A genome-scale analysis of the cis-regulatory circuitry underlying sonic hedgehog-mediated patterning of the mammalian limb
cis- 1 Department of Molecular and Cellular Biology, Harvard University, Cambridge, Massachusetts 02138, USA;
- 2 Department of Biostatistics, Johns Hopkins Bloomberg School of Public Health, Baltimore, Maryland 21205, USA;
- 3 Department of Statistics, Stanford University, Stanford, California, 94305, USA;
- 4 Harvard Stem Cell Institute, Harvard University, Cambridge, Massachusetts 02138, USA
Abstract
Sonic hedgehog (Shh) signals via Gli transcription factors to direct digit number and identity in the vertebrate limb. We characterized the Gli-dependent cis-regulatory network through a combination of whole-genome chromatin immunoprecipitation (ChIP)-on-chip and transcriptional profiling of the developing mouse limb. These analyses identified ∼5000 high-quality Gli3-binding sites, including all known Gli-dependent enhancers. Discrete binding regions exhibit a higher-order clustering, highlighting the complexity of cis-regulatory interactions. Further, Gli3 binds inertly to previously identified neural-specific Gli enhancers, demonstrating the accessibility of their cis-regulatory elements. Intersection of DNA binding data with gene expression profiles predicted 205 putative limb target genes. A subset of putative cis-regulatory regions were analyzed in transgenic embryos, establishing Blimp1 as a direct Gli target and identifying Gli activator signaling in a direct, long-range regulation of the BMP antagonist Gremlin. In contrast, a long-range silencer cassette downstream from Hand2 likely mediates Gli3 repression in the anterior limb. These studies provide the first comprehensive characterization of the transcriptional output of a Shh-patterning process in the mammalian embryo and a framework for elaborating regulatory networks in the developing limb.
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Footnotes
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↵5 Present address: Section of Molecular Cell and Developmental Biology, Institute for Cellular and Molecular Biology, The University of Texas at Austin, 2500 Speedway, Austin, TX 78712, USA.
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↵6 Corresponding author.
↵6 E-MAIL mcmahon{at}mcb.harvard.edu; FAX (617) 496-3763.
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Supplemental material is available at http://www.genesdev.org.
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Article is online at http://www.genesdev.org/cgi/doi/10.1101/gad.1693008.
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- Received May 8, 2008.
- Accepted August 5, 2008.
- Copyright © 2008, Cold Spring Harbor Laboratory Press
