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Staurosporine-induced collapse of cochlear hair bundles.

Goodyear RJ, Ratnayaka HS, Warchol ME, Richardson GP - J. Comp. Neurol. (2014)

Bottom Line: Staurosporine exposure results in the fusion of the hair bundle's stereocilia, a resorption of the parallel actin bundles of the stereocilia into the cytoplasm of the hair cell, a detachment of the apical, non-stereociliary membrane of the hair cell from the underlying cuticular plate, and a severing of the hair-bundle's rootlets from the actin cores of the stereocilia.It does not block membrane retrieval at the apical pole of the hair cells, nor does it elicit the externalization of phosphatidylserine.Staurosporine treatment causes a reduction in levels of the phosphorylated forms of ezrin, radixin, and moesin in cochlear cultures during the period of hair-bundle loss, indicating the integrity of the hair bundle may be actively maintained by the phosphorylation status of these proteins.

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Affiliation: Sussex Neuroscience and School of Life Sciences, University of Sussex, Brighton, BN1 9QG, UK.

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Phospho-ERM levels in staurosporine-treated cochlear cultures. Immunolabeling with anti-phospho-ERM (A–D) and anti-radixin (E–H) antibodies following incubation in control medium (A,B,E,F) or medium containing 10 nM staurosporine (C,D,G,H) for 5 hours. Phalloidin staining is also shown in (B,D,F,H). Staurosporine causes a dramatic loss of phospho-ERM (C) but not of radixin labeling (G). I: Western blots of lysates from cochlear cultures stained for phospho-ERM or radixin following 14 hours in control medium or medium containing 10 nM staurosporine. J: Quantitative analysis of levels of radixin (red bars) and phospho-radixin (green bars) in cochlear cultures that were treated with 10 nM staurosporine for 1, 2.5, 5, 10, and 14 hours. Numbers of independent experiments are indicated. A magenta-green version of this figure is available as Supporting Figure 2. Scale bar = 10 μm in H (applies to A–H).
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fig08: Phospho-ERM levels in staurosporine-treated cochlear cultures. Immunolabeling with anti-phospho-ERM (A–D) and anti-radixin (E–H) antibodies following incubation in control medium (A,B,E,F) or medium containing 10 nM staurosporine (C,D,G,H) for 5 hours. Phalloidin staining is also shown in (B,D,F,H). Staurosporine causes a dramatic loss of phospho-ERM (C) but not of radixin labeling (G). I: Western blots of lysates from cochlear cultures stained for phospho-ERM or radixin following 14 hours in control medium or medium containing 10 nM staurosporine. J: Quantitative analysis of levels of radixin (red bars) and phospho-radixin (green bars) in cochlear cultures that were treated with 10 nM staurosporine for 1, 2.5, 5, 10, and 14 hours. Numbers of independent experiments are indicated. A magenta-green version of this figure is available as Supporting Figure 2. Scale bar = 10 μm in H (applies to A–H).

Mentions: The effects of staurosporine observed by transmission electron microscopy had, in some instances, a striking resemblance to the changes in hair-bundle structure described in radixin mutant mice (see fig. 5 in Kitajiri et al., 2004). The distribution of radixin and phosphorylated forms of the ERM proteins were therefore examined in control cultures and cultures that had been treated with 10 nM staurosporine for 5 hours (Fig. 8). Strong labeling of the microvilli on the surfaces of the supporting cells and a much weaker labeling of hair bundles was observed in control cultures with antibodies to phospho-ERM (Fig. 8A). Staurosporine treatment resulted in almost complete loss of all labeling (Fig. 8C). A polyclonal antibody to human radixin stained the sensory hair bundles and the microvilli of the surrounding supporting cells (Fig. 8E), and this distribution remained unchanged in staurosporine-treated cultures (Fig. 8G) despite collapse of the hair bundles.


Staurosporine-induced collapse of cochlear hair bundles.

