Evidence for additional FREM1 heterogeneity in Manitoba oculotrichoanal syndrome

PURPOSE: Manitoba Oculotrichoanal (MOTA) syndrome is an autosomal recessive disorder present in First Nations families that is characterized by ocular (cryptophthalmos), facial, and genital anomalies. At the commencement of this study, its genetic basis was undefined. METHODS: Homozygosity analysis was employed to map the causative locus using DNA samples from four probands of Cree ancestry. After single nucleotide polymorphism (SNP) genotyping, data were analyzed and exported to PLINK to identify regions identical by descent (IBD) and common to the probands. Candidate genes within and adjacent to the IBD interval were sequenced to identify pathogenic variants, with analyses of potential deletions or duplications undertaken using the B-allele frequency and log(2) ratio of SNP signal intensity. RESULTS: Although no shared IBD region >1 Mb was evident on preliminary analysis, adjusting the criteria to permit the detection of smaller homozygous IBD regions revealed one 330 Kb segment on chromosome 9p22.3 present in all 4 probands. This interval comprising 152 SNPs, lies 16 Kb downstream of FRAS1-related extracellular matrix protein 1 (FREM1), and no copy number variations were detected either in the IBD region or FREM1. Subsequent sequencing of both genes in the IBD region, followed by FREM1, did not reveal any mutations. CONCLUSIONS: This study illustrates the utility of studying geographically isolated populations to identify genomic regions responsible for disease through analysis of small numbers of affected individuals. The location of the IBD region 16 kb from FREM1 suggests the phenotype in these patients is attributable to a variant outside of FREM1, potentially in a regulatory element, whose identification may prove tractable to next generation sequencing. In the context of recent identification of FREM1 coding mutations in a proportion of MOTA cases, characterization of such additional variants offers scope both to enhance understanding of FREM1’s role in cranio-facial biology and may facilitate genetic counselling in populations with high prevalences of MOTA to reduce the incidence of this disorder.