Identification of spontaneous mutations within the long-range limb-specific Sonic hedgehog enhancer (ZRS) that alter Sonic hedgehog expression in the chicken limb mutants oligozeugodactyly and silkie breed

Authors

  • Sarah A. Maas,

    1. Department of Biomolecular Chemistry, University of Wisconsin-Madison, Madison, Wisconsin
    2. Department of Anatomy, University of Wisconsin-Madison, Madison, Wisconsin
    Current affiliation:
    1. Adnexus, a Bristol-Myers Squibb R&D Company, 100 Beaver St., Waltham, MA 02453
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    • Sarah A. Maas and Takayuki Suzuki contributed equally to this article.

  • Takayuki Suzuki,

    1. Department of Anatomy, University of Wisconsin-Madison, Madison, Wisconsin
    Current affiliation:
    1. Division of Biological Science, Nagoya University, Furo-cho, Chikusa-ku, Nagoya 464-8602, Japan and Japan Science and Technology Agency (JST), Saitama 332-0012, Japan
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    • Sarah A. Maas and Takayuki Suzuki contributed equally to this article.

  • John F. Fallon

    Corresponding author
    1. Department of Anatomy, University of Wisconsin-Madison, Madison, Wisconsin
    • Department of Cell and Regenerative Biology, University of Wisconsin-Madison, 1300 University Ave., Madison, WI 53706
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Abstract

The evolutionarily conserved, non-coding ∼800-base-pair (bp) zone of polarizing activity (ZPA) regulatory sequence (ZRS) controls Shh expression in the posterior limb. We report that the chicken mutant oligozeugodactyly (ozd), which lacks limb Shh expression, has a large deletion within the ZRS. Furthermore, the preaxial polydactylous, Silkie Breed chicken, which develops ectopic anterior limb Shh expression, has a single bp change within the ZRS. Using an in vivo reporter assay to examine enhancer function in the chick limb, we demonstrate that the wild-type ZRS drives β-galactosidase reporter expression in the ZPA of both wild-type and ozd limbs. The Silkie ZRS drives β-galactosidase in both posterior and anterior Shh domains in wild-type limb buds. These results support the hypothesis that the ZRS integrates positive and negative prepatterned regulatory inputs in the chicken model system and demonstrate the utility of the chicken limb as an efficient genetic system for gene regulatory studies. Developmental Dynamics 240:1212–1222, 2011. © 2011 Wiley-Liss, Inc.

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