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Frauke Coppieters, Kristof Van Schil, Steve Lefever, Julie De Zaeytijd, Bart P. Leroy, Ingele Casteels, Thomy de Ravel, Elfride De Baere; Combined Identity-by-descent Mapping And Exome Sequencing Identifies A Novel MERTK Mutation In Autosomal Recessive Retinitis Pigmentosa. Invest. Ophthalmol. Vis. Sci. 2012;53(14):4521.
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© ARVO (1962-2015); The Authors (2016-present)
Autosomal recessive retinitis pigmentosa (ARRP) is a genetically heterogeneous disorder, with 35 disease genes and 3 loci associated thus far. The goal of this study was to identify the molecular defect in a consanguineous ARRP family with one affected individual and three unaffected siblings (parents: first cousins once removed).
Genomewide identity-by-descent (IBD) mapping was performed in the patient and three unaffected siblings using GeneChip Human Mapping 250K Nsp arrays (Affymetrix). Subsequently, the patient underwent exome enrichment (TruSeq Exome Enrichment Kit, Illumina) and sequencing (2x100 cycles, HiSeq, Illumina). The CLC Genomics Workbench (CLC bio) was employed for read mapping and variant calling. Variants were confirmed using Sanger sequencing (ABI 3730xl, Applied Biosystems).
IBD mapping revealed 5 homozygous regions larger than 10 Mb in the patient. However, 4 of these were also homozygous in one or more healthy siblings, thus leaving a 37.6 Mb region on chromosome 2 as the best candidate.In total, 80/89 million reads could be mapped against the human genome reference sequence (NCBI, GRCh37.p5), resulting in an average coverage of 45x. The candidate region on chromosome 2 contained 836 substitutions and 24 deletions/insertions with a coverage and a variant allele frequency equal to or above 10x and 75%, respectively. After candidate gene analysis of all RetNet genes, the following novel homozygous missense mutation was identified in MERTK: c.2180G>A (p.Arg727Gln) (NM_006343.2). This mutation affects a highly conserved nucleotide and amino acid, is located in the active site of the tyrosine protein kinase domain, and is predicted to abolish protein function by both SIFT and PolyPhen. Moreover, the carbonyl oxygen of the Arg727 residue is known to be involved in a hydrogen-bond with the hydroxyl group of the ligand Compound-52 (Huang et al., 2009, J Struct Biol). In addition, the mutation was heterozygous in the parents and heterozygous or absent in the healthy siblings. Notably, the age-of-onset (12 years), photophobia, and macular pigment alterations observed in the patient are in agreement with previously reported MERTK-associated phenotypes.
In this study, we identified the causal genetic defect in a consanguineous family with a single ARRP patient by combining IBD mapping with exome sequencing. This approach is a powerful tool to establish a molecular diagnosis in genetic heterogeneous conditions such as most retinal dystrophies.
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