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Neurod1 suppresses hair cell differentiation in ear ganglia and regulates hair cell subtype development in the cochlea.

Jahan I, Pan N, Kersigo J, Fritzsch B - PLoS ONE (2010)

Bottom Line: Our data suggest that the long noted cross-regulation of Atoh1 expression by Neurog1 might actually be mediated in large part by Neurod1.We suggest that Neurod1 is regulated by both Neurog1 and Atoh1 and provides a negative feedback for either gene.Through this and other feedback, Neurod1 suppresses alternate fates of neurons to differentiate as hair cells and regulates hair cell subtypes.

View Article: PubMed Central - PubMed

Affiliation: Department of Biology, University of Iowa, Iowa City, Iowa, United States of America.

ABSTRACT

Background: At least five bHLH genes regulate cell fate determination and differentiation of sensory neurons, hair cells and supporting cells in the mammalian inner ear. Cross-regulation of Atoh1 and Neurog1 results in hair cell changes in Neurog1 mice although the nature and mechanism of the cross-regulation has not yet been determined. Neurod1, regulated by both Neurog1 and Atoh1, could be the mediator of this cross-regulation.

Methodology/principal findings: We used Tg(Pax2-Cre) to conditionally delete Neurod1 in the inner ear. Our data demonstrate for the first time that the absence of Neurod1 results in formation of hair cells within the inner ear sensory ganglia. Three cell types, neural crest derived Schwann cells and mesenchyme derived fibroblasts (neither expresses Neurod1) and inner ear derived neurons (which express Neurod1) constitute inner ear ganglia. The most parsimonious explanation is that Neurod1 suppresses the alternative fate of sensory neurons to develop as hair cells. In the absence of Neurod1, Atoh1 is expressed and differentiates cells within the ganglion into hair cells. We followed up on this effect in ganglia by demonstrating that Neurod1 also regulates differentiation of subtypes of hair cells in the organ of Corti. We show that in Neurod1 conditional mice there is a premature expression of several genes in the apex of the developing cochlea and outer hair cells are transformed into inner hair cells.

Conclusions/significance: Our data suggest that the long noted cross-regulation of Atoh1 expression by Neurog1 might actually be mediated in large part by Neurod1. We suggest that Neurod1 is regulated by both Neurog1 and Atoh1 and provides a negative feedback for either gene. Through this and other feedback, Neurod1 suppresses alternate fates of neurons to differentiate as hair cells and regulates hair cell subtypes.

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Related in: MedlinePlus

Fgf8 misexpression correlates with formation of ‘ectopic inner hair cells’.In situ hybridization shows a persistent expression of Fgf8 in neurosensory precursors in Neurod1 mutant cochlea as early as E12.5 (arrow in B’) with premature expression in particular in the apex of the cochlea (C’) in comparison to wild-type (C). Fgf8 is expressed transiently in the prosensory domain in wild-type mice (arrow in A) and later is shown to be upregulated in the cochlea with a base-to-apex gradient (A,B,C). In newborn mice, Fgf8 is uniformly expressed in all inner hair cells almost along the entire length of the cochlea in wild-type mice (D, D’). In contrast, Neurod1 CKO mice display an increased expression level in the apex (E, F, H, I, J) and deviate from the single row labeling of only inner hair cells seen in the base (G). Two or more rows of inner hair cells are positive for Fgf8 and scattered single and multiple cells are interspersed among the multiple rows of outer hair cells which are also positive for Fgf8 (arrows in I). The apical tip shows up to three rows of Fgf8 positive cells (J). Radial sections through the apex of Fgf8 ISH reacted and Myo VIIa immunostained cochlea reveals co-localization of Fgf8 and Myo VIIa in inner as well as ‘ectopic inner hair cells’ scattered among outer hair cells (arrows in K–K”). SEM of P30 Neurod1 CKO mice reveals a normal organization of inner hair cells in a single row and three rows of outer hair cells in the base of Neurod1 CKO mice (L). In contrast, the apex shows inner hair cell sized stereocilia (arrows) interspersed among normal sized stereocilia bearing outer hair cells (arrowhead; M,N). Dotted line in H indicates border between normal and disorganized organ of Corti. IHC, inner hair cells; OHC, outer hair cells. Bar indicates 100 µm in A–F except D’; 10 µm in D’,G–J, K–K” and 1 µm in L–N.
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pone-0011661-g008: Fgf8 misexpression correlates with formation of ‘ectopic inner hair cells’.In situ hybridization shows a persistent expression of Fgf8 in neurosensory precursors in Neurod1 mutant cochlea as early as E12.5 (arrow in B’) with premature expression in particular in the apex of the cochlea (C’) in comparison to wild-type (C). Fgf8 is expressed transiently in the prosensory domain in wild-type mice (arrow in A) and later is shown to be upregulated in the cochlea with a base-to-apex gradient (A,B,C). In newborn mice, Fgf8 is uniformly expressed in all inner hair cells almost along the entire length of the cochlea in wild-type mice (D, D’). In contrast, Neurod1 CKO mice display an increased expression level in the apex (E, F, H, I, J) and deviate from the single row labeling of only inner hair cells seen in the base (G). Two or more rows of inner hair cells are positive for Fgf8 and scattered single and multiple cells are interspersed among the multiple rows of outer hair cells which are also positive for Fgf8 (arrows in I). The apical tip shows up to three rows of Fgf8 positive cells (J). Radial sections through the apex of Fgf8 ISH reacted and Myo VIIa immunostained cochlea reveals co-localization of Fgf8 and Myo VIIa in inner as well as ‘ectopic inner hair cells’ scattered among outer hair cells (arrows in K–K”). SEM of P30 Neurod1 CKO mice reveals a normal organization of inner hair cells in a single row and three rows of outer hair cells in the base of Neurod1 CKO mice (L). In contrast, the apex shows inner hair cell sized stereocilia (arrows) interspersed among normal sized stereocilia bearing outer hair cells (arrowhead; M,N). Dotted line in H indicates border between normal and disorganized organ of Corti. IHC, inner hair cells; OHC, outer hair cells. Bar indicates 100 µm in A–F except D’; 10 µm in D’,G–J, K–K” and 1 µm in L–N.

