Purpose: The zone of polarizing activity regulatory sequence (ZRS) is an enhancer that regulates sonic hedgehog during embryonic limb development. Recently, mutations in a noncoding evolutionary conserved sequence 500 bp upstream of the ZRS, termed the pre-ZRS (pZRS), have been associated with polydactyly in dogs and humans. Here, we report the first case of triphalangeal thumb–polysyndactyly syndrome (TPT-PS) to be associated with mutations in this region and show via mouse enhancer assays how this mutation leads to ectopic expression throughout the developing limb bud. Methods: We used linkage analysis, whole-exome sequencing, Sanger sequencing, fluorescence in situ hybridization, multiplex ligation-dependent probe amplification, single-nucleotide polymorphism array, and a mouse transgenic enhancer assay. Results: Ten members of a TPT-PS family were included in this study. The mutation was linked to chromosome 7q36 (LOD score 3.0). No aberrations in the ZRS could be identified. A point mutation in the pZRS (chr7:156585476G>C; GRCh37/hg19) was detected in all affected family members. Functional characterization using a mouse transgenic enhancer essay showed extended ectopic expression dispersed throughout the entire limb bud (E11.5). Conclusion: Our work describes the first mutation in the pZRS to be associated with TPT-PS and provides functional evidence that this mutation leads to ectopic expression of this enhancer within the developing limb.

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Keywords congenital limb deformities, genetic enhancer elements, hedgehog protein, polydactyly, syndactyly
Persistent URL dx.doi.org/10.1038/gim.2018.18, hdl.handle.net/1765/113013
Journal Genetics in Medicine
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Potuijt, J.W.P. (Jacob W P), Baas, M, Sukenik-Halevy, R. (Rivka), Douben, H, Nguyen, P. (Picard), Venter, D, … de Klein, A. (2018). A point mutation in the pre-ZRS disrupts sonic hedgehog expression in the limb bud and results in triphalangeal thumb–polysyndactyly syndrome. Genetics in Medicine, 20(11), 1405–1413. doi:10.1038/gim.2018.18