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Highly conserved non-coding elements on either side of SOX9 associated with Pierre Robin sequence


Pierre Robin sequence (PRS) is an important subgroup of cleft palate. We report several lines of evidence for the existence of a 17q24 locus underlying PRS, including linkage analysis results, a clustering of translocation breakpoints 1.06–1.23 Mb upstream of SOX9, and microdeletions both 1.5 Mb centromeric and 1.5 Mb telomeric of SOX9. We have also identified a heterozygous point mutation in an evolutionarily conserved region of DNA with in vitro and in vivo features of a developmental enhancer. This enhancer is centromeric to the breakpoint cluster and maps within one of the microdeletion regions. The mutation abrogates the in vitro enhancer function and alters binding of the transcription factor MSX1 as compared to the wild-type sequence. In the developing mouse mandible, the 3-Mb region bounded by the microdeletions shows a regionally specific chromatin decompaction in cells expressing Sox9. Some cases of PRS may thus result from developmental misexpression of SOX9 due to disruption of very-long-range cis-regulatory elements.

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Figure 1: Family trees and highly conserved noncoding element (HCNE) rearrangements.
Figure 2: HCNEs at the PRS1 locus have tissue-specific enhancer activity.
Figure 3: HCNE-F2 has enhancer activity.
Figure 4: Chromatin characteristics of the region around SOX9.

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NCBI Reference Sequence


  1. Robin, P. A fall of the base of the tongue considered as a new cause of nasopharyngeal respiratory impairment: Pierre Robin sequence, a translation. 1923. Plast. Reconstr. Surg. 93, 1301–1303 (1994).

    Article  CAS  Google Scholar 

  2. Etchevers, H.C., Amiel, J. & Lyonnet, S. Molecular bases of human neurocristopathies. Adv. Exp. Med. Biol. 589, 213–234 (2006).

    Article  CAS  Google Scholar 

  3. Vintiner, G.M., Temple, I.K., Middleton-Price, H.R., Baraitser, M. & Malcolm, S. Genetic and clinical heterogeneity of Stickler syndrome. Am. J. Med. Genet. 41, 44–48 (1991).

    Article  CAS  Google Scholar 

  4. Jamshidi, N. et al. Isolated Robin sequence associated with a balanced t(2;17) chromosomal translocation. J. Med. Genet. 41, e1 (2004).

    Article  CAS  Google Scholar 

  5. Jakobsen, L.P. et al. Pierre Robin sequence may be caused by dysregulation of SOX9 and KCNJ2. J. Med. Genet. 44, 381–386 (2007).

    Article  CAS  Google Scholar 

  6. Ahituv, N., Prabhakar, S., Poulin, F., Rubin, E.M. & Couronne, O. Mapping cis-regulatory domains in the human genome using multi-species conservation of synteny. Hum. Mol. Genet. 14, 3057–3063 (2005).

    Article  CAS  Google Scholar 

  7. Dermitzakis, E.T., Reymond, A. & Antonarakis, S.E. Conserved non-genic sequences—an unexpected feature of mammalian genomes. Nat. Rev. Genet. 6, 151–157 (2005).

    Article  CAS  Google Scholar 

  8. Woolfe, A. et al. Highly conserved non-coding sequences are associated with vertebrate development. PLoS Biol. 3, e7 (2005).

    Article  Google Scholar 

  9. Velagaleti, G.V. et al. Position effects due to chromosome breakpoints that map approximately 900 Kb upstream and approximately 1.3 Mb downstream of SOX9 in two patients with campomelic dysplasia. Am. J. Hum. Genet. 76, 652–662 (2005).

    Article  CAS  Google Scholar 

  10. Cai, J. et al. Gene expression in pharyngeal arch 1 during human embryonic development. Hum. Mol. Genet. 14, 903–912 (2005).

    Article  CAS  Google Scholar 

  11. Satokata, I. & Maas, R. Msx1 deficient mice exhibit cleft palate and abnormalities of craniofacial and tooth development. Nat. Genet. 6, 348–356 (1994).

    Article  CAS  Google Scholar 

  12. Cirulli, E.T. & Goldstein, D.B. In vitro assays fail to predict in vivo effects of regulatory polymorphisms. Hum. Mol. Genet. 16, 1931–1939 (2007).

    Article  CAS  Google Scholar 

  13. Morey, C., Da Silva, N.R., Perry, P. & Bickmore, W.A. Nuclear reorganisation and chromatin decondensation are conserved, but distinct, mechanisms linked to Hox gene activation. Development 134, 909–919 (2007).

    Article  CAS  Google Scholar 

  14. Bagheri-Fam, S., Ferraz, C., Demaille, J., Scherer, G. & Pfeifer, D. Comparative genomics of the SOX9 region in human and Fugu rubripes: conservation of short regulatory sequence elements within large intergenic regions. Genomics 78, 73–82 (2001).

    Article  CAS  Google Scholar 

  15. Bagheri-Fam, S. et al. Long-range upstream and downstream enhancers control distinct subsets of the complex spatiotemporal Sox9 expression pattern. Dev. Biol. 291, 382–397 (2006).

    Article  CAS  Google Scholar 

  16. Bien-Willner, G.A., Stankiewicz, P. & Lupski, J.R. SOX9cre1, a cis-acting regulatory element located 1.1 Mb upstream of SOX9, mediates its enhancement through the SHH pathway. Hum. Mol. Genet. 16, 1143–1156 (2007).