Mentions: In mice, FGF3, FGF8 and FGF10 play a role in the early inductive events of the otic vesicle formation [8], [39], [46], [55], [56], [57], [58]. We observed Fgf8 expression in the delaminating sensory neuron in both wild-type and Neurod1 CKO mice as early as E10.5 (Fig. 4). Consistent with expression changes in Atoh1 and other downstream hair cell specific genes, Fgf8 was also expressed prematurely in the apex of the Neurod1 mutant mice (Fig. 8B’). Fgf8 was transiently expressed in the apex of wild-type mice but disappeared after E11.5 (compare Fig. 8A and B). However, absence of Neurod1 resulted in continued Fgf8 expression in the apex from E11.5 onward and thus resulted in premature and reversed expression pattern (Fig. 8A’, B’, C’). In contrast, in wild-type mice, Fgf8 expression started at E14.5 from the base of the cochlea progressing over time to the apex (Fig. 8C).


Neurod1 suppresses hair cell differentiation in ear ganglia and regulates hair cell subtype development in the cochlea.

Jahan I, Pan N, Kersigo J, Fritzsch B - PLoS ONE (2010)

Fgf8 misexpression correlates with formation of ‘ectopic inner hair cells’.In situ hybridization shows a persistent expression of Fgf8 in neurosensory precursors in Neurod1 mutant cochlea as early as E12.5 (arrow in B’) with premature expression in particular in the apex of the cochlea (C’) in comparison to wild-type (C). Fgf8 is expressed transiently in the prosensory domain in wild-type mice (arrow in A) and later is shown to be upregulated in the cochlea with a base-to-apex gradient (A,B,C). In newborn mice, Fgf8 is uniformly expressed in all inner hair cells almost along the entire length of the cochlea in wild-type mice (D, D’). In contrast, Neurod1 CKO mice display an increased expression level in the apex (E, F, H, I, J) and deviate from the single row labeling of only inner hair cells seen in the base (G). Two or more rows of inner hair cells are positive for Fgf8 and scattered single and multiple cells are interspersed among the multiple rows of outer hair cells which are also positive for Fgf8 (arrows in I). The apical tip shows up to three rows of Fgf8 positive cells (J). Radial sections through the apex of Fgf8 ISH reacted and Myo VIIa immunostained cochlea reveals co-localization of Fgf8 and Myo VIIa in inner as well as ‘ectopic inner hair cells’ scattered among outer hair cells (arrows in K–K”). SEM of P30 Neurod1 CKO mice reveals a normal organization of inner hair cells in a single row and three rows of outer hair cells in the base of Neurod1 CKO mice (L). In contrast, the apex shows inner hair cell sized stereocilia (arrows) interspersed among normal sized stereocilia bearing outer hair cells (arrowhead; M,N). Dotted line in H indicates border between normal and disorganized organ of Corti. IHC, inner hair cells; OHC, outer hair cells. Bar indicates 100 µm in A–F except D’; 10 µm in D’,G–J, K–K” and 1 µm in L–N.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2908541&req=5

