Plastic Surgery Research Council
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Presenter: Lisa Gferer, MD
Co-Authors: Kamel G; Liao EC
Massachusetts General Hospital

Background: Cleft lip and/or palate (CL/P) is the most frequent craniofacial birth defect. Transcription factor IRF6 has been confirmed as a key locus for syndromic and non-syndromic CL/P. In order to understand irf6 function during palate development and its role in cleft malformation, we are generating transgenic irf6 reporter line for use in mechanistic analysis.

Methods: A 7.185 kb zebrafish irf6 promoter sequence was amplified from the irf6 bacterial artificial chromosome. The Tg:irf6:eGFP transgenic animal was generated using Gateway system, with Tol2 transposase mediating germline integration. Progeny of stable transgenic lines were analyzed by compound and confocal microscopy.

Results: Expression of irf6 was detected at single cell stage, confirming the presence of irf6 as a maternal transcript (Figure. 1). Irf6 expression continued throughout gastrulation, then localized in the otic placode and migrating cranial neural crest cells. Later in embryogenesis, irf6 expression was found along pharyngeal arches and facial prominences, including the maxillary / trabeculae extensions. By 72 hpf, irf6 was noted in the lower jaw, ceratohyal and ceratobranchials. Importantly, in addition to expression in epithelial tissues, irf6 expression was found in chondrocytes forming the jaw structures.

Discussion: We have generated a stable Tg:irf6:eGFP transgenic animal model that faithfully recapitulates irf6 expression. The unexpected observation of irf6 expression in chondrocytes emphasizes its importance in the development of craniofacial mesenchyme and explains why disruption results in CL/P phenotypes. Mating of the irf6 and sox10 transgenic lines will allow us to identify the subset of cranial neural crest cells that are specifically important to palate development.

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