes Microtubules and stimulates their assembly. Current descriptions of the tubulin-interacting regions of Tau involve Microtubules as the target and result mainly from deletions of Tau domains based on sequence analysis and from NMR spectroscopy experiments. Here, instead of Microtubules, we use the complex of two tubulin heterodimers with the stathmin-like domain of the RB3 protein (T2R) to identify interacting Tau fragments generated by limited proteolysis. We show that fragments in the proline-rich region and in the microtubule-binding repeats domain each interact on their own not only with T2R but also with Microtubules, albeit with moderate affinity. NMR analysis of the interaction with T2R of constructs in these two regions leads to a fragment, composed of adjacent parts of the microtubule-binding repeat domain and of the proline-rich region, that binds tightly to stabilized Microtubules. This demonstrates the synergy of the two Tau regions we identified in the Tau-microtubule interaction. Moreover, we show that this fragment, which binds to two tubulin heterodimers, stimulates efficiently microtubule assembly.

  • Systematic identification of tubulin interacting fragments of the microtubule-associated protein TAU leads to a highly efficient promoter of microtubule assembly.
    Journal of Biological Chemistry, 2011
    Co-Authors: Caroline Fauquant, Dries Verdegem, Virginie Redeker, Benoit Gigant, Isabelle Landrieu, Guy Lippens, Olivier Laprévote, Jean-michel Wieruzseski, Marcel Knossow
    Abstract:

    Tau is a microtubule-associated protein which stabilizes Microtubules and stimulates their assembly. Current descriptions of the tubulin-interacting regions of Tau involve Microtubules as the target and result mainly from deletions of Tau domains based on sequence analysis and from NMR spectroscopy experiments. Here, instead of Microtubules, we use the complex of two tubulin heterodimers with the stathmin-like domain of the RB3 protein (T2R) to identify interacting Tau fragments generated by limited proteolysis. We show that fragments in the proline rich region and in the microtubule binding repeats domain each interact on their own not only with T2R but also with Microtubules, albeit with moderate affinity. NMR analysis of the interaction with T2R of constructs in these two regions leads to a fragment composed of adjacent parts of the microtubule binding repeat domain and of the proline rich region that binds tightly to stabilized Microtubules. This demonstrates the synergy of the two Tau regions we identified in the Tau-microtubule interaction. Moreover, we show that this fragment binds two tubulin heterodimers in a complex that incorporates in Microtubules more readily than a single heterodimer and in this way stimulates microtubule assembly.