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Altered Twist1 and Hand2 dimerization is associated with Saethre-Chotzen syndrome and limb abnormalities

Nature Genetics volume 37, pages 373–381 (2005)Cite this article

Abstract

Autosomal dominant mutations in the gene encoding the basic helix-loop-helix transcription factor Twist1 are associated with limb and craniofacial defects in humans with Saethre-Chotzen syndrome. The molecular mechanism underlying these phenotypes is poorly understood. We show that ectopic expression of the related basic helix-loop-helix factor Hand2 phenocopies Twist1 loss of function in the limb and that the two factors have a gene dosage–dependent antagonistic interaction. Dimerization partner choice by Twist1 and Hand2 can be modulated by protein kinase A– and protein phosphatase 2A–regulated phosphorylation of conserved helix I residues. Notably, multiple Twist1 mutations associated with Saethre-Chotzen syndrome alter protein kinase A–mediated phosphorylation of Twist1, suggesting that misregulation of Twist1 dimerization through either stoichiometric or post-translational mechanisms underlies phenotypes of individuals with Saethre-Chotzen syndrome.

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Figure 1: Conservation of helix I threonine and serine residues in the Twist family of bHLH proteins.
Figure 2: Hand2 and Twist1 are phosphorylated by PKA and B56δ-containing PP2A on conserved helix I residues.
Figure 3: Twist1 point mutations in the basic domain have altered phosphorylation by PKA.
Figure 4: Regulated heterodimerization between Twist1 and Hand proteins.
Figure 5: Coexpression of Twist1 and Hand2 in the developing limb.
Figure 6: Chick limb phenotypes caused by Hand2 misexpression.
Figure 7: Genetic antagonism between Twist1 and Hand2.

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References

  1. Castanon, I. & Baylies, M.K. A Twist in fate: evolutionary comparison of Twist structure and function. Gene 287, 11–22 (2002).

    Article  CAS  Google Scholar 

  2. O'Rourke, M.P. & Tam, P.P. Twist functions in mouse development. Int. J. Dev. Biol. 46, 401–413 (2002).

    CAS  PubMed  Google Scholar 

  3. Firulli, A.B. A HANDful of questions: the molecular biology of the heart and neural crest derivatives (HAND)-subclass of basic helix-loop-helix transcription factors. Gene 312, 27–40 (2003).

    Article  CAS  Google Scholar 

  4. Chen, Z.F. & Behringer, R.R. Twist is required in head mesenchyme for cranial neural tube morphogenesis. Genes Dev. 9, 686–699 (1995).

    Article  CAS  Google Scholar 

  5. O'Rourke, M.P., Soo, K., Behringer, R.R., Hui, C.C. & Tam, P.P. Twist plays an essential role in FGF and SHH signal transduction during mouse limb development. Dev. Biol. 248, 143–156 (2002).

    Article  CAS  Google Scholar 

  6. Tavares, A.T., Izpisua-Belmonte, J.C. & Rodriguez-Leon, J. Developmental expression of chick twist and its regulation during limb patterning. Int. J. Dev. Biol. 45, 707–713 (2001).

    CAS  PubMed  Google Scholar 

  7. Wolf, C. et al. The M-twist gene of Mus is expressed in subsets of mesodermal cells and is closely related to the Xenopus X-twi and the Drosophila twist genes. Dev. Biol. 143, 363–373 (1991).

    Article  CAS  Google Scholar 

  8. el Ghouzzi, V. et al. Mutations of the TWIST gene in the Saethre-Chotzen syndrome. Nat. Genet. 15, 42–46 (1997).

    Article  CAS  Google Scholar 

  9. Bourgeois, P. et al. The variable expressivity and incomplete penetrance of the twist-null heterozygous mouse phenotype resemble those of human Saethre-Chotzen syndrome. Hum. Mol. Genet. 7, 945–957 (1998).

    Article  CAS  Google Scholar 

  10. Gripp, K.W., Zackai, E.H. & Stolle, C.A. Mutations in the human TWIST gene. Hum. Mutat. 15, 150–155 (2000).

    Article  CAS  Google Scholar 

  11. Howard, T.D. et al. Mutations in TWIST, a basic helix-loop-helix transcription factor, in Saethre-Chotzen syndrome. Nat. Genet. 15, 36–41 (1997).

    Article  Google Scholar 

  12. Jabs, E.W. TWIST and Saethre-Chotzen Syndrome. in Inborn Errors of Development: The Molecular Basis of Clinical Disorders of Morphogenesis (ed. Epstein, C.J., Erickson, R.P. and Wynshaw-Boris, A.) 401–409 (Oxford University Press, Oxford, 2004).

    Google Scholar 

  13. El Ghouzzi, V. et al. Mutations in the basic domain and the loop-helix II junction of TWIST abolish DNA binding in Saethre-Chotzen syndrome. FEBS Lett. 492, 112–118 (2001).

