title: Direct functional consequences of ZRS enhancer mutation combine with secondary long range SHH signalling effects to cause preaxial polydactyly.
abstract: Sonic hedgehog (SHH) plays a central role in patterning numerous embryonic tissues including, classically, the developing limb bud where it controls digit number and identity. This study utilises the polydactylous Silkie (Slk) chicken breed, which carries a mutation in the long range limb-specific regulatory element of SHH, the ZRS. Using allele specific SHH expression analysis combined with quantitative protein analysis, we measure allele specific changes in SHH mRNA and concentration of SHH protein over time. This confirms that the Slk ZRS enhancer mutation causes increased SHH expression in the posterior leg mesenchyme. Secondary consequences of this increased SHH signalling include increased FGF pathway signalling and growth as predicted by the SHH/GREM1/FGF feedback loop and the Growth/Morphogen models. Manipulation of Hedgehog, FGF signalling and growth demonstrate that anterior-ectopic expression of SHH and induction of preaxial polydactyly is induced secondary to increased SHH signalling and Hedgehog-dependent growth directed from the posterior limb. We predict that increased long range SHH signalling acts in combination with changes in activation of SHH transcription from the Slk ZRS allele. Through analysis of the temporal dynamics of anterior SHH induction we predict a gene regulatory network which may contribute to activation of anterior SHH expression from the Slk ZRS.
authors.affiliation: Division of Developmental Biology, The Roslin Institute and R(D)SVS, University of Edinburgh, Easter Bush, Midlothian EH25 9RG, UK.
authors.forename: Edward J
authors.initials: EJ
authors.lastname: Johnson
authors.affiliation: Division of Developmental Biology, The Roslin Institute and R(D)SVS, University of Edinburgh, Easter Bush, Midlothian EH25 9RG, UK.
authors.forename: David M
authors.initials: DM
authors.lastname: Neely
authors.affiliation: Division of Genetics and Genomics, The Roslin Institute and R(D)SVS, University of Edinburgh, Easter Bush, Midlothian EH25 9RG, UK.
authors.forename: Ian C
authors.initials: IC
authors.lastname: Dunn
authors.affiliation: Division of Developmental Biology, The Roslin Institute and R(D)SVS, University of Edinburgh, Easter Bush, Midlothian EH25 9RG, UK. Electronic address: megan.davey@roslin.ed.ac.uk.
authors.forename: Megan G
authors.initials: MG
authors.lastname: Davey
mesh_terms: D000818:Animals; D002642:Chick Embryo; D019295:Computational Biology; D017931:DNA Primers / Q000235:genetics; D024202:Electrophoretic Mobility Shift Assay; D004742:Enhancer Elements, Genetic / Q000235:genetics*; D005346:Fibroblast Growth Factors / Q000378:metabolism; D020869:Gene Expression Profiling; D018507:Gene Expression Regulation, Developmental / Q000502:physiology*; D053263:Gene Regulatory Networks / Q000235:genetics; D053823:Hedgehog Proteins / Q000235:genetics* / Q000502:physiology; D017403:In Situ Hybridization; D008954:Models, Biological; D009154:Mutation / Q000235:genetics*; D017689:Polydactyly / Q000235:genetics*; D012150:Polymorphism, Restriction Fragment Length; D060888:Real-Time Polymerase Chain Reaction; D015398:Signal Transduction / Q000502:physiology*
chemical_list: D017931:DNA Primers; D053823:Hedgehog Proteins; D005346:Fibroblast Growth Factors
keywords: Cyclopamine; HOXA13; Polydactyly; SAG; Sonic hedgehog; ZRS