Screening of the arrestin gene in dogs afflicted with generalized progressive retinal atrophy

BACKGROUND: Intronic DNA sequences of the canine arrestin (SAG) gene was screened to identify potential disease causing mutations in dogs with generalized progressive retinal atrophy (gPRA). The intronic sequences flanking each of the 16 exons were obtained from clones of a canine genomic library. RESULTS: Using polymerase chain reaction-single strand conformation polymorphism (PCR-SSCP) and DNA sequence analyses we screened affected and unaffected dogs of 23 breeds with presumed autosomal recessively (ar) transmitted gPRA. In the coding region of the SAG gene 12 nucleotide exchanges were identified, 5 of which lead to amino acid substitutions (H14C; A111V; A113T; D259T; A379E). 7 other exonic substitutions represent silent polymorphisms (C132C; Q199Q; H225H; V247V; P264P; T288T and L293L). 16 additional sequence variations were observed in intronic regions of different dog breeds. CONCLUSIONS: In several breeds, these polymorphisms were found in homozygous state in unaffected and in heterozygous state in affected animals. Consequently these informative substitutions provide evidence to exclude mutations in the SAG gene as causing retinal degeneration in 14 of the 23 dog breeds with presumed ar transmitted gPRA

Indirect exclusion of four candidate genes for generalized progressive retinal atrophy in several breeds of dogs

BACKGROUND: Generalized progressive retinal atrophy (gPRA) is a hereditary ocular disorder with progressive photoreceptor degeneration in dogs. Four retina-specific genes, ATP binding cassette transporter retina (ABCA4), connexin 36 (CX36), c-mer tyrosin kinase receptor (MERTK) and photoreceptor cell retinol dehydrogenase (RDH12) were investigated in order to identify mutations leading to autosomal recessive (ar) gPRA in 29 breeds of dogs. RESULTS: Mutation screening was performed initially by PCR and single strand conformation polymorphism (SSCP) analysis, representing a simple method with comparatively high reliability for identification of sequence variations in many samples. Conspicuous banding patterns were analyzed via sequence analyses in order to detect the underlying nucleotide variations. No pathogenetically relevant mutations were detected in the genes ABCA4, CX36, MERTK and RDH12 in 71 affected dogs of 29 breeds. Yet 30 new sequence variations were identified, both, in the coding regions and intronic sequences. Many of the sequence variations were in heterozygous state in affected dogs. CONCLUSION: Based on the ar transmittance of gPRA in the breeds investigated, informative sequence variations provide evidence allowing indirect exclusion of pathogenetic mutations in the genes ABCA4 (for 9 breeds), CX36 (for 12 breeds), MERTK (for all 29 breeds) and RDH12 (for 9 breeds)

Exclusion of PINK1 as candidate gene for the late-onset form of Parkinson's disease in two European populations

BACKGROUND: Parkinson's disease (PD) is the second most common neurodegenerative disorder. Recently, mutations in the PINK1 (PARK6) gene were shown to rarely cause autosomal-recessively transmitted, early-onset parkinsonism. In order to evaluate whether PINK1 contributes to the risk of common late-onset PD we analysed PINK1 sequence variations. A German (85 patients) and a Norwegian cohort (90 patients) suffering from late-onset PD were screened for mutations and single nucleotide polymorphisms (SNPs) in the PINK1 gene. Both cohorts consist of well-characterized patients presenting a positive family history of PD in ~17%. Investigations were performed by single strand conformation polymorphism (SSCP), denaturating high performance liquid chromatography (DHPLC) and sequencing analyses. SNP frequencies were compared by the χ(2 )test RESULTS: Several common SNPs were identified in our cohorts, including a recently identified coding variant (Q115L) in exon 1. Genotyping of the Q115L variation did not reveal significant frequency differences between patients and controls. Pathogenic mutations in the PINK1 gene were not identified, neither in the German nor in the Norwegian cohort. CONCLUSION: Sequence variation in the PINK1 gene appears to play a marginal quantitative role in the pathogenesis of the late-onset form of PD, in German and Norwegian cohorts, if at all

Kandidatengenanalyse für generalisierte progressive Retina-Atrophie in dreißig Hunderassen

Die generalisierte progressive Retina-Atrophie (gPRA) ist eine meist autosomal rezessiv vererbte, genetisch heterogene Augenerkrankung bei Hunden, die im Endstadium immer zur Blindheit führt. Während der Dissertation wurden anhand mehrerer Strategien Mutationen in Kandidatengenen (deren Produkte am Sehvorgang beteiligt sind) definiert, die jeweils für gPRA in den zur Verfügung stehenden 30 verschiedenen Rassen verantwortlich sind. Teilweise wurden Gene für die vollständige Kandidatengenanalyse aus einer Lambda Phagengenombank isoliert, sequenziert und charakterisiert. Insgesamt wurden 13 Kandidatengene untersucht. Dabei wurden eine Vielzahl von Sequenzvariationen in diesen Genen nachgewiesen mit deren Hilfe diese Gene als Ursache der gPRA in verschiedenen Rassen anhand von Stammbaumanalysen ausgeschlossen werden konnten. Eine gPRA-verursachende Mutation (8-bp Insertion) im Exon 21 des PDE6 Gens wurde in Sloughis identifiziert und ein DNA-Test für diese Rasse etabliert

Analysis of PDE6D

Abstract The δ and γ subunits of the cGMP-phosphodiesterase (PDE6D, PDE6G) genes were screened in order to identify mutations causing generalised progressive retinal atrophy (gPRA) in dogs. In the PDE6D gene, single nucleotide polymorphisms (SNP) were observed in exon 4, in introns 2 and 3 and in the 3' untranslated region (UTR) of different dog breeds. In the coding region of the PDE6G gene, exclusively healthy Labrador Retrievers showed an A → G transition in exon 4 without amino acid exchange. SNP were also observed in introns 1 and 2 in different dog breeds. The different SNP were used as intragenic markers to investigate the involvement of both genes in gPRA. The informative substitutions allowed us to exclude mutations in the PDE6D and PDE6G genes as causing retinal degeneration in 15 of the 22 dog breeds with presumed autosomal recessively transmitted (ar) gPRA.</p

Analysis of PDE6D and PDE6G genes for generalised progressive retinal atrophy (gPRA) mutations in dogs

The $\delta$ and $\gamma$ subunits of the cGMP-phosphodiesterase (PDE6D, PDE6G) genes were screened in order to identify mutations causing generalised progressive retinal atrophy (gPRA) in dogs. In the PDE6D gene, single nucleotide polymorphisms (SNP) were observed in exon 4, in introns 2 and 3 and in the $3'$ untranslated region (UTR) of different dog breeds. In the coding region of the PDE6G gene, exclusively healthy Labrador Retrievers showed an A $\rightarrow$ G transition in exon 4 without amino acid exchange. SNP were also observed in introns 1 and 2 in different dog breeds. The different SNP were used as intragenic markers to investigate the involvement of both genes in gPRA. The informative substitutions allowed us to exclude mutations in the PDE6D and PDE6G genes as causing retinal degeneration in 15 of the 22 dog breeds with presumed autosomal recessively transmitted (ar) gPRA