    Article  CAS  Google Scholar 

  14. El Ghouzzi, V. et al. Saethre-Chotzen mutations cause TWIST protein degradation or impaired nuclear location. Hum. Mol. Genet. 9, 813–819 (2000).

    Article  CAS  Google Scholar 

  15. Charite, J., McFadden, D.G. & Olson, E.N. The bHLH transcription factor dHAND controls Sonic hedgehog expression and establishment of the zone of polarizing activity during limb development. Development 127, 2461–2470 (2000).

    CAS  PubMed  Google Scholar 

  16. Fernandez-Teran, M. et al. Role of dHAND in the anterior-posterior polarization of the limb bud: implications for the Sonic hedgehog pathway. Development 127, 2133–2142 (2000).

    CAS  PubMed  Google Scholar 

  17. Yelon, D. et al. The bHLH transcription factor hand2 plays parallel roles in zebrafish heart and pectoral fin development. Development 127, 2573–2582 (2000).

    CAS  PubMed  Google Scholar 

  18. Massari, M.E. & Murre, C. Helix-loop-helix proteins: regulators of transcription in eucaryotic organisms. Mol. Cell. Biol. 20, 429–440 (2000).

    Article  CAS  Google Scholar 

  19. Firulli, B.A., Hadzic, D.B., McDaid, J.R. & Firulli, A.B. The basic helix-loop-helix transcription factors dHAND and eHAND exhibit dimerization characteristics that suggest complex regulation of function. J. Biol. Chem. 275, 33567–33573 (2000).

    Article  CAS  Google Scholar 

  20. Firulli, B.A. et al. PKA, PKC, and the protein phosphatase 2A influence HAND factor function: a mechanism for tissue-specific transcriptional regulation. Mol. Cell 12, 1225–1237 (2003).

    Article  CAS  Google Scholar 

  21. Castanon, I., Von Stetina, S., Kass, J. & Baylies, M.K. Dimerization partners determine the activity of the Twist bHLH protein during Drosophila mesoderm development. Development 128, 3145–3159 (2001).

    CAS  PubMed  Google Scholar 

  22. Johnson, D. et al. A comprehensive screen for TWIST mutations in patients with craniosynostosis identifies a new microdeletion syndrome of chromosome band 7p21.1. Am. J. Hum. Genet. 63, 1282–1293 (1998).

    Article  CAS  Google Scholar 

  23. Kasparcova, V.S. et al. Molecular analysis of patients with Saethre-Chotzen syndrome: novel mutations and polymorphisms in the TWIST gene. Am. J. Hum. Genet. Suppl. 63, A367 (1998).

    Google Scholar 

  24. Corsi, A.K., Brodigan, T.M., Jorgensen, E.M. & Krause, M. Characterization of a dominant negative C. elegans Twist mutant protein with implications for human Saethre-Chotzen syndrome. Development 129, 2761–2772 (2002).

    CAS  PubMed  Google Scholar 

  25. Centonze, V.E., Sun, M., Masuda, A., Gerritsen, H. & Herman, B. Fluorescence resonance energy transfer imaging microscopy. Methods Enzymol. 360, 542–560 (2003).

    Article  CAS  Google Scholar 

  26. Hamburger, V. & Hamilton, H.L. A series of normal stages in the development of the chick embryo. J. Exp. Morph. 88, 49–92 (1951).

    Article  CAS  Google Scholar 

  27. Yang, Y. et al. Relationship between dose, distance and time in Sonic Hedgehog-mediated regulation of anteroposterior polarity in the chick limb. Development 124, 4393–4404 (1997).

    CAS  Google Scholar 

  28. Tickle, C. The number of polarizing region cells required to specify additional digits in the developing chick wing. Nature 289, 295–298 (1981).

    Article  CAS  Google Scholar 

  29. Olson, E.N. Regulation of muscle transcription by the MyoD family. The heart of the matter. Circ. Res. 72, 1–6 (1993).

    Article  CAS  Google Scholar 

  30. Hamamori, Y., Wu, H.Y., Sartorelli, V. & Kedes, L. The basic domain of myogenic basic helix-loop-helix (bHLH) proteins is the novel target for direct inhibition by another bHLH protein, Twist. Mol. Cell. Biol. 17, 6563–6573 (1997).

    Article  CAS  Google Scholar 

  31. Spicer, D.B., Rhee, J., Cheung, W.L. & Lassar, A.B. Inhibition of myogenic bHLH and MEF2 transcription factors by the bHLH protein Twist. Science 272, 1476–1480 (1996).

    Article  CAS  Google Scholar 

  32. McFadden, D.G., McAnally, J., Richardson, J.A., Charite, J. & Olson, E.N. Misexpression of dHAND induces ectopic digits in the developing limb bud in the absence of direct DNA binding. Development 129, 3077–3088 (2002).

