Identification of the causal variants for different inherited retinal phenotypes in a complex consanguineous family from Pakistan

Abstract

Purpose: A large consanguineous Pakistani family with multiple loops in which the affected individuals suffered from Leber congenital amaurosis (LCA), autosomal recessive retinitis pigmentosa (arRP), or cone dysfunction (CD) was analyzed for the identification of causal genetic defects in this family by using homozygosity mapping and whole exome sequencing (WES).

Methods: We performed homozygosity mapping in two LCA siblings using the Illumina 700k single nucleotide polymorphism microarray. Six retinal disease genes (IQCB1, KLHL7, MERTK, NPHP3, RBP3, and RHO) residing in the shared homozygous regions larger than 2 Mb were sequenced. One LCA patient was subsequently analyzed using WES and variants with a PhyloP >2.7 and dbSNP frequency <0.5% were selected. Segregation analysis was carried out using restriction fragment length polymorphism analysis or Sanger sequencing in family members of all loops.

Results: Sanger sequencing of six candidate genes residing in shared homozygous regions of two LCA siblings revealed no causal variants. After filtering of WES data, a previously published mutation in RPGRIP1 (p.R827L) was detected in the LCA proband. This mutation was found to segregate with the disease phenotypes in homozygous state in the LCA and CD loops, but not in the arRP loop. As the LCA siblings are closely related to the arRP patients in the neighboring loop, WES data of the LCA patient was further analyzed to identify additional pathogenic variants, that could potentially cause arRP. A heterozygous nonsense mutation in SAG (p.R292*) was detected in the LCA proband, and this mutation was found homozygously in all three arRP cases.

Conclusions: We found that the variability in inherited retinal dystrophies in a Pakistani family is due to variations in two genes that encode components of different structures in photoreceptor cells, i.e. the connecting cilium involved in transport processes (RPGRIP1), and the photoreceptor outer segments harboring the phototransduction cascade (SAG). Four patients carry two RPGRIP1 variants and one SAG variant and one patient carries two SAG variants and one RPGRIP1 variant. To investigate whether the additional alleles modify the respective phenotypes, these patients will be clinically re-evaluated to establish detailed genotype-phenotype correlations.