Goodyear RJ, Ratnayaka HS, Warchol ME, Richardson GP - J. Comp. Neurol. (2014)

Phospho-ERM levels in staurosporine-treated cochlear cultures. Immunolabeling with anti-phospho-ERM (A–D) and anti-radixin (E–H) antibodies following incubation in control medium (A,B,E,F) or medium containing 10 nM staurosporine (C,D,G,H) for 5 hours. Phalloidin staining is also shown in (B,D,F,H). Staurosporine causes a dramatic loss of phospho-ERM (C) but not of radixin labeling (G). I: Western blots of lysates from cochlear cultures stained for phospho-ERM or radixin following 14 hours in control medium or medium containing 10 nM staurosporine. J: Quantitative analysis of levels of radixin (red bars) and phospho-radixin (green bars) in cochlear cultures that were treated with 10 nM staurosporine for 1, 2.5, 5, 10, and 14 hours. Numbers of independent experiments are indicated. A magenta-green version of this figure is available as Supporting Figure 2. Scale bar = 10 μm in H (applies to A–H).
© Copyright Policy - open-access
Related In: Results  -  Collection

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Show All Figures
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fig08: Phospho-ERM levels in staurosporine-treated cochlear cultures. Immunolabeling with anti-phospho-ERM (A–D) and anti-radixin (E–H) antibodies following incubation in control medium (A,B,E,F) or medium containing 10 nM staurosporine (C,D,G,H) for 5 hours. Phalloidin staining is also shown in (B,D,F,H). Staurosporine causes a dramatic loss of phospho-ERM (C) but not of radixin labeling (G). I: Western blots of lysates from cochlear cultures stained for phospho-ERM or radixin following 14 hours in control medium or medium containing 10 nM staurosporine. J: Quantitative analysis of levels of radixin (red bars) and phospho-radixin (green bars) in cochlear cultures that were treated with 10 nM staurosporine for 1, 2.5, 5, 10, and 14 hours. Numbers of independent experiments are indicated. A magenta-green version of this figure is available as Supporting Figure 2. Scale bar = 10 μm in H (applies to A–H).
Mentions: The effects of staurosporine observed by transmission electron microscopy had, in some instances, a striking resemblance to the changes in hair-bundle structure described in radixin mutant mice (see fig. 5 in Kitajiri et al., 2004). The distribution of radixin and phosphorylated forms of the ERM proteins were therefore examined in control cultures and cultures that had been treated with 10 nM staurosporine for 5 hours (Fig. 8). Strong labeling of the microvilli on the surfaces of the supporting cells and a much weaker labeling of hair bundles was observed in control cultures with antibodies to phospho-ERM (Fig. 8A). Staurosporine treatment resulted in almost complete loss of all labeling (Fig. 8C). A polyclonal antibody to human radixin stained the sensory hair bundles and the microvilli of the surrounding supporting cells (Fig. 8E), and this distribution remained unchanged in staurosporine-treated cultures (Fig. 8G) despite collapse of the hair bundles.

Bottom Line: Staurosporine exposure results in the fusion of the hair bundle's stereocilia, a resorption of the parallel actin bundles of the stereocilia into the cytoplasm of the hair cell, a detachment of the apical, non-stereociliary membrane of the hair cell from the underlying cuticular plate, and a severing of the hair-bundle's rootlets from the actin cores of the stereocilia.It does not block membrane retrieval at the apical pole of the hair cells, nor does it elicit the externalization of phosphatidylserine.Staurosporine treatment causes a reduction in levels of the phosphorylated forms of ezrin, radixin, and moesin in cochlear cultures during the period of hair-bundle loss, indicating the integrity of the hair bundle may be actively maintained by the phosphorylation status of these proteins.

View Article: PubMed Central - PubMed

Affiliation: Sussex Neuroscience and School of Life Sciences, University of Sussex, Brighton, BN1 9QG, UK.

Show MeSH
Related in: MedlinePlus