    CAS  PubMed  Google Scholar 

  33. Zuniga, A., Quillet, R., Perrin-Schmitt, F. & Zeller, R. Mouse Twist is required for fibroblast growth factor-mediated epithelial-mesenchymal signalling and cell survival during limb morphogenesis. Mech. Dev. 114, 51–59 (2002).

    Article  CAS  Google Scholar 

  34. Bialek, P. et al. A twist code determines the onset of osteoblast differentiation. Dev. Cell 6, 423–435 (2004).

    Article  CAS  Google Scholar 

  35. Yoshida, C.A. et al. Runx2 and Runx3 are essential for chondrocyte maturation, and Runx2 regulates limb growth through induction of Indian hedgehog. Genes Dev. 18, 952–963 (2004).

    Article  CAS  Google Scholar 

  36. Otto, F. et al. Cbfa1, a candidate gene for cleidocranial dysplasia syndrome, is essential for osteoblast differentiation and bone development. Cell 89, 765–771 (1997).

    Article  CAS  Google Scholar 

  37. Okuda, T., van Deursen, J., Hiebert, S.W., Grosveld, G. & Downing, J.R. AML1, the target of multiple chromosomal translocations in human leukemia, is essential for normal fetal liver hematopoiesis. Cell 84, 321–330 (1996).

    Article  CAS  Google Scholar 

  38. Komori, T. et al. Targeted disruption of Cbfa1 results in a complete lack of bone formation owing to maturational arrest of osteoblasts. Cell 89, 755–764 (1997).

    Article  CAS  Google Scholar 

  39. Logan, M. & Tabin, C. Targeted gene misexpression in chick limb buds using avian replication-competent retroviruses. Methods 14, 407–420 (1998).

    Article  CAS  Google Scholar 

  40. Laufer, E. et al. Expression of Radical fringe in limb-bud ectoderm regulates apical ectodermal ridge formation. Nature 386, 366–373 (1997).

    Article  CAS  Google Scholar 

  41. Srivastava, D. et al. Regulation of cardiac mesodermal and neural crest development by the bHLH transcription factor, dHAND. Nat. Genet. 16, 154–160 (1997).

    Article  CAS  Google Scholar 

  42. Srivastava, D., Cserjesi, P. & Olson, E.N. A subclass of bHLH proteins required for cardiac morphogenesis. Science 270, 1995–1999 (1995).

    Article  CAS  Google Scholar 

  43. Pathi, S. et al. Comparative biological responses to human Sonic, Indian, and Desert hedgehog. Mech. Dev. 106, 107–117 (2001).

    Article  CAS  Google Scholar 

  44. Kaufman, M. The Atlas of Mouse Development (Academic, San Diego, 1992).

    Google Scholar 

Download references

Acknowledgements

We thank K. Dionne and I. Messina for technical assistance; S. Weiner for help with in situ analyses; and L. Field, C. Tabin, T. Jessell, A. Kania, D. Vasiliauskas, L. Zeltser and members of the laboratory of E.L. for comments on the manuscript. This work was supported by the National Institutes of Health (to A.B.F., D.M.V. and D.K.), March of Dimes Birth Defects Foundation (to A.B.F.), American Cancer Society (to P.C.), American Heart Association (to P.C.) and Howard Hughes Medical Institute Research Resources Program for Medical Schools (to E.L.).

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Author notes

  1. Neil Vargesson

    Present address: Section of Cell and Molecular Biology, Imperial College London, Sir Alexander Fleming Building, London, SW7 2AZ, England

  2. Beth A Firulli and Dayana Krawchuk: These authors contributed equally to this work.

Authors and Affiliations

  1. Departments of Pediatrics and Medical & Molecular Genetics, Wells Center for Pediatric Research, James Whitcomb Riley Hospital for Children, Indiana Medical School, 1044 W. Walnut R4 371, Indianapolis, 46202-5225, Indiana, USA

    Beth A Firulli, Simon J Conway & Anthony B Firulli

  2. Department of Genetics and Development, College of Physicians and Surgeons, Columbia University, 701 West 168th Street, New York, 10032, New York, USA

    Dayana Krawchuk, Neil Vargesson & Ed Laufer

  3. Departments of Cellular and Structural Biology, University of Texas Health Science Center at San Antonio, San Antonio, 78229, Texas, USA

    Victoria E Centonze

  4. Huntsman Cancer Institute Center for Children, University of Utah, Salt Lake City, 84112, Utah, USA

    David M Virshup

  5. Department of Cell & Molecular Biology, Tulane University, 2000 Percival Stern Hall, New Orleans, 70118, Louisiana, USA

    Peter Cserjesi

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Correspondence to Ed Laufer or Anthony B Firulli.

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Firulli, B., Krawchuk, D., Centonze, V. et al. Altered Twist1 and Hand2 dimerization is associated with Saethre-Chotzen syndrome and limb abnormalities. Nat Genet 37, 373–381 (2005). https://doi.org/10.1038/ng1525

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