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The Rho GTPase Cdc42 regulates hair cell planar polarity and cellular patterning in the developing cochlea.

Kirjavainen A, Laos M, Anttonen T, Pirvola U - Biol Open (2015)

Bottom Line: Atypical protein kinase C (aPKC), a putative Cdc42 effector, colocalized with Cdc42 at the hair cell apex, and aPKC expression was altered upon Cdc42 depletion.Our data suggest that Cdc42 together with aPKC is part of the machinery establishing hair cell planar polarity and that Cdc42 acts on polarity through the cell-surface microtubule network.In addition, our data demonstrates that Cdc42 is required for stereociliogenesis in the immature cochlea.

View Article: PubMed Central - PubMed

Affiliation: Department of Biosciences, Viikinkaari 1, 00014 University of Helsinki, Finland.

No MeSH data available.


Related in: MedlinePlus

Impaired stereociliogenesis in outer hair cells of Cdc42loxP/loxP;Fgfr3-iCre-ERT2 mice postnatally.Confocal views of whole mount specimens from the medial coil of the cochlea. Boxes in A,B,E,F are enlarged in A',B',E',F', respectively. (A,A′) Phalloidin labeling shows the typical hair bundle morphology in the control specimen at P6. (B,B′) In the mutant cochlea at P6, all OHCs with stereociliary bundles are present, but some bundles are misoriented and have a fragmented or stunted appearance. (C,D) Degeneration of OHC bundles is also seen in transverse views where DAPI labels nuclei and acetylated tubulin marks kinocilia and microtubules. Arrows point to outer hair cells. (E,E′) Phalloidin labeling reveals the normal cellular organization and hair bundle morphology in the control specimen at P18. (F,F′) In the mutant cochlea, scattered OHC loss is seen (asterisks in F). Also, OHC bundles are fragmented and often composed of very short stereocilia (arrowheads in F′). (G,H) Unrecombined IHCs of mutants show stereociliary bundles comparable to controls. Abbreviations: OHCs, outer hair cells; IHC, inner hair cell; IP, inner pillar cell. Scale bar shown in H: A–D, 5 µm; E′,F′ 3 µm; A′,B′,G,H, 2 µm.
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f09: Impaired stereociliogenesis in outer hair cells of Cdc42loxP/loxP;Fgfr3-iCre-ERT2 mice postnatally.Confocal views of whole mount specimens from the medial coil of the cochlea. Boxes in A,B,E,F are enlarged in A',B',E',F', respectively. (A,A′) Phalloidin labeling shows the typical hair bundle morphology in the control specimen at P6. (B,B′) In the mutant cochlea at P6, all OHCs with stereociliary bundles are present, but some bundles are misoriented and have a fragmented or stunted appearance. (C,D) Degeneration of OHC bundles is also seen in transverse views where DAPI labels nuclei and acetylated tubulin marks kinocilia and microtubules. Arrows point to outer hair cells. (E,E′) Phalloidin labeling reveals the normal cellular organization and hair bundle morphology in the control specimen at P18. (F,F′) In the mutant cochlea, scattered OHC loss is seen (asterisks in F). Also, OHC bundles are fragmented and often composed of very short stereocilia (arrowheads in F′). (G,H) Unrecombined IHCs of mutants show stereociliary bundles comparable to controls. Abbreviations: OHCs, outer hair cells; IHC, inner hair cell; IP, inner pillar cell. Scale bar shown in H: A–D, 5 µm; E′,F′ 3 µm; A′,B′,G,H, 2 µm.

Mentions: To study the fate of OHCs with abnormal apical polarity, cochleas of the Cdc42loxP/loxP;Fgfr3-iCre-ERT2 mice (tamoxifen administration at E13.5 and E14.5) were analyzed at P6. Phalloidin labeling showed a normal complement of auditory hair cells in these animals, demonstrating that Cdc42 inactivation does not abrogate the survival of juvenile hair cells. OHCs with misoriented and dysmorphic stereociliary bundles were maintained in these cochleas. Acetylated tubulin immunostaining revealed the presence of the kinocilium in OHCs of both mutant and control mice. Notably, the staircase pattern of stereociliary bundles was in many cases disrupted. Bundles also appeared fragmented, suggesting that stereociliogenesis itself was impaired (Fig. 9A–D), consistent with strong Cdc42 expression in stereocilia (Fig. 2D). To confirm the involvement of Cdc42 in stereociliogenesis, P18 mutant mice were analyzed. In contrast to OHCs of control specimens that showed long and precisely arranged stereocilia, OHCs of the mutant mice had fragmented hair bundles consisting of stereocilia of variable lengths, some being very short (Fig. 9E,F). The unrecombined IHCs with normal bundles served as controls (Fig. 9G,H). Interestingly, at P18, besides the defects in OHC stereociliogenesis, scattered OHC loss was observed (Fig. 9F). Together, in addition to the control of hair cell patterning, shape and planar polarity, these results give evidence that Cdc42 regulates postnatal stereociliogenesis.


