Acetylated tubulin is essential for touch sensation in mice
- EMBL Mouse Biology Unit, Italy
- Molecular Medicine Partnership Unit (MMPU), Germany
- Centre for Integrative Neuroscience, Germany
- Istituto Officina dei Materiali-CNR, Italy
- Max Planck Institute of Molecular Cell Biology and Genetics, Germany
- European Molecular Biology Laboratory, Germany
- Ecole Polytechnique Federale de Lausanne, Switzerland
- Consiglio Nazionale delle Ricerche, Italy
Figures
Figure 1 with 1 supplement
Behavioural analysis of Atat1cKOmice.
(a) Bar-chart summarising the results of a tape test to assay low threshold mechanosensation. Atat1cKO mice demonstrated significantly less response events over the 5 min counting period (t-Test, p<0…
Figure 1—figure supplement 1
Behavioral analysis of Atat1cKO mice.
(a) Bar-chart showing the latency of response from Atat1Control and Atat1cKO mice to immersion in a water bath at 46, 48 and 50°C respectively (unpaired t test, p>0.05 for all temperatures tested). …
Figure 2 with 3 supplements
αTAT1 is required for mechanosensitivity of low threshold mechanoreceptors.
Slowly adapting mechanoreceptor fibers (SAM): Typical responses (top) and stimulus-response functions (bottom) to increasing displacement for the ramp (a) and static (b) phase (two-way ANOVA with pos…
Figure 2—figure supplement 1
Electrical excitability and stimulus response properties of SAM fibres in Atat1Control and Atat1cKO mice.
(a) Mean discharge rates (1 s bins) during the course of ramp and static stimuli. Note the significant reduction of dynamic ramp discharge recorded from Atat1cKO mice. (b) A cumulative sum plot of …
Figure 2—figure supplement 2
Electrical excitability and stimulus response properties of RAM fibres in Atat1Control and Atat1cKO mice.
(a) A cumulative sum plot of displacement stimulus threshold. (b) Mechanical latency as a function of displacement (two-way ANOVA with post-hoc Bonferroni’s test, p>0.05). (c) A cumulative sum plot …
Figure 2—figure supplement 3
Electrical excitability and stimulus response properties of D-hair fibres in Atat1Control and Atat1cKO mice.
(a) A cumulative sum plot of displacement stimulus threshold. (b) Mechanical latency as a function of displacement (two-way ANOVA with post-hoc Bonferroni’s test, p>0.05). (c) A cumulative sum plot …
Figure 3 with 3 supplements
αTAT1 is required for mechanosensitivity of nociceptors.
Typical responses (top) and mean discharge rates (1 s bins) during 10 s 150 µm stimulus of Aδ-mechanonociceptors (AM) (a) and C-fibre nociceptors (c) from αTAT1control and αTAT1cko mice (two-way …
Figure 3—figure supplement 1
Electrical excitability and stimulus response properties of AM fibres in Atat1Control and Atat1cKO mice.
(a) A cumulative sum plot of displacement stimulus threshold. (b) Mechanical latency as a function of displacement (two-way ANOVA with post-hoc Bonferroni’s test, p<0.05). (c) Mean von Frey …
Figure 3—figure supplement 2
Electrical excitability and stimulus response properties of C fibres in Atat1Control and Atat1cKO mice.
(a) A cumulative sum plot of displacement stimulus threshold. (b) Mechanical latency as a function of displacement (two-way ANOVA with post-hoc Bonferroni’s test, p<0.01). (c) Mean von Frey …
Figure 3—figure supplement 3
Heat response of C fibers in Atat1Control and Atat1cKO mice.
(a) Proportions of C fibers responding to noxious heat. C-M, C mechanonociceptor; C-MH, C mechano/heat receptor, responding both to noxious heat and mechanical stimuli. No significant difference was …
Figure 4 with 2 supplements
Morphological analysis of the peripheral nervous system in Atat1cKO mice.
(a and b) Electron micrographs of a sectioned saphenous nerve from an Atat1Control (a) and Atat1cKO (b) mouse (Scale bar 5 μm). (c and d) Graph summarizing the number of myelinated fibres (c) and …
Figure 4—figure supplement 1
Spinal cord staining in Atat1Control and Atat1cKO mice.
