A CNGB1 frameshift mutation in Papillon and Phalène dogs with progressive retinal atrophy

Abstract

Progressive retinal degenerations are the most common causes of complete blindness both in human and in dogs. Canine progressive retinal atrophy (PRA) or degeneration resembles human retinitis pigmentosa (RP) and is characterized by a progressive loss of rod photoreceptor cells followed by a loss of cone function. The primary clinical signs are detected as vision impairment in a dim light. Although several genes have been associated with PRAs, there are still PRAs of unknown genetic cause in many breeds, including Papillons and Phalènes. We have performed a genome wide association and linkage studies in cohort of 6 affected Papillons and Phalènes and 14 healthy control dogs to map a novel PRA locus on canine chromosome 2, with a 1.9 Mb shared homozygous region in the affected dogs. Parallel exome sequencing of a trio identified an indel mutation, including a 1-bp deletion, followed by a 6-bp insertion in the CNGB1 gene. This mutation causes a frameshift and premature stop codon leading to probable nonsense mediated decay (NMD) of the CNGB1 mRNA. The mutation segregated with the disease and was confirmed in a larger cohort of 145 Papillons and Phalènes (PFisher = 1.4×10(-8)) with a carrier frequency of 17.2 %. This breed specific mutation was not present in 334 healthy dogs from 10 other breeds or 121 PRA affected dogs from 44 other breeds. CNGB1 is important for the photoreceptor cell function its defects have been previously associated with retinal degeneration in both human and mouse. Our study indicates that a frameshift mutation in CNGB1 is a cause of PRA in Papillons and Phalènes and establishes the breed as a large functional animal model for further characterization of retinal CNGB1 biology and possible retinal gene therapy trials. This study enables also the development of a genetic test for breeding purposes.

Figure 1. Pedigree of PRA affected Papillons and Phalènes.

Pedigree indicates the affected dogs that were used in the study. Samples from six affected dogs were available for genotyping. Disease segregation is consistent with autosomal recessive mode of inheritance as all affected dogs are born from healthy parents and both sexes are affected. Obligate carrier parents of affected dogs are marked in the pedigree. Obligate carriers genotyped as heterozygous for CNGB1 mutation are marked with a yellow background.

Figure 2. Genome wide association and linkage analyses.

A) A Manhattan plot of genome-wide case-control association analysis performed using 6 cases and 14 controls indicate the most highly associated region in CFA2. B) The PRA associated region on chromosome 2 spans from 61.4 Mb to 63.3 Mb based on association, linkage and joint analyses. C) Genotypes at the PRA associated region on CFA2. All cases share a 1.9 Mb homozygous block, and within this block SNPs BICF2S23238410, BICF2P309315 and BICF2P75954 show complete recessive segregation with the disease. D) Chromatograms of the c.2685delA (arrow), c.2687_2688insTAGCTA (shadowed) mutations in CNGB1 gene in an affected (2) and normal (1) dog. The CNGB1 gene is located between the two segregating SNPs (BICF2P309315, BICF2P75954).

Figure 3. CNGB1 protein alignments.

A) CNGB1 amino acid alignment of the normal and affected dogs. The p.Tyr889Serfs*5 mutation in the affected dog results in a loss of a significant part of the C-terminus of the protein and probable NMD of the CNGB1 mRNA B) CNGB1 sequence alignment between different vertebrates. The mutation is located in a highly conserved region across species. The arrows mark the first mutated amino acid caused by the frameshift and the premature stop codon.