Abstract
Otofaciocervical syndrome (OFCS) is an autosomal recessively inherited disorder characterized by facial dysmorphism, external ear anomalies with preauricular pits and hearing impairment, branchial cysts or fistulas, anomalies of the vertebrae and the shoulder girdle, and mild intellectual disability. In a large consanguineous family with OFCS from Turkey, we performed whole-exome sequencing (WES) of a single pooled DNA sample of four affected individuals. Filtering for variants with a percentage of alternate reads ≥90 % and a coverage of at least five reads identified only a single novel homozygous variant, c.497G>T, located in PAX1 that co-segregated with the disease in the family. PAX1 encodes a transcription factor with a critical role in pattern formation during embryogenesis in vertebrates. The mutation is predicted to substitute the glycine at position 166 to valine (p.G166V) within the highly conserved paired-box domain of the PAX1 protein. We performed a dual luciferase reporter assay to examine the transactivation of a regulatory sequence in the Nkx3-2 promoter region, which is a direct target of mouse Pax1 transcriptional regulation. We observed a significantly reduced transactivation in HEK293T cells overexpressing Pax1G157V in comparison to Pax1WT expressing cells, indicating a reduced DNA-binding affinity of the mutant protein. Taken together, our results show that the strategy of pooling DNA is a powerful, cost-effective application for WES in consanguineous families and establish PAX1 as a new disease-causing gene for OFCS and as part of the EYA-DACH-SIX-PAX network, important in early embryogenesis.
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Acknowledgments
We are grateful to all family members who participated in this study, to Esther Milz for excellent technical assistance, to Andrea Pannes for her technical advice in slow-down PCR, to Simon von Ameln for cochlea preparation, and to Karin Boss for critically reading the manuscript. This work was supported by the German Federal Ministry of Education and Research (BMBF) by grant number 01GM1211A (E-RARE network CRANIRARE-2) to B.W.
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The authors declare that they have no conflict of interest.
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Pohl, E., Aykut, A., Beleggia, F. et al. A hypofunctional PAX1 mutation causes autosomal recessively inherited otofaciocervical syndrome. Hum Genet 132, 1311–1320 (2013). https://doi.org/10.1007/s00439-013-1337-9
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DOI: https://doi.org/10.1007/s00439-013-1337-9