A unique stylopod patterning mechanism by Shox2-controlled osteogenesis

Development. 2016 Jul 15;143(14):2548-60. doi: 10.1242/dev.138750. Epub 2016 Jun 10.


Vertebrate appendage patterning is programmed by Hox-TALE factor-bound regulatory elements. However, it remains unclear which cell lineages are commissioned by Hox-TALE factors to generate regional specific patterns and whether other Hox-TALE co-factors exist. In this study, we investigated the transcriptional mechanisms controlled by the Shox2 transcriptional regulator in limb patterning. Harnessing an osteogenic lineage-specific Shox2 inactivation approach we show that despite widespread Shox2 expression in multiple cell lineages, lack of the stylopod observed upon Shox2 deficiency is a specific result of Shox2 loss of function in the osteogenic lineage. ChIP-Seq revealed robust interaction of Shox2 with cis-regulatory enhancers clustering around skeletogenic genes that are also bound by Hox-TALE factors, supporting a lineage autonomous function of Shox2 in osteogenic lineage fate determination and skeleton patterning. Pbx ChIP-Seq further allowed the genome-wide identification of cis-regulatory modules exhibiting co-occupancy of Pbx, Meis and Shox2 transcriptional regulators. Integrative analysis of ChIP-Seq and RNA-Seq data and transgenic enhancer assays indicate that Shox2 patterns the stylopod as a repressor via interaction with enhancers active in the proximal limb mesenchyme and antagonizes the repressive function of TALE factors in osteogenesis.

Keywords: Limb; Patterning; Shox2; Skeleton; Stylopod.

MeSH terms

  • Animals
  • Base Sequence
  • Binding Sites / genetics
  • Body Patterning* / genetics
  • Cell Lineage
  • Core Binding Factor Alpha 1 Subunit / metabolism
  • Enhancer Elements, Genetic
  • Extremities / embryology*
  • Gene Deletion
  • Gene Expression Regulation, Developmental
  • Genome
  • Homeodomain Proteins / genetics
  • Homeodomain Proteins / metabolism*
  • Mesenchymal Stem Cells / cytology
  • Mesenchymal Stem Cells / metabolism
  • Mice
  • Models, Biological
  • Nucleotide Motifs / genetics
  • Osteogenesis* / genetics
  • Protein Binding


  • Core Binding Factor Alpha 1 Subunit
  • Homeodomain Proteins
  • Shox2 protein, mouse