Frameshift Variant in AMPD2 in Cirneco dell'Etna Dogs with Retinopathy and Tremors

Abstract

While the manifestations of many inherited retinal disorders are limited to loss of vision, others are part of a syndrome that affects multiple tissues, particularly the nervous system. Most syndromic retinal disorders are thought to be recessively inherited. Two dogs out of a litter of Cirneco dell' Etna dogs, both males, showed signs of retinal degeneration, along with tremors and signs described as either atypical seizures or paroxysmal dyskinesias, while the other two male littermates were normal. We named this oculo-neurological syndrome CONS (Cirneco oculo-neurological syndrome), and undertook homozygosity mapping and whole-genome sequencing to determine its potential genetic etiology. Notably, we detected a 1-bp deletion in chromosome 6 that was predicted to cause a frameshift and premature stop codon within the canine AMPD2 gene, which encodes adenosine monophosphate deaminase, an enzyme that converts adenosine 5'-monophosphate (AMP) to inosine 5'-monophosphate (IMP). Genotyping of the available Cirneco population suggested perfect segregation between cases and controls for the variant. Moreover, this variant was absent in canine genomic databases comprised of thousands of unaffected dogs. The AMPD2 genetic variant we identified in dogs presents with retinal manifestations, adding to the spectrum of neurological manifestations associated with AMPD2 variants in humans.

Keywords: animal model; inherited canine disease; oculo-neurological syndrome; syndromic retinal condition.

Figure 1

Retinal and cerebral phenotype. Left (A) and right (B) fundus pictures of a case (CRN4). Hyperreflectivity and marked attenuation/loss of retinal vessels along with pale atrophic optic discs indicate advanced stages of retinal degeneration. Sclerosis of the choroidal vessels is also present in the inferior non-tapetal region, accompanied by segmental choroid retinal atrophy. MRI of CRN4 (C) and CRN6 (D). (C) Transverse T2 FLAIR image at the level of the thalamus, where bilateral and symmetrical hyperintensities affecting the periventricular white matter tracts are depicted (arrows). (D): Transverse T2 weighted image at the level of the thalamus. In this case, no abnormalities were observed in the brain parenchyma. Moderate-to-severe left temporalis muscle atrophy is evident (arrow).

Figure 2

Family tree and mapping. (A) Family tree of the cases. The four siblings shown include the two cases. Homozygous cases CRN4 and CRN6 are shown with a red-filled shape. Carriers (CRN1 and CRN5) and wild-type (CRN7) controls are shown with a half-filled and empty shape, respectively. Males are indicated with a square, females with a circle. Missing samples that we were unable to genotype are indicated with a diagonal bar. The carrier grandmother CRN1 is shared by both parents. (B) Mapping of the candidate regions. Homozygous regions exclusive for the cases are shown in red. The interval containing the candidate variant is indicated with an asterisk.

Figure 3

Sequencing of the putative causative variant. (A) Whole genome sequencing of two affected Cirneco (CRN4 and CRN6) and of unrelated retinal RNA-seq (confirming the exon and transcript in retina) shown in Integrate Genome Viewer. Note the 1-bp deletion (g.42,698,170delC). (B) Sanger sequencing of a control (top) a carrier (middle) and a case (bottom) showing the deletion of the coding C. Variant indicated with a red arrow.

Figure 4

Position of the proposed causative variant. (A) Position of the c.2,131delG variant in the 15th exon of the canine AMPD2 transcript. Exons shown as re-annotated from canine retinal RNA-seq. (B) Human AMPD2 protein with the position of the AMPD2 mutations associated with neurological diseases in Homo sapiens as reported in the literature [52,53,54,55,56,57]. The canine AMPD2 variant (position as realigned to the human protein) is reported in red and with the larger arrow. As for most pathological human variants, the mutation we report occurs within the AMPD domain.

Figure 5

(A) Representation of the first two principal components (PCs) of the multidimensional scaling analysis of the individual identity-by-state distances. (B) Best-fitting model (number of clusters K = 23) obtained from the admixture analysis. Each bar represents a subject and each color a different cluster.