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Math1 regulates development of the sensory epithelium in the mammalian cochlea

Nature Neuroscience volume 7, pages 1310–1318 (2004)Cite this article

Abstract

The transcription factor Math1 (encoded by the gene Atoh1, also called Math1) is required for the formation of mechanosensory hair cells in the inner ear; however, its specific molecular role is unknown. Here we show that absence of Math1 in mice results in a complete disruption of formation of the sensory epithelium of the cochlea, including the development of both hair cells and associated supporting cells. In addition, ectopic expression of Math1 in nonsensory regions of the cochlea is sufficient to induce the formation of sensory clusters that contain both hair cells and supporting cells. The formation of these clusters is dependent on inhibitory interactions mediated, most probably, through the Notch pathway, and on inductive interactions that recruit cells to develop as supporting cells through a pathway independent of Math1. These results show that Math1 functions in the developing cochlea to initiate both inductive and inhibitory signals that regulate the overall formation of the sensory epithelia.

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Figure 1: Expression of Math1 in the developing cochlea.
Figure 2: Cross-sections through the cochlear duct in wild-type and Math1-null mice at P0.
Figure 3: Expression of supporting cell markers in the cochleae of wild-type and Math1-null mice.
Figure 4: Supporting cell differentiation is disrupted in Math1-null mice.
Figure 5: Math1ORF expression induces formation of hair cells and supporting cells.
Figure 6: Cells that transiently express Math1ER can develop as supporting cells.
Figure 7: Components of the Notch signaling pathway are expressed in ectopic hair cells and inhibition of γ-secretase influences cell fate decisions by supporting cells.

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Acknowledgements

We thank R. Hertzano for technical advice; H. Zoghbi and K. Barald for the Math1 mutant mice; T. Friedman, R. Wenthold, N. Sans, A. Chitnis and R. Hertzano for reading an earlier version of this manuscript; M. Pryor for technical assistance; A. Capehart, L. Bianchi, M. Cohen-Salmon and C. Petit for antibodies; and M. Rivolta and A. McMahon for cDNAs. This research was supported by the Intramural Program at the National Institute on Deafness and Other Communication Disorders.

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Author notes

  1. Chad Woods and Mireille Montcouquiol: These authors contributed equally to this work.

Authors and Affiliations

  1. Section on Developmental Neuroscience, National Institute on Deafness and other Communication Disorders, National Institutes of Health, Porter Neuroscience Research Center, Building 35, Bethesda, 20892, Maryland, USA

    Chad Woods, Mireille Montcouquiol & Matthew W Kelley

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Supplementary information

Supplementary Fig. 1

Transfection with Math1ORF induces hair cell formation in the GER. (a) Low magnification view of an E13 explant transfected with Math1ORF after 6 DIV. Hair cells located in the sensory epithelium and in KO are labeled with an antibody for Myosin VI (red). Note the presence of ectopic hair cells in KO (arrows). (b) The same image as in a, except with cells transfected with Math1ORF labeled in green. (c) Merged image of panels a and b. (d) The same image as in a, however each ectopic hair cell as been circled in white. (e) The same image as in b, with each transfected cell circled. (f) The same image as in c, except with circles. Only three transfected cells (purple circles) are not positive for Myosin 6. (g) High magnification view of Myosin VI-positive cells (red) in an E13 explant transfected as in a. The edge of the sensory epithelium is located in the upper left hand corner of the image. Ectopic hair cells are present in KO. (h) The same image as in g, except with cells transfected with Math1ORF labeled in green. (i) Merged image of panels h and i. (j) The same image as in g, except with ecotpic hair cells circled in white. (k) The same image as in h, except with transfected cells circled in white. (l) The same image as in i, except with circles. Only a single transfected cell (purple circle) is not positive for Myosin VI. Scale bar in a (same in b-f), 200 µm. Scale bar in g (same in h-l), 40 µm. (PDF 432 kb)

Supplementary Fig. 2

Transfection with Math1ER induces hair cells only in the presence of tamoxifen. (a) Low magnification image of an E13 explant transfected with Math1ERand maintained for 6 DIV in the absence of tamoxifen. Myosin VI-positive hair cells in the sensory epithelium are labeled in red. (b) The same image as in a. Cells expressing Math1ER are illustrated in green. (c) Merged image of a and b. Note that none of the transfected cells in KO have developed as hair cells. (d) Low magnification image of an E13 explant transfected with Math1ERand maintained for 6 DIV in the absence of tamoxifen. Jagged1-positive supporting cells in the sensory epithelium are labeled in red. (e) The same image as in d. Cells expressing Math1ER are illustrated in green. (f) Merged image of d and e. Note that none of the transfected cells in KO have developed as supporting cells. (g) Low magnification image of an E13 explant transfected with Math1ER and maintained for 6 DIV in the presence of 15 nM tamoxifen. Myosin VI-positive hair cells (red) are present in both the sensory epithelium and KO. (h) The same image as in g. Cells expressing Math1ER are illustrated in green. (i) Merged image of g and h. Note that transfected cells in KO are also positive for Myosin VI, indicating that these cells have developed as hair cells. (j) Magnified image of KO from the sample illustrated in g. Transfected cells that have developed as hair cells are circled. Note that many of the green cells express Myosin VI at the lumenal surface. See Figure 6a-d for a higher magnification view. Scale bar in b (same in a, c-f), 100 µm. Scale bar in h (same in g,i) 200 µm. Scale bar in j, 200 µm. (PDF 364 kb)

Supplementary Fig. 3

Detection of Math1-promoter activity using anti-β-galactosidase. Confocal image of the lumenal surface of the sensory epithelium from an E13.5 explant established from a Math1+/bgal embryo and maintained for 6 DIV. Inner (IHC) and outer (numbered) hair cells are positive for β-galactosidase, while supporting cells are negative. Scale bar, 20 µm. (PDF 119 kb)

Supplementary Fig. 4

Expression of Math1ORF induces expression of supporting cell markers in explants from Math1-nulls. (a) Low magnification image of a large region of Math1ORF-transfected cells (green). Jag1 (red) is expressed in cells surrounding the transfected cells. (b) High magnification image of a small cluster of Math1ORF-transfected cells (green). Cells surrounding the cluster are positive for Jag1 (red). (c) High magnification image of a small cluster of Math1ORF-transfected cells (green). Surrounding cells express Otogelin (red). Scale bar in a 100 µ. Scale bar in b (same in c), 20 µm. (PDF 229 kb)

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Woods, C., Montcouquiol, M. & Kelley, M. Math1 regulates development of the sensory epithelium in the mammalian cochlea. Nat Neurosci 7, 1310–1318 (2004). https://doi.org/10.1038/nn1349

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