title: Long-range conserved non-coding SHOX sequences regulate expression in developing chicken limb and are associated with short stature phenotypes in human patients.
abstract: Defects in long-range regulatory elements have recently emerged as previously underestimated factors in the genesis of human congenital disorders. Léri-Weill dyschondrosteosis is a dominant skeletal malformation syndrome caused by mutations in the short stature homeobox gene SHOX. We have analysed four families with Léri-Weill dyschondrosteosis with deletions in the pseudoautosomal region but still with an intact SHOX coding region. Using fluorescence in situ hybridization and single nucleotide polymorphism studies, we identified an interval of approximately 200 kb that was deleted in all tested affected family members but retained in the unaffected members and in 100 control individuals. Comparative genomic analysis of this interval revealed eight highly conserved non-genic elements between 48 and 215 kb downstream of the SHOX gene. As mice do not have a Shox gene, we analysed the enhancer potential in chicken embryos using a green fluorescent protein reporter construct driven by the beta-globin promoter, by in ovo electroporation of the limb bud. We observed cis-regulatory activity in three of the eight non-genic elements in the developing limbs arguing for an extensive control region of this gene. These findings are consistent with the idea that the deleted region in the affected families contains several distinct elements that regulate Shox expression in the developing limb. Furthermore, the deletion of these elements in humans generates a phenotype apparently undistinguishable to those patients identified with mutations in the SHOX coding region and, for the first time, demonstrates the potential of an in vivo assay in chicken to monitor putative enhancer activity in relation to human disease.
authors.affiliation: Department of Molecular Human Genetics, University of Heidelberg, Im Neuenheimer Feld 366, 69120 Heidelberg, Germany.
authors.forename: Nitin
authors.initials: N
authors.lastname: Sabherwal
authors.forename: Fiona
authors.initials: F
authors.lastname: Bangs
authors.forename: Ralph
authors.initials: R
authors.lastname: Röth
authors.forename: Birgit
authors.initials: B
authors.lastname: Weiss
authors.forename: Karin
authors.initials: K
authors.lastname: Jantz
authors.forename: Eva
authors.initials: E
authors.lastname: Tiecke
authors.forename: Georg K
authors.initials: GK
authors.lastname: Hinkel
authors.forename: Christiane
authors.initials: C
authors.lastname: Spaich
authors.forename: Berthold P
authors.initials: BP
authors.lastname: Hauffa
authors.forename: Hetty
authors.initials: H
authors.lastname: van der Kamp
authors.forename: Johannes
authors.initials: J
authors.lastname: Kapeller
authors.forename: Cheryll
authors.initials: C
authors.lastname: Tickle
authors.forename: Gudrun
authors.initials: G
authors.lastname: Rappold
mesh_terms: D000015:Abnormalities, Multiple / Q000235:genetics*; D000293:Adolescent; D000328:Adult; D000368:Aged; D000818:Animals; D001483:Base Sequence; D001827:Body Height / Q000235:genetics; D002642:Chick Embryo; D002648:Child; D002874:Chromosome Mapping; D017124:Conserved Sequence / Q000235:genetics*; D004252:DNA Mutational Analysis; D017931:DNA Primers; D021901:DNA, Intergenic / Q000235:genetics*; D018274:Electroporation; D005260:Female; D040461:Gene Components; D005786:Gene Expression Regulation; D023281:Genomics / Q000379:methods; D006614:Hindlimb / Q000196:embryology / Q000378:metabolism*; D018398:Homeodomain Proteins / Q000235:genetics*; D006801:Humans; D017404:In Situ Hybridization, Fluorescence; D008297:Male; D008875:Middle Aged; D008969:Molecular Sequence Data; D010009:Osteochondrodysplasias / Q000235:genetics*; D010375:Pedigree; D020641:Polymorphism, Single Nucleotide / Q000235:genetics; D017384:Sequence Deletion / Q000235:genetics*; D000074122:Short Stature Homeobox Protein; D013577:Syndrome
chemical_list: D017931:DNA Primers; D021901:DNA, Intergenic; D018398:Homeodomain Proteins; C106071:SHOX protein, human; D000074122:Short Stature Homeobox Protein