RESUMO
Over 200 mutations in the retina specific member of the ATP-binding cassette transporter superfamily (ABCA4) have been associated with a diverse group of human retinal diseases. The disease mechanisms, and genotype-phenotype associations, nonetheless, remain elusive in many cases. As orthologous genes are commonly mutated in canine models of human blinding disorders, canine ABCA4 appears to be an ideal candidate gene to identify and study sequence changes in dogs affected by various forms of inherited retinal degeneration. However, the size of the gene and lack of haplotype assignment significantly limit targeted association and/or linkage approaches. This study assessed the naturally observed sequence diversity of ABCA4 in the dog, identifying 80% of novel variations. While none of the observed polymorphisms have been associated with blinding disorders to date, breed and potentially disease specific haplotypes have been identified. Moreover, a tag SNP map of 17 (15) markers has been established that accurately predicts common ABCA4 haplotypes (frequency > 5%) explaining >85% (>80%) of the observed genetic diversity and will considerably advance future studies. Our sequence analysis of the complete canine ABCA4 coding region will clearly provide a baseline and tools for future association studies and comparative genomics to further delineate the role of ABCA4 in canine blinding disorders.
Assuntos
Transportadores de Cassetes de Ligação de ATP/genética , Polimorfismo de Nucleotídeo Único , Doenças Retinianas/genética , Substituição de Aminoácidos , Animais , Doenças do Cão/genética , Cães/genética , Feminino , Variação Genética , Humanos , Masculino , Modelos Animais , Mutação , Degeneração Retiniana/genética , Degeneração Retiniana/veterinária , Especificidade da EspécieRESUMO
PURPOSE: To identify the genomic location of previously uncharacterized canine retina-expressed expressed sequence tags (ESTs), and thus identify potential candidate genes for heritable retinal disorders. METHODS: A set of over 500 retinal canine ESTs were mapped onto the canine genome using the RHDF(5000-2) radiation hybrid (RH) panel, and the resulting map positions were compared to their respective localization in the CanFam2 assembly of the canine genome sequence. RESULTS: Unique map positions could be assigned for 99% of the mapped clones, of which only 29% showed significant homology to known RefSeq sequences. A comparison between RH map and sequence assembly indicated some areas of discrepancy. Retinal expressed genes were not concentrated in particular areas of the canine genome, and also were located on the canine Y chromosome (CFAY). Several of the EST clones were located within areas of conserved synteny to human retinal disease loci. CONCLUSIONS: RH mapping of canine retinal ESTs provides insight into the location of potential candidate genes for hereditary retinal disorders, and, by comparison with the assembled canine genome sequence, highlights inconsistencies with the current assembly. Regions of conserved synteny between the canine and the human genomes allow this information to be extrapolated to identify potential positional candidate genes for mapped human retinal disorders. Furthermore, these ESTs can help identify novel or uncharacterized genes of significance for better understanding of retinal morphology, physiology, and pathology.
Assuntos
Cães/genética , Etiquetas de Sequências Expressas , Proteínas do Olho/genética , Mapeamento de Híbridos Radioativos , Retina/metabolismo , Doenças Retinianas/genética , Animais , Bases de Dados de Proteínas , Doenças do Cão/genética , Genoma , Genoma Humano , Humanos , Projetos de Pesquisa , Doenças Retinianas/veterináriaRESUMO
Specific cone-directed therapy is of high priority in the treatment of human hereditary retinal diseases. However, not much information exists about the specific targeting of photoreceptor subclasses. Three versions of the human red cone opsin promoter (PR0.5, 3LCR-PR0.5 and PR2.1), and the human blue cone opsin promoter HB569, were evaluated for their specificity and robustness in targeting green fluorescent protein (GFP) gene expression to subclasses of cones in the canine retina when used in recombinant adeno-associated viral vectors of serotype 5. The vectors were administered by subretinal injection. The promoter PR2.1 led to most effective and specific expression of GFP in the long- and medium-wavelength-absorbing cones (L/M cones) of normal and diseased retinas. The PR0.5 promoter was not effective. Adding three copies of the 35-bp LCR in front of PR0.5 lead to weak GFP expression in L/M cones. The HB569 promoter was not specific, and GFP was expressed in a few L/M cones, some rods and the retinal pigment epithelium. These results suggest that L/M cones, the predominant class of cone photoreceptors in the retinas of dogs and most mammalian species can be successfully targeted using the human red cone opsin promoter.
Assuntos
Terapia Genética/métodos , Regiões Promotoras Genéticas , Células Fotorreceptoras Retinianas Cones/metabolismo , Opsinas de Bastonetes/genética , Reparo Gênico Alvo-Dirigido , Animais , Defeitos da Visão Cromática/metabolismo , Defeitos da Visão Cromática/terapia , Dependovirus/genética , Cães , Expressão Gênica , Vetores Genéticos/administração & dosagem , Proteínas de Fluorescência Verde/genética , Humanos , Injeções , Modelos Animais , Transdução Genética/métodos , TransgenesRESUMO
Strains of silver foxes, selectively bred at the Institute of Cytology and Genetics of the Russian Academy of Sciences, are a well established, novel model for studying the genetic basis of behavior, and the processes involved in canine domestication. Here we describe a method to measure fox behavior as quantitative phenotypes which distinguish populations and resegregate in experimental pedigrees. We defined 50 binary observations that nonredundantly and accurately distinguished behaviors in reference populations and cross-bred pedigrees. Principal-component analysis dissected out the independent elements underlying these behaviors. PC1 accounted for >44% of the total variance in measured traits. This system clearly distinguished tame foxes from aggressive and wildtype foxes. F1 foxes yield intermediate values that extend into the ranges of both the tame and aggressive foxes, while the scores of the backcross generation resegregate. These measures can thus be used for QTL mapping to explore the genetic basis of tame and aggressive behavior in foxes, which should provide new insights into the mechanisms of mammalian behavior and canine domestication.