    Article  CAS  Google Scholar 

  17. Suzuki, T., Sakai, D., Osumi, N., Wada, H. & Wakamatsu, Y. Sox genes regulate type 2 collagen expression in avian neural crest cells. Dev. Growth Differ. 48, 477–486 (2006).

    Article  CAS  Google Scholar 

  18. Akiyama, H. et al. Interactions between Sox9 and beta-catenin control chondrocyte differentiation. Genes Dev. 18, 1072–1087 (2004).

    Article  CAS  Google Scholar 

  19. Bi, W., Deng, J.M., Zhang, Z., Behringer, R.R. & de Crombrugghe, B. Sox9 is required for cartilage formation. Nat. Genet. 22, 85–89 (1999).

    Article  CAS  Google Scholar 

  20. Holder-Espinasse, M. et al. Pierre Robin sequence: a series of 117 consecutive cases. J. Pediatr. 139, 588–590 (2001).

    Article  CAS  Google Scholar 

  21. Mori-Akiyama, Y., Akiyama, H., Rowitch, D.H. & de Crombrugghe, B. Sox9 is required for determination of the chondrogenic cell lineage in the cranial neural crest. Proc. Natl. Acad. Sci. USA 100, 9360–9365 (2003).

    Article  CAS  Google Scholar 

  22. Andelfinger, G. et al. KCNJ2 mutation results in Andersen syndrome with sex-specific cardiac and skeletal muscle phenotypes. Am. J. Hum. Genet. 71, 663–668 (2002).

    Article  CAS  Google Scholar 

  23. Wagner, T. et al. Autosomal sex reversal and campomelic dysplasia are caused by mutations in and around the SRY-related gene SOX9. Cell 79, 1111–1120 (1994).

    Article  CAS  Google Scholar 

  24. Kleinjan, D.A. & van Heyningen, V. Long-range control of gene expression: emerging mechanisms and disruption in disease. Am. J. Hum. Genet. 76, 8–32 (2005).

    Article  CAS  Google Scholar 

  25. Lettice, L.A. et al. A long-range Shh enhancer regulates expression in the developing limb and fin and is associated with preaxial polydactyly. Hum. Mol. Genet. 12, 1725–1735 (2003).

    Article  CAS  Google Scholar 

  26. Hong, C.S. & Saint-Jeannet, J.P. Sox proteins and neural crest development. Semin. Cell Dev. Biol. 16, 694–703 (2005).

    Article  CAS  Google Scholar 

  27. McCauley, D.W. & Bronner-Fraser, M. Importance of SoxE in neural crest development and the evolution of the pharynx. Nature 441, 750–752 (2006).

    Article  CAS  Google Scholar 

  28. Pheasant, M. & Mattick, J.S. Raising the estimate of functional human sequences. Genome Res. 17, 1245–1253 (2007).

    Article  CAS  Google Scholar 

  29. Sharpe, J. et al. Optical projection tomography as a tool for 3D microscopy and gene expression studies. Science 296, 541–545 (2002).

    Article  CAS  Google Scholar 

  30. Essafi, A. et al. Direct transcriptional regulation of Bim by FoxO3a mediates STI571-induced apoptosis in Bcr-Abl-expressing cells. Oncogene 24, 2317–2329 (2005).

    Article  CAS  Google Scholar 

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We are grateful to the affected individuals and their families who participated in this study, to the Associations Françaises du Syndrome de Robin, to the Centres de Références Anomalies Cranio-Faciales Rares (AP-HP, Necker and Trousseau hospitals), and to C. Ozilou, G. Staub and G. Guédu-Molina for assistance. We thank T. Attié-Bitach, G. Couly and L. Legeai-Mallet (Necker) and V. van Heyningen and R. Hill (MRC HGU) for useful discussion. This study was underwritten by grants from the Agence Nationale de la Recherche (ERARE grant CraniRare), EUROCRAN FP5, the Fondation pour la Recherche Médicale (FRM), the MRC (UK) and the National Health and Medical Research Council (Australia). S.T. was supported in part by grant NS039818 from the US National Institutes of Health and S.B. by the FRM.

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Authors and Affiliations



S.B., J.A.F. and A.P. performed molecular genetics studies. J.A.F., C.T.G. and N.J. performed chromosomal studies. S.B. and J.R. performed the in vitro enhancer activity experiments. S.B., S.T., C.G., M.V. and H.C.E. performed human expression studies. J.R., S.B. and A.E. performed immunoprecipitation experiments. D.-J.K. performed transgenic assays. J.A.F., S.H., P.P. and D.B. performed the in vivo chromatin compaction studies. M.F. did the OPT image analysis. S.B. and H.R.C. performed the comparative genomic analysis. J.A., V.A., C.A., M.H.-E., N.K., M.M.L., A.P., I.K.T., M.V., P.T., M.-P.V., D.R.F. and S.L. recruited individuals and families affected with PRS. S.L., D.R.F., P.G.F. and J.A. contributed to the concept, strategy, study design and project management. S.B., H.C.E., S.T., P.G.F., D.R.F. and S.L. contributed to the writing of the manuscript. All authors discussed the results.

Corresponding authors

Correspondence to David R FitzPatrick or Stanislas Lyonnet.

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Benko, S., Fantes, J., Amiel, J. et al. Highly conserved non-coding elements on either side of SOX9 associated with Pierre Robin sequence. Nat Genet 41, 359–364 (2009).

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