Differential gene expression in mice with misexpression of Six2 associated with frontonasal dysplasia

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2012-08

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[Honolulu] : [University of Hawaii at Manoa], [August 2012]

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Abstract

We have previously described the Br mutant mouse displaying heritable frontonasal dysplasia. Linkage analysis mapped the mutation near the homeobox transcription factor Six2, normally expressed in the facial and metanephric mesenchyme during development. The purpose of this study is to determine expression patterns of Six2, as well as possible downstream targets of Six2, in the developing midface. The three sets of facial prominences (medial, lateral, and maxillary) from embryos at gestational day 11.5 (E11.5) were dissected and RNA extracted for qRT-PCR assays and microarray analysis. Medial nasal prominences (MNP) and E13.5 kidneys were also taken for cell culture. Results from qRT-PCR indicated Six2 expression is highest in the MNP at E11.5 and demonstrated haploinsufficient down-regulation in each of the three facial prominence sets in the Br mouse at this age. Microarray results suggested the misregulation of several genes in the Br midface, including Six3, another member of the Six family of transcription factors. MNP and kidney qRT-PCR and immunohistochemistry for Six3 substantiated its upregulation in the microarray. Additionally, Shh and Flrt2 were confirmed misexpressed in the developing midface, both of which have been previously shown to play critical roles in craniofacial development. RNA interference on Six2 in E11.5 MNP and E13.5 embryonic kidney cultures did not demonstrate misexpression of Six3, suggesting Six2 is not a direct regulator of Six3 and that the Br mutation may be located in a transcriptional activation domain of Six2 that also inhibits Six3 transcription. Further sequencing analysis will be needed to confirm the type and location of the Br mutation. This work was supported, in part, by NIH R01DK064752 & NCRR 5P20RR024206.

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Ph.D. University of Hawaii at Manoa 2012.
Includes bibliographical references.

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Six2, frontonasal dysplasia, Br mouse, Six3

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Theses for the degree of Doctor of Philosophy (University of Hawaii at Manoa). Biomedical Sciences (Physiology).

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