title: A hypofunctional PAX1 mutation causes autosomal recessively inherited otofaciocervical syndrome.
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 Pax1(G157V) in comparison to Pax1(WT) 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.
authors.affiliation: Institute of Human Genetics, University Medical Faculty, University of Cologne, Kerpener Str. 34, 50931, Cologne, Germany.
authors.forename: Esther
authors.initials: E
authors.lastname: Pohl
authors.forename: Ayca
authors.initials: A
authors.lastname: Aykut
authors.forename: Filippo
authors.initials: F
authors.lastname: Beleggia
authors.forename: Emin
authors.initials: E
authors.lastname: Karaca
authors.forename: Burak
authors.initials: B
authors.lastname: Durmaz
authors.forename: Katharina
authors.initials: K
authors.lastname: Keupp
authors.forename: Esra
authors.initials: E
authors.lastname: Arslan
authors.forename: Melis
authors.initials: M
authors.lastname: Palamar
authors.forename: Melis Palamar
authors.initials: MP
authors.lastname: Onay
authors.forename: Gökhan
authors.initials: G
authors.lastname: Yigit
authors.forename: Ferda
authors.initials: F
authors.lastname: Özkinay
authors.forename: Bernd
authors.initials: B
authors.lastname: Wollnik
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D019280:Branchio-Oto-Renal Syndrome / Q000235:genetics*; D005260:Female; D018507:Gene Expression Regulation, Developmental; D057809:HEK293 Cells; D018398:Homeodomain Proteins / Q000235:genetics / Q000378:metabolism; D006801:Humans; D008297:Male; D051379:Mice; D008969:Molecular Sequence Data; D009154:Mutation; D051761:Paired Box Transcription Factors / Q000235:genetics* / Q000378:metabolism; D010375:Pedigree; D011401:Promoter Regions, Genetic; D017422:Sequence Analysis, DNA; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism; D014421:Turkey
chemical_list: D018398:Homeodomain Proteins; C491536:NKX3-2 protein, human; C107965:Nkx3-2 protein, mouse; D051761:Paired Box Transcription Factors; D014157:Transcription Factors; C070835:PAX1 transcription factor