Deciphering intrafamilial phenotypic variability by exome sequencing in a Bardet-Biedl family

Abstract

Bardet-Biedl syndrome (BBS) is a model ciliopathy characterized by a wide range of clinical variability. The heterogeneity of this condition is reflected in the number of underlying gene defects and the epistatic interactions between the proteins encoded. BBS is generally inherited in an autosomal recessive trait. However, in some families, mutations across different loci interact to modulate the expressivity of the phenotype. In order to investigate the magnitude of epistasis in one BBS family with remarkable intrafamilial phenotypic variability, we designed an exome sequencing-based approach using SOLID 5500xl platform. This strategy allowed the reliable detection of the primary causal mutations in our family consisting of two novel compound heterozygous mutations in McKusick-Kaufman syndrome (MKKS) gene (p.D90G and p.V396F). Additionally, exome sequencing enabled the detection of one novel heterozygous NPHP4 variant which is predicted to activate a cryptic acceptor splice site and is only present in the most severely affected patient. Here, we provide an exome sequencing analysis of a BBS family and show the potential utility of this tool, in combination with network analysis, to detect disease-causing mutations and second-site modifiers. Our data demonstrate how next-generation sequencing (NGS) can facilitate the dissection of epistatic phenomena, and shed light on the genetic basis of phenotypic variability.

Keywords: Bardet–Biedl Syndrome; MKKS; NGS; NPHP4; intrafamilial variability.

Figure 1

Family cosegregation analysis. RP42 family tree showing the segregation of the sequence variants identified during the molecular analysis of MKKS and NPHP4. [M];[M]: homozygous; [M];[=]: heterozygous.

Figure 2

Detection of novel mutations in the MKKS gene. (A) Chromosome overview of the chromosome 20, MKKS is mapped on region 20p12.2 (black bar). (B) MKKS spans approximately 29 Kb and is composed of six exons. Filled boxes reflect coding exons (3–6) and unfilled boxes reflect UTR. (C) Electropherogram depiction of the index patient (II:4) confirming the heterozygous mutations in exons 3 and 5 of MKKS gene. IUPAC SNP codes used to designate heterozygous substitutions (“k” from Keto for G/T and “r” from puRine for A/G). MKKS Genbank accession number: NG_009109.1.

Figure 3

MKKS protein depiction. (A) Schematic representation of the identified variants within the MKKS domains, including the typical chaperonin group II domains (equatorial, intermediate, and apical). The domain organization was modified from (Stoetzel et al. 2007). (B) Alignment of the orthologs from different species showing conservation of the mutated residues. An * (asterisk) indicates positions which have a single, fully conserved residue. A: (colon) indicates conservation between groups of strongly similar properties. A. (period) indicates conservation between groups of weakly similar properties.

Figure 4

Significant protein–protein interaction network: Network analysis allows relating NPHP4 to BBS proteins, supporting in this way its' possible role as modifier gene.