An Evolutionarily Conserved Enhancer Regulates Bmp4 Expression in Developing Incisor and Limb Bud

Dolrudee Jumlongras; Salil A. Lachke; Daniel J. O’Connell; Anton Aboukhalil; Xiao Li; Sung E. Choe; Joshua W. K. Ho; Annick Turbe-Doan; Erin A. Robertson; Bjorn R. Olsen; Martha L. Bulyk; Brad A. Amendt; Richard L. Maas

doi:10.1371/journal.pone.0038568

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An Evolutionarily Conserved Enhancer Regulates Bmp4 Expression in Developing Incisor and Limb Bud

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An Evolutionarily Conserved Enhancer Regulates Bmp4 Expression in Developing Incisor and Limb Bud

Dolrudee Jumlongras, Salil A. Lachke, Daniel J. O’Connell, Anton Aboukhalil, Xiao Li, Sung E. Choe, Joshua W. K. Ho, Annick Turbe-Doan, Erin A. Robertson, Bjorn R. Olsen, …

PloS one, Vol.7(6), pp.e38568-e38568

06/12/2012

DOI: 10.1371/journal.pone.0038568

PMCID: PMC3373496

PMID: 22701669

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Abstract

To elucidate the transcriptional regulation of Bmp4 expression during organogenesis, we used phylogenetic footprinting and transgenic reporter analyses to identify Bmp4 c is - r egulatory m odules (CRMs). These analyses identified a regulatory region located ∼46 kb upstream of the mouse Bmp4 transcription start site that had previously been shown to direct expression in lateral plate mesoderm. We refined this regulatory region to a 396-bp minimal enhancer, and show that it recapitulates features of endogenous Bmp4 expression in developing mandibular arch ectoderm and incisor epithelium during the initiation-stage of tooth development. In addition, this enhancer directs expression in the apical ectodermal ridge (AER) of the developing limb and in anterior and posterior limb mesenchyme. Transcript profiling of E11.5 mouse incisor dental lamina, together with protein binding microarray (PBM) analyses, allowed identification of a conserved DNA binding motif in the Bmp4 enhancer for Pitx homeoproteins, which are also expressed in the developing mandibular and incisor epithelium. In vitro electrophoretic mobility shift assays (EMSA) and in vivo transgenic reporter mutational analyses revealed that this site supports Pitx binding and that the site is necessary to recapitulate aspects of endogenous Bmp4 expression in developing craniofacial and limb tissues. Finally, Pitx2 chromatin immunoprecipitation (ChIP) demonstrated direct binding of Pitx2 to this Bmp4 enhancer site in a dental epithelial cell line. These results establish a direct molecular regulatory link between Pitx family members and Bmp4 gene expression in developing incisor epithelium.

Biology

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Title: Subtitle: An Evolutionarily Conserved Enhancer Regulates Bmp4 Expression in Developing Incisor and Limb Bud
Creators: Dolrudee Jumlongras - Brigham and Women's Hospital
Salil A. Lachke - University of Delaware
Daniel J. O’Connell - Brigham and Women's Hospital
Anton Aboukhalil - Brigham and Women's Hospital
Xiao Li - Texas A&M Health Science Center
Sung E. Choe - Brigham and Women's Hospital
Joshua W. K. Ho - Brigham and Women's Hospital
Annick Turbe-Doan - Brigham and Women's Hospital
Erin A. Robertson - Brigham and Women's Hospital
Bjorn R. Olsen - Harvard University
Martha L. Bulyk - Brigham and Women's Hospital
Brad A. Amendt - Texas A&M Health Science Center
Richard L. Maas - Brigham and Women's Hospital
Resource Type: Journal article
Publication Details: PloS one, Vol.7(6), pp.e38568-e38568
DOI: 10.1371/journal.pone.0038568
PMID: 22701669
PMCID: PMC3373496
NLM abbreviation: PLoS One
ISSN: 1932-6203
eISSN: 1932-6203
Publisher: Public Library of Science
Alternative title: Regulation of Bmp4 Expression during Organogenesis
Language: English
Date published: 06/12/2012
Academic Unit: Orthodontics; Anatomy and Cell Biology; Craniofacial Anomalies Research Center; Dental Research
Record Identifier: 9984284452102771

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