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Single-molecule tracking of tau reveals fast kiss-and-hop interaction with microtubules in living neurons.

Janning D, Igaev M, Sündermann F, Brühmann J, Beutel O, Heinisch JJ, Bakota L, Piehler J, Junge W, Brandt R - Mol. Biol. Cell (2014)

Bottom Line: Furthermore, we observed by quantitative imaging using fluorescence decay after photoactivation recordings of photoactivatable GFP-tagged tubulin that, despite this rapid dynamics, tau is capable of regulating the tubulin-microtubule balance.Our data imply a novel kiss-and-hop mechanism by which tau promotes neuronal microtubule assembly.The rapid kiss-and-hop interaction explains why tau, although binding to microtubules, does not interfere with axonal transport.

View Article: PubMed Central - PubMed

Affiliation: Department of Neurobiology, University of Osnabrück, D-49076 Osnabrück, Germany.

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Single-molecule tracking reveals undirected fast movement of tau in processes of neural cells. (A) Schematic representation of the Halo-tagged tau fusion construct (Halo-tau) and the visualization of a thin cellular layer by TIRF microscopy. The microtubule-binding repeats (R1–R4) are indicated by the gray boxes, and the evanescent field close to the cellular attachment site is shown in yellow. Bottom, colocalization of Halo-tau (left) and monomeric enhanced GFP–tagged α-tubulin (EGFP-tub; right) as visualized by TIRF microscopy of the cell body of living PC12 cells. Sum projections of 1000 consecutive frames were recorded. Scale bar, 10 μm. MTOC, microtubule-organizing center. (B) Time series of individual Halo-tau molecules moving in a process of a neuronally differentiated PC12 cell. Motion during the first 85 ms. Dashed lines indicate the border of the process. A trajectory generated from the complete time series (2.2 s = 440 frames) is shown below, indicating undirected fast movement both longitudinally and transversally. The starting point is indicated by a circle and the end by a square. Scale bar, 0.5 μm.
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Figure 1: Single-molecule tracking reveals undirected fast movement of tau in processes of neural cells. (A) Schematic representation of the Halo-tagged tau fusion construct (Halo-tau) and the visualization of a thin cellular layer by TIRF microscopy. The microtubule-binding repeats (R1–R4) are indicated by the gray boxes, and the evanescent field close to the cellular attachment site is shown in yellow. Bottom, colocalization of Halo-tau (left) and monomeric enhanced GFP–tagged α-tubulin (EGFP-tub; right) as visualized by TIRF microscopy of the cell body of living PC12 cells. Sum projections of 1000 consecutive frames were recorded. Scale bar, 10 μm. MTOC, microtubule-organizing center. (B) Time series of individual Halo-tau molecules moving in a process of a neuronally differentiated PC12 cell. Motion during the first 85 ms. Dashed lines indicate the border of the process. A trajectory generated from the complete time series (2.2 s = 440 frames) is shown below, indicating undirected fast movement both longitudinally and transversally. The starting point is indicated by a circle and the end by a square. Scale bar, 0.5 μm.

Mentions: To assess the behavior of tau in neuronal processes, single, fluorescently labeled tau molecules were tracked with a temporal resolution of 5 ms. HaloTag was fused to the amino terminus of human tau, where it does not interfere with the MT interaction that is mediated by tau's carboxy-terminal half (Gauthier-Kemper et al., 2011; Figure 1A, top). The focal plane of the total internal reflection fluorescence (TIRF) microscope was adjusted to capture MTs (Figure 1A).


Single-molecule tracking of tau reveals fast kiss-and-hop interaction with microtubules in living neurons.

Janning D, Igaev M, Sündermann F, Brühmann J, Beutel O, Heinisch JJ, Bakota L, Piehler J, Junge W, Brandt R - Mol. Biol. Cell (2014)

Single-molecule tracking reveals undirected fast movement of tau in processes of neural cells. (A) Schematic representation of the Halo-tagged tau fusion construct (Halo-tau) and the visualization of a thin cellular layer by TIRF microscopy. The microtubule-binding repeats (R1–R4) are indicated by the gray boxes, and the evanescent field close to the cellular attachment site is shown in yellow. Bottom, colocalization of Halo-tau (left) and monomeric enhanced GFP–tagged α-tubulin (EGFP-tub; right) as visualized by TIRF microscopy of the cell body of living PC12 cells. Sum projections of 1000 consecutive frames were recorded. Scale bar, 10 μm. MTOC, microtubule-organizing center. (B) Time series of individual Halo-tau molecules moving in a process of a neuronally differentiated PC12 cell. Motion during the first 85 ms. Dashed lines indicate the border of the process. A trajectory generated from the complete time series (2.2 s = 440 frames) is shown below, indicating undirected fast movement both longitudinally and transversally. The starting point is indicated by a circle and the end by a square. Scale bar, 0.5 μm.
© Copyright Policy - creative-commons
Related In: Results  -  Collection

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Figure 1: Single-molecule tracking reveals undirected fast movement of tau in processes of neural cells. (A) Schematic representation of the Halo-tagged tau fusion construct (Halo-tau) and the visualization of a thin cellular layer by TIRF microscopy. The microtubule-binding repeats (R1–R4) are indicated by the gray boxes, and the evanescent field close to the cellular attachment site is shown in yellow. Bottom, colocalization of Halo-tau (left) and monomeric enhanced GFP–tagged α-tubulin (EGFP-tub; right) as visualized by TIRF microscopy of the cell body of living PC12 cells. Sum projections of 1000 consecutive frames were recorded. Scale bar, 10 μm. MTOC, microtubule-organizing center. (B) Time series of individual Halo-tau molecules moving in a process of a neuronally differentiated PC12 cell. Motion during the first 85 ms. Dashed lines indicate the border of the process. A trajectory generated from the complete time series (2.2 s = 440 frames) is shown below, indicating undirected fast movement both longitudinally and transversally. The starting point is indicated by a circle and the end by a square. Scale bar, 0.5 μm.
Mentions: To assess the behavior of tau in neuronal processes, single, fluorescently labeled tau molecules were tracked with a temporal resolution of 5 ms. HaloTag was fused to the amino terminus of human tau, where it does not interfere with the MT interaction that is mediated by tau's carboxy-terminal half (Gauthier-Kemper et al., 2011; Figure 1A, top). The focal plane of the total internal reflection fluorescence (TIRF) microscope was adjusted to capture MTs (Figure 1A).

Bottom Line: Furthermore, we observed by quantitative imaging using fluorescence decay after photoactivation recordings of photoactivatable GFP-tagged tubulin that, despite this rapid dynamics, tau is capable of regulating the tubulin-microtubule balance.Our data imply a novel kiss-and-hop mechanism by which tau promotes neuronal microtubule assembly.The rapid kiss-and-hop interaction explains why tau, although binding to microtubules, does not interfere with axonal transport.

View Article: PubMed Central - PubMed

Affiliation: Department of Neurobiology, University of Osnabrück, D-49076 Osnabrück, Germany.

Show MeSH
Related in: MedlinePlus