UCSF

Congenital craniofacial anomalies represent one-third of all structural birth defects, but although the genetic causes of many syndromes are known, the molecular and cellular mechanisms often remain poorly understood. This is the case for craniofrontonasal syndrome (CFNS), an X-linked syndrome caused by mutations in EFNB1 and characterized by craniofacial, skeletal, and neurological anomalies. EFNB1 encodes ephrin-B1, a member of the Eph/ephrin family of signaling molecules that regulate cell positioning and tissue morphology during embryonic development. Unlike most X-linked diseases, CFNS manifests very mildly in hemizygous males with no functional ephrin-B1, whereas females heterozygous for ephrin-B1 mutations are severely affected. The increased severity that occurs in the presence of mosaicism of ephrin-B1 expressing and non-expressing cells after random X chromosome inactivation has been named “cellular interference.” In Efnb1+/- mice, ephrin-B1 expressing and non-expressing cells aberrantly segregate from one another in the neural plate neuroepithelium, providing a general potential mechanism for cellular interference; however, whether and how ephrin-B1-mediated cell segregation contributes to craniofacial phenotypes in human patients was not known. Using mouse models and the first human induced pluripotent stem cell model of a human craniofacial disease, we established that ephrin-B1 mosaicism drives pathogenic cell segregation in human cell types relevant to CFNS and that ephrin-B1 is a potent regulator of segregation not only in the early neuroepithelium, but also in the craniofacial mesenchyme, correlating with dysmorphology of facial structures. Ephrin-B1 also regulates EphB receptor levels when these molecules are coexpressed, and aberrant EphB receptor accumulation in ephrin-B1 mosaic cell populations may influence segregation or phenotypic outcome and disease severity. These discoveries contribute to a greater understanding of the role of ephrin-B1 in regulating cell positioning during development, as well as how disruption of its function can result in congenital craniofacial disease.