Frontonasal Displazi Tip 3 Ailelerinde Malformasyondan Sorumlu Genin Araştırılması

Abstract

Frontonasal dysplasia type 3 (FND3, MIM 613456) is an autosomal recessive severe facial malformation characterized by biletaral extreme microphthalmia, bilateral oblique facial cleft, complete cleft palate, hypertelorism, wide nasal bridge with hypoplasia of the ala nasi, large cranial bone defect s associated with caudal appendage. Both a microdeletion on chromosome 12q21 region containing ALX1 and a point mutation in the ALX1 produce similar phenotype. In this study, a large autosomal recessive Turkish FND3 family and two additional singleton families from Sri Lanka and Egypt were studied. DNA sequencing analysis of ALX1 coding region failed to identify additional mutations in these three families. Putative deletions or duplications within ALX1 gene were excluded using both quantitative real time PCR (qPCR) and multiple ligation probe amplification (MLPA) analyses. 250K SNP chip genotyping followed by homozygosity mapping revealed a single 7Mb homozygote region on chromosome 12q21 harboring ALX1 in two out of three families. The 12q21 region was excluded in the third family providing evidence for genetic heterogeneity of FND3 malformation. Identification of homozygosity to ALX1 region suggested that ALX1 mutations might be located at non-coding segments such as promoter or other regulatory elements at least in some families. In order to explore this possibility approximately 22.000 base pairs from both proximal and distal parts of the ALX1 as well as intronic regions were screened by Sanger sequencing. No causative variant was identified. Whole exome (WES) and Whole genome (WGS) sequencing followed by variant filtration analysis identified a total of 464 variants in the critical region. Region–based regulatory annotation using intuitive interface of UCSC genome browser revealed a total of 27 critical variants residing on putative transcription factor binding sites. This study clearly showed that as yet unidentified mutations lying in the non-coding/regulatory regions of ALX1 gene contribute autosomal recessive FND3 malformation in some families.