pone-0011661-g008: Fgf8 misexpression correlates with formation of ‘ectopic inner hair cells’.In situ hybridization shows a persistent expression of Fgf8 in neurosensory precursors in Neurod1 mutant cochlea as early as E12.5 (arrow in B’) with premature expression in particular in the apex of the cochlea (C’) in comparison to wild-type (C). Fgf8 is expressed transiently in the prosensory domain in wild-type mice (arrow in A) and later is shown to be upregulated in the cochlea with a base-to-apex gradient (A,B,C). In newborn mice, Fgf8 is uniformly expressed in all inner hair cells almost along the entire length of the cochlea in wild-type mice (D, D’). In contrast, Neurod1 CKO mice display an increased expression level in the apex (E, F, H, I, J) and deviate from the single row labeling of only inner hair cells seen in the base (G). Two or more rows of inner hair cells are positive for Fgf8 and scattered single and multiple cells are interspersed among the multiple rows of outer hair cells which are also positive for Fgf8 (arrows in I). The apical tip shows up to three rows of Fgf8 positive cells (J). Radial sections through the apex of Fgf8 ISH reacted and Myo VIIa immunostained cochlea reveals co-localization of Fgf8 and Myo VIIa in inner as well as ‘ectopic inner hair cells’ scattered among outer hair cells (arrows in K–K”). SEM of P30 Neurod1 CKO mice reveals a normal organization of inner hair cells in a single row and three rows of outer hair cells in the base of Neurod1 CKO mice (L). In contrast, the apex shows inner hair cell sized stereocilia (arrows) interspersed among normal sized stereocilia bearing outer hair cells (arrowhead; M,N). Dotted line in H indicates border between normal and disorganized organ of Corti. IHC, inner hair cells; OHC, outer hair cells. Bar indicates 100 µm in A–F except D’; 10 µm in D’,G–J, K–K” and 1 µm in L–N.
Mentions: In mice, FGF3, FGF8 and FGF10 play a role in the early inductive events of the otic vesicle formation [8], [39], [46], [55], [56], [57], [58]. We observed Fgf8 expression in the delaminating sensory neuron in both wild-type and Neurod1 CKO mice as early as E10.5 (Fig. 4). Consistent with expression changes in Atoh1 and other downstream hair cell specific genes, Fgf8 was also expressed prematurely in the apex of the Neurod1 mutant mice (Fig. 8B’). Fgf8 was transiently expressed in the apex of wild-type mice but disappeared after E11.5 (compare Fig. 8A and B). However, absence of Neurod1 resulted in continued Fgf8 expression in the apex from E11.5 onward and thus resulted in premature and reversed expression pattern (Fig. 8A’, B’, C’). In contrast, in wild-type mice, Fgf8 expression started at E14.5 from the base of the cochlea progressing over time to the apex (Fig. 8C).

Bottom Line: Our data suggest that the long noted cross-regulation of Atoh1 expression by Neurog1 might actually be mediated in large part by Neurod1.We suggest that Neurod1 is regulated by both Neurog1 and Atoh1 and provides a negative feedback for either gene.Through this and other feedback, Neurod1 suppresses alternate fates of neurons to differentiate as hair cells and regulates hair cell subtypes.

View Article: PubMed Central - PubMed

Affiliation: Department of Biology, University of Iowa, Iowa City, Iowa, United States of America.

ABSTRACT

Background: At least five bHLH genes regulate cell fate determination and differentiation of sensory neurons, hair cells and supporting cells in the mammalian inner ear. Cross-regulation of Atoh1 and Neurog1 results in hair cell changes in Neurog1 mice although the nature and mechanism of the cross-regulation has not yet been determined. Neurod1, regulated by both Neurog1 and Atoh1, could be the mediator of this cross-regulation.

Methodology/principal findings: We used Tg(Pax2-Cre) to conditionally delete Neurod1 in the inner ear. Our data demonstrate for the first time that the absence of Neurod1 results in formation of hair cells within the inner ear sensory ganglia. Three cell types, neural crest derived Schwann cells and mesenchyme derived fibroblasts (neither expresses Neurod1) and inner ear derived neurons (which express Neurod1) constitute inner ear ganglia. The most parsimonious explanation is that Neurod1 suppresses the alternative fate of sensory neurons to develop as hair cells. In the absence of Neurod1, Atoh1 is expressed and differentiates cells within the ganglion into hair cells. We followed up on this effect in ganglia by demonstrating that Neurod1 also regulates differentiation of subtypes of hair cells in the organ of Corti. We show that in Neurod1 conditional mice there is a premature expression of several genes in the apex of the developing cochlea and outer hair cells are transformed into inner hair cells.

Conclusions/significance: Our data suggest that the long noted cross-regulation of Atoh1 expression by Neurog1 might actually be mediated in large part by Neurod1. We suggest that Neurod1 is regulated by both Neurog1 and Atoh1 and provides a negative feedback for either gene. Through this and other feedback, Neurod1 suppresses alternate fates of neurons to differentiate as hair cells and regulates hair cell subtypes.

Show MeSH
Related in: MedlinePlus