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Analysis of Zfhx1a mutant mice reveals palatal shelf contact-independent medial edge epithelial differentiation during palate fusion

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Abstract

Cleft palate is a common birth defect that involves disruptions in multiple developmental steps such as growth, differentiation, elevation, and fusion. Medial edge epithelial (MEE) differentiation is essential for palate fusion. An important question is whether the MEE differentiation that occurs during fusion is induced by palate shelf contact or is programmed intrinsically by the palate shelf itself. Here, we report that the loss of Zfhx1a function in mice leads to a cleft palate phenotype that is mainly attributable to a delay in palate elevation. Zfhx1a encodes a transcription regulatory protein that modulates several signaling pathways including those activated by members of the transforming growth factor-β (TGF-β) superfamily. Loss of Zfhx1a function in mice leads to a complete cleft palate with 100% penetrance. Zfhx1a mutant palatal shelves display normal cell differentiation and proliferation and are able to fuse in an in vitro culture system. The only defect detected was a delay of 24–48 h in palatal shelf elevation. Using the Zfhx1a mutant as a model, we studied the relationship between MEE differentiation and palate contact/adhesion. We found that down-regulation of Jag2 expression in the MEE cells, a key differentiation event establishing palate fusion competence, was independent of palate contact/adhesion. Moreover, the expression of several key factors essential for fusion, such as TGF-β3 and MMP13, was also down-regulated at embryonic stage 16.5 in a contact-independent manner, suggesting that differentiation of the medial edge epithelium was largely programmed through an intrinsic mechanism within the palate shelf.

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Acknowledgements

We thank Dr. Yiping Chen at the Ohio State University, Dr. Rulang Jiang at the University of Rochester, and Dr. Mary B. Goldring at Harvard Medical School for providing in situ probes, and Dr. Dennis Warner for comments on the manuscript.

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Correspondence to Douglas S. Darling or Jixiang Ding.

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Correspondence should be addressed to Dr. Douglas S. Darling and Dr. Jixiang Ding.

This work is supported by the COBRE program of the National Center for Research Resource (P20RR017702 to the University of Louisville Birth Defects Center) and by research grants (EY017869 and DE13614 to D.D.; DE015565 and DE016845 to J.D.) from the National Institutes of Health, USA.

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supplemental figure

Whole-mount in situ hybridization followed by sectioning and showing expression of Zfhx1a mRNA in mouse secondary palate on E12.5 and E13.5. a, b Zfhx1a mRNA was detected in the mouse secondary palate on E12.5 (arrows in a), and the expression was decreased on E13.5 (arrows in b). c, d Sections of whole-mount in situ hybridization at E12.5 (c) and E13.5 (d) showing the expression of Zfhx1a mRNA only in mesenchymal cells (arrows). Bars 200 μm (a, b), 100 μm (c, d) (GIF 443 kb)

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Jin, JZ., Li, Q., Higashi, Y. et al. Analysis of Zfhx1a mutant mice reveals palatal shelf contact-independent medial edge epithelial differentiation during palate fusion. Cell Tissue Res 333, 29–38 (2008). https://doi.org/10.1007/s00441-008-0612-x

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