This is a proposal to investigate the morphogenesis of the coronal suture and mechanisms of coronal synostosis. The long-term aim is to understand how boundaries between cellular compartments form, and how such boundaries function in growth control and patterning. The cellular mechanisms underlying craniosynostosis are largely unknown. Prevalent views, based largely on ex vivo approaches, seek to explain craniosynostosis in terms of the timing of differentiation or survival of osteoblast populations. Our results suggest that understanding the fundamental causes of craniosynostosis requires an analysis at a higher level of biological organization. Through an examination of the Twist mutant mouse, we found that synostosis of the coronal suture is associated with a defect in the boundary between neural crest-derived mesenchyme that forms the frontal bone and mesodermal-derived mesenchyme that forms the parietal bone. In an effort to identify other genes that function together with Twist in coronal suture development, we have uncovered evidence that perturbations in Eph-ephrin and Bmp signaling may contribute to these defects. That ephrin signaling may have a part in the Twist mutant phenotype is suggested by our observations (i) that ephrins A2 and A4, as well as their receptor, EphA4, are expressed in a highly localized manner in the developing coronal suture, (ii) and that their expression is altered in Twist mutant mice and restored to their wild type pattern in Msx2-Twist double mutants. That BMP signaling is involved is suggested by our observation that the gene encoding the Bmp antagonist, noggin, is upregulated in the prospective coronal suture of Twist mutant mice, and that transgenic overexpression of noggin causes a sutural defect similar to that seen in Twist mutants. Moreover, inactivation of Bmp4 in neural crest also causes fusion of the frontal and parietal bones at the coronal suture. These data lead us to the hypothesis that craniosynostosis in the Twist mutant is caused in part by a perturbations in gene networks that control boundary formation, and that these networks are likely to include elements of the Eph-ephrin and BMP pathways. Here we propose (i) to carry out a series of genetic experiments aimed at testing the hypothesis that Twist interacts functionally with the Eph-ephrin and BMP pathways in the patterning of the coronal suture; (ii) to test the hypothesis that Msx2 and Twist function cooperatively in a genetic cascade that regulates boundary formation and coronal suture development. This will entail an analysis both of coronal suture development and the expression of ephrinA2 and EphA4 in Msx2-Twist double mutants. Finally, we will examine the regulatory relationship between Msx2 and Twist. We will investigate the mechanism by which reduced Twist function leads to expanded expression of Msx2, and we will determine whether upregulation of Msx2 is sufficient to cause a boundary defect and synostosis in the coronal suture. The significance of the proposed studies is, first, that they will contribute information on the pathophysiology of an important class of craniofacial disorders-the craniosynostoses. Second, they will address the biological significance of boundary formation in patterning and growth control-a fundamental problem in developmental biology. ? ?

National Institute of Health (NIH)
National Institute of Dental & Craniofacial Research (NIDCR)
Research Project (R01)
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Skeletal Biology Development and Disease Study Section (SBDD)
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Small, Rochelle K
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University of Southern California
Schools of Medicine
Los Angeles
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Sun, Jingjing; Ting, Man-Chun; Ishii, Mamoru et al. (2016) Msx1 and Msx2 function together in the regulation of primordial germ cell migration in the mouse. Dev Biol 417:11-24
Ishii, Mamoru; Sun, Jingjing; Ting, Man-Chun et al. (2015) The Development of the Calvarial Bones and Sutures and the Pathophysiology of Craniosynostosis. Curr Top Dev Biol 115:131-56
Cheng, Su-Li; Behrmann, Abraham; Shao, Jian-Su et al. (2014) Targeted reduction of vascular Msx1 and Msx2 mitigates arteriosclerotic calcification and aortic stiffness in LDLR-deficient mice fed diabetogenic diets. Diabetes 63:4326-37
Sharma, Vikram P; Fenwick, Aimée L; Brockop, Mia S et al. (2013) Mutations in TCF12, encoding a basic helix-loop-helix partner of TWIST1, are a frequent cause of coronal craniosynostosis. Nat Genet 45:304-7
Sun, Jingjing; Ishii, Mamoru; Ting, Man-Chun et al. (2013) Foxc1 controls the growth of the murine frontal bone rudiment by direct regulation of a Bmp response threshold of Msx2. Development 140:1034-44
Cha, Jeeyeon; Sun, Xiaofei; Bartos, Amanda et al. (2013) A new role for muscle segment homeobox genes in mammalian embryonic diapause. Open Biol 3:130035
Ishii, Mamoru; Arias, Athena C; Liu, Liqiong et al. (2012) A stable cranial neural crest cell line from mouse. Stem Cells Dev 21:3069-80
Ting, Man-Chun; Liao, Chun-Peng; Yan, Chunli et al. (2012) An enhancer from the 8q24 prostate cancer risk region is sufficient to direct reporter gene expression to a subset of prostate stem-like epithelial cells in transgenic mice. Dis Model Mech 5:366-74
Daikoku, Takiko; Cha, Jeeyeon; Sun, Xiaofei et al. (2011) Conditional deletion of Msx homeobox genes in the uterus inhibits blastocyst implantation by altering uterine receptivity. Dev Cell 21:1014-25
Roybal, Paul G; Wu, Nancy L; Sun, Jingjing et al. (2010) Inactivation of Msx1 and Msx2 in neural crest reveals an unexpected role in suppressing heterotopic bone formation in the head. Dev Biol 343:28-39

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