Identification and in silico analyses of novel TGFBR1 and TGFBR2 mutations in Marfan syndrome-related disorders

Very recently, heterozygous mutations in the genes encoding transforming growth factor beta receptors I (TGFBR1) and II (TGFBR2) have been reported in Loeys-Dietz aortic aneurysm syndrome (LDS). In addition, dominant TGFBR2 mutations have been identified in Marfan syndrome type 2 (MFS2) and familial thoracic aortic aneurysms and dissections (TAAD). In the past, mutations of these genes were associated with atherosclerosis and several human cancers. Here, we report a total of nine novel and one known heterozygous sequence variants in the TGFBR1 and TGFBR2 genes in nine of 70 unrelated individuals with MFS-like phenotypes who previously tested negative for mutations in the gene encoding the extracellular matrix protein fibrillin-1 (FBN1). To assess the pathogenic impact of these sequence variants, in silico analyses were performed by the PolyPhen, SIFT, and Fold-X algorithms and by means of a 3D homology model of the TGFBR2 kinase domain. Our results showed that in all but one of the patients the pathogenic effect of at least one sequence variant is highly probable (c.722C > T, c.799A > C, and c.1460G > A in TGFBR1 and c.773T > G, c.1106G > T, c.1159G > A, c.1181G > A, and c.1561T > C in TGFBR2). These deleterious alleles occurred de novo or segregated with the disease in the families, indicating a causative association between the sequence variants and clinical phenotypes. Since TGFBR2 mutations found in patients with MFS-related disorders cannot be distinguished from heterozygous TGFBR2 mutations reported in tumor samples, we emphasize the importance of segregation analysis in affected families. In order to be able to find the mutation that is indeed responsible for a MFS-related phenotype, we also propose that genetic testing for sequence alterations in TGFBR1 and TGFBR2 should be complemented by mutation screening of the FBN1 gene.

Whole-Genome Sequencing of a Canine Family Trio Reveals a FAM83G Variant Associated with Hereditary Footpad Hyperkeratosis

Over 250 Mendelian traits and disorders, caused by rare alleles have been mapped in the canine genome. Although each disease is rare in the dog as a species, they are collectively common and have major impact on canine health. With SNP-based genotyping arrays, genome-wide association studies (GWAS) have proven to be a powerful method to map the genomic region of interest when 10-20 cases and 10-20 controls are available. However, to identify the genetic variant in associated regions, fine-mapping and targeted re-sequencing is required. Here we present a new approach using whole-genome sequencing (WGS) of a family trio without prior GWAS. As a proof-of-concept, we chose an autosomal recessive disease known as hereditary footpad hyperkeratosis (HFH) in Kromfohrl änder dogs. To our knowledge, this is the first time this family trio WGS-approach, has successfully been used to identify a genetic variant that perfectly segregates with a canine disorder. The sequencing of three Kromfohrl änder dogs from a family trio (an affected offspring and both its healthy parents) resulted in an average genome coverage of 9.2X per individual. After applying stringent filtering criteria for candidate causative coding variants, 527 single nucleotide variants (SNVs) and 15 indels were found to be homozygous in the affected offspring and heterozygous in the parents. Using the computer software packages ANNOVAR and SIFT to functionally annotate coding sequence differences and to predict their functional effect, resulted in seven candidate variants located in six different genes. Of these, only FAM83G:c155G>C (p.R52P) was found to be concordant in eight additional cases and 16 healthy Kromfohrl änder dogs.