Interestingly, both families were flagged by the corresponding research group because exome sequencing did not reveal a likely deleterious bi-allelic variant in any of the established JBTS-related genes. Through an investigator-initiated collaboration, an attempt was made to exploit the consanguineous nature of both families, which can readily reveal a potentially unifying etiology if they have an overlapping autozygome, as previously described.7 In brief, we performed genome-wide genotyping with the Axiom SNP Chip platform from Affymetrix and the Sure Select V4 platform from Agilent Technologies and then determined autozygomes by using HomozygosityMapper on all available family members. This revealed a single critical locus (chr10: 101,569,997–109,106,128, UCSC Genome Browser hg 19) (Figure 2A). This locus spans 57 genes, none of which is known to be linked to a ciliopathy phenotype. After re-analyzing the exome variants by only considering variants within this locus (Tables S1 and S2), we found a single previously unreported variant in ARL3 in each index individual: c.445C>T (p.Arg149Cys) (GenBank: NM_004311.3) in family 1 and c.446G>A (p.Arg149His) (GenBank: NM_004311.3) in family 2 (Figure 2B). Both homozygous variants fully co-segregated with the JBTS phenotype in each family.
Figure 2 Molecular Genetic Investigations of the Two JBTS-Affected Families
(A) Genome-wide homozygosity mapping shows the shared homozygous region between the affected members of the two families on chromosome 10 (blue rectangle). Regions of homozygosity are shown in red, and the position of ARL3 is marked with a black arrow.
(B) Schematic representation to ARL3 with the homozygous missense variants located in exon 5.
(C) Evolutionary conservation of residue Arg149, which is highly conserved throughout all species shown except D. melanogaster.
(D) Sequence chromatograms of the two different ARL3 variants described in this study.