The Rho GTPase Cdc42 regulates hair cell planar polarity and cellular patterning in the developing cochlea.

Kirjavainen A, Laos M, Anttonen T, Pirvola U - Biol Open (2015)

Impaired stereociliogenesis in outer hair cells of Cdc42loxP/loxP;Fgfr3-iCre-ERT2 mice postnatally.Confocal views of whole mount specimens from the medial coil of the cochlea. Boxes in A,B,E,F are enlarged in A',B',E',F', respectively. (A,A′) Phalloidin labeling shows the typical hair bundle morphology in the control specimen at P6. (B,B′) In the mutant cochlea at P6, all OHCs with stereociliary bundles are present, but some bundles are misoriented and have a fragmented or stunted appearance. (C,D) Degeneration of OHC bundles is also seen in transverse views where DAPI labels nuclei and acetylated tubulin marks kinocilia and microtubules. Arrows point to outer hair cells. (E,E′) Phalloidin labeling reveals the normal cellular organization and hair bundle morphology in the control specimen at P18. (F,F′) In the mutant cochlea, scattered OHC loss is seen (asterisks in F). Also, OHC bundles are fragmented and often composed of very short stereocilia (arrowheads in F′). (G,H) Unrecombined IHCs of mutants show stereociliary bundles comparable to controls. Abbreviations: OHCs, outer hair cells; IHC, inner hair cell; IP, inner pillar cell. Scale bar shown in H: A–D, 5 µm; E′,F′ 3 µm; A′,B′,G,H, 2 µm.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4400594&req=5

f09: Impaired stereociliogenesis in outer hair cells of Cdc42loxP/loxP;Fgfr3-iCre-ERT2 mice postnatally.Confocal views of whole mount specimens from the medial coil of the cochlea. Boxes in A,B,E,F are enlarged in A',B',E',F', respectively. (A,A′) Phalloidin labeling shows the typical hair bundle morphology in the control specimen at P6. (B,B′) In the mutant cochlea at P6, all OHCs with stereociliary bundles are present, but some bundles are misoriented and have a fragmented or stunted appearance. (C,D) Degeneration of OHC bundles is also seen in transverse views where DAPI labels nuclei and acetylated tubulin marks kinocilia and microtubules. Arrows point to outer hair cells. (E,E′) Phalloidin labeling reveals the normal cellular organization and hair bundle morphology in the control specimen at P18. (F,F′) In the mutant cochlea, scattered OHC loss is seen (asterisks in F). Also, OHC bundles are fragmented and often composed of very short stereocilia (arrowheads in F′). (G,H) Unrecombined IHCs of mutants show stereociliary bundles comparable to controls. Abbreviations: OHCs, outer hair cells; IHC, inner hair cell; IP, inner pillar cell. Scale bar shown in H: A–D, 5 µm; E′,F′ 3 µm; A′,B′,G,H, 2 µm.
Mentions: To study the fate of OHCs with abnormal apical polarity, cochleas of the Cdc42loxP/loxP;Fgfr3-iCre-ERT2 mice (tamoxifen administration at E13.5 and E14.5) were analyzed at P6. Phalloidin labeling showed a normal complement of auditory hair cells in these animals, demonstrating that Cdc42 inactivation does not abrogate the survival of juvenile hair cells. OHCs with misoriented and dysmorphic stereociliary bundles were maintained in these cochleas. Acetylated tubulin immunostaining revealed the presence of the kinocilium in OHCs of both mutant and control mice. Notably, the staircase pattern of stereociliary bundles was in many cases disrupted. Bundles also appeared fragmented, suggesting that stereociliogenesis itself was impaired (Fig. 9A–D), consistent with strong Cdc42 expression in stereocilia (Fig. 2D). To confirm the involvement of Cdc42 in stereociliogenesis, P18 mutant mice were analyzed. In contrast to OHCs of control specimens that showed long and precisely arranged stereocilia, OHCs of the mutant mice had fragmented hair bundles consisting of stereocilia of variable lengths, some being very short (Fig. 9E,F). The unrecombined IHCs with normal bundles served as controls (Fig. 9G,H). Interestingly, at P18, besides the defects in OHC stereociliogenesis, scattered OHC loss was observed (Fig. 9F). Together, in addition to the control of hair cell patterning, shape and planar polarity, these results give evidence that Cdc42 regulates postnatal stereociliogenesis.

Bottom Line: Atypical protein kinase C (aPKC), a putative Cdc42 effector, colocalized with Cdc42 at the hair cell apex, and aPKC expression was altered upon Cdc42 depletion.Our data suggest that Cdc42 together with aPKC is part of the machinery establishing hair cell planar polarity and that Cdc42 acts on polarity through the cell-surface microtubule network.In addition, our data demonstrates that Cdc42 is required for stereociliogenesis in the immature cochlea.

View Article: PubMed Central - PubMed

Affiliation: Department of Biosciences, Viikinkaari 1, 00014 University of Helsinki, Finland.

No MeSH data available.


Related in: MedlinePlus