(a) Representative fluorescent image of a 100 μM section of spinal cord taken from an Atat1Control mouse stained with IB4 (green) and NF200 (red) (Scale bar 100 μM). (b) Representative fluorescent …
Figure 4—figure supplement 2
Axonal transport in DRG neurons from Atat1cKO mice.
(a) Single molecule tracking of NGF molecules linked to Quantum dots in neurites from Atat1Control and Atat1cKO (b) DRG grown in microfluidic devices. (c) Average response. Error bars indicate SEM.
Figure 5 with 4 supplements
Atat1-mediated acetylated microtubules regulate mechanosensitivity in sensory neurons.
(a) Stacked histograms showing the proportion of different mechano-gated currents activated by neurite indentation in sensory neurons from control Atat1Control and Atat1cKO mice (χ2 test, p<0.05). …
Figure 5—figure supplement 1
Absence of Atat1 in sensory neurons does not alter kinetic properties of RA currents.
(a) The proportion of neurons that display a mechano-gated response as function of displacement was plotted for Atat1Control (n = 15) and Atat1cKO (n = 31) sensory neurons. Quantitative comparison …
Figure 5—figure supplement 2
Absence of Atat1 in sensory neurons does not alter membrane electrophysiological properties and proton activated currents.
(a, b) Absence of Atat1 in sensory neurons does not alter voltage gated currents. Mean whole-cell inward and outward currents are measured at different test potentials for Atat1Control and Atat1cKO …
Figure 5—figure supplement 3
Capsaicin response of DRG neurons in Atat1Control and Atat1cKO mice.
(a) The proportion of small neurons (diameter < 25 μm) displaying capsaicin evoked calcium transients was not different between Atat1Control and Atat1cKO mice (100 nM capsaicin: Atat1Control, n = 169…
Figure 5—figure supplement 4
FRAP analysis of Piezo2 in transfected DRG from Atat1Control and Atat1cKO mice.
(a) Graph of recovery from photobleaching of Piezo2-GFP construct transfected into DRG neurons from Atat1Control and Atat1cKO mice. (b) Halftime of recovery in Atat1Control and Atat1cKO mice (p>0.05,…
Figure 6 with 1 supplement
Superresolution imaging of microtubules in DRG from Atat1Control and Atat1cKO mice.
(a and b) Superresolution image of anti α-tubulin staining of Atat1Control (a) and Atat1cKO (b) DRG colour coded by depth (red close to objective, Scale bar 5 μm). (c) Graphical representation of …
Figure 6—figure supplement 1
Actin cytoskeletal organization in DRG from Atat1cKO mice.
(a) Fluorescent image of a cultured Atat1Control DRG stained with phalloidin (green) and Hoechst (blue) (Scale bar 10 μm). (b) Fluorescent image of a cultured Atat1cKO DRG stained with phalloidin …
Figure 7 with 2 supplements
Microtubule organization in peripheral sensory neurons.
(a) Anti-acetylated-α-tubulin staining of Atat1Control cultured DRG cells (corresponding surface plot below). Note the prominent sub-membrane localisation of acetylated tubulin (Scale bar 5 μm). (b) …
Figure 7—figure supplement 1
Acetylated microtubule distribution in sensory neurons from Atat1cKO mice.
(a) Low magnification image of anti-acetylated tubulin stained (red) cultured DRG. Note the prominent sub-membrane localization of the stain in many neuronal soma (Scale bar 30 μm). (b) …
Figure 7—figure supplement 2
Atomic Force Microscopy of DRG from Atat1cKO mice.
(a) Sketch of AFM set-up used for indentation measurements including a bead 4.5 μm diameter glued at the end of a tipless cantilever. The cantilever is moved by a piezo-system: the bending of the …
Videos
Video 1
Tape test assay.
Video recording of mice showing attempts to remove a piece of tape by Atat1Control (left) and Atat1cKO (right), over a 5 min period.
Video 2
Tail clip assay.
Video of Atat1Control (top) and Atat1cKO (bottom) mice to measure the time taken until the first response of the animals to the clip.
Video 3
Microfluidics.
Movement of NGF labelled quantum dots along the neurites of cultured DRG. Atat1Control (left) and Atat1cKO (right).
Video 4
Acetylated tubulin distribution in cornea.
3D rendering of a whole mount cornea preparation showing membrane bound SNAP labeling in red and acetylated tubulin in green.