Assuntos
Agressão , Raposas/genética , Genoma , Locos de Características Quantitativas , Animais , Animais Domésticos/genética , Animais Selvagens/genética , Cruzamentos Genéticos , Cães/genética , Humanos , Modelos Genéticos , Linhagem , Gravação em VídeoRESUMO
B chromosomes are often considered to be one of the most mysterious elements of karyotypes (Camacho, 2004). It is generally believed that mammalian B chromosomes do not contain any protein coding genes. The discovery of a conserved KIT gene in Canidae B chromosomes has changed this view. Here we performed analysis of sequences surrounding KIT in B chromosomes of the fox and raccoon dog. The presence of the RPL23A pseudogene was shown in canid B chromosomes. The 3' end fragment of the KDR gene was found in raccoon dog B chromosomes. The size of the B-specific fragment homologous to the autosome fragment was estimated to be a minimum of 480 kbp in both species. The origin and evolution of B chromosomes in Canidae are discussed.
Assuntos
Mapeamento Cromossômico/métodos , Cromossomos/ultraestrutura , Proteínas Proto-Oncogênicas c-kit/genética , Animais , Bandeamento Cromossômico , Cães , Raposas , Biblioteca Gênica , Hibridização in Situ Fluorescente , Cariotipagem , Modelos Genéticos , Cães GuaxininsRESUMO
Recent advances in genomics resources and tools are facilitating quantitative trait locus mapping. We developed a crossbreed pedigree for mapping quantitative trait loci for hip dysplasia in dogs by crossing dysplastic Labrador Retrievers and normal Greyhounds. We show that one advantage to using a crossbreed pedigree is the increased marker informativeness in the backcross/F2 population relative to the founder populations. We also discuss three factors that affect the detection power in the context of this crossbreed pedigree: being able to detect and correct genotyping errors, increasing marker density for chromosomes with a sparse coverage, and adding individuals to the mapping population as soon as they become available.
Assuntos
Alelos , Displasia Pélvica Canina/genética , Repetições de Microssatélites/genética , Polimorfismo Genético/genética , Animais , Mapeamento Cromossômico/métodos , Cães , Feminino , Hibridização Genética , Masculino , Linhagem , Reprodutibilidade dos Testes , Projetos de Pesquisa , Estatística como Assunto/métodosRESUMO
The silver fox, a variant of the red fox (Vulpes vulpes), is a close relative of the dog (Canis familiaris). Cytogenetic differences and similarities between these species are well understood, but their genomic organizations have not been compared at higher resolution. Differences in their behavior also remain unexplained. Two silver fox strains demonstrating markedly different behavior have been generated at the Institute of Cytology and Genetics of the Russian Academy of Sciences. Foxes selected for tameness are friendly, like domestic dogs, while foxes selected for aggression resist human contact. To refine our understanding of the comparative genomic organization of dogs and foxes, and enable a study of the genetic basis of behavior in these fox strains, we need a meiotic linkage map of the fox. Towards this goal we generated a primary set of fox microsatellite markers. Four hundred canine microsatellites, evenly distributed throughout the canine genome, have been identified that amplify robustly from fox DNA. Polymorphism information content (PIC) values were calculated for a representative subset of these markers and population inbreeding coefficients were determined for tame and aggressive foxes. To begin to identify fox-specific single nucleotide polymorphisms (SNPs) in genes involved in the neurobiology of behavior, fox and dog orthologs of serotonin 5-HT1A and 5-HT1B receptor genes have been cloned. Sequence comparison of these genes from tame and aggressive foxes reveal several SNPs. The close relationship of the fox and dog enables canine genomic tools to be utilized in developing a fox meiotic map and mapping behavioral traits in the fox.
Assuntos
Agressão/fisiologia , Mapeamento Cromossômico , Raposas/genética , Animais , Sequência de Bases , Cromossomos de Mamíferos/genética , Primers do DNA , Raposas/fisiologia , Marcadores Genéticos , Genética Populacional , Repetições de Microssatélites/genética , Dados de Sequência Molecular , Polimorfismo Genético/genética , Polimorfismo de Nucleotídeo Único/genética , Receptores 5-HT1 de Serotonina/genética , Alinhamento de Sequência , Análise de Sequência de DNARESUMO
Progressive retinal atrophies (PRA) are a heterogeneous group of inherited eye diseases common to both dogs and man. Over 100 individual canine breeds display some sort of retinal degeneration, making the dog an extremely valuable resource both for finding the genetic determinants of inherited blindness and for developing naturally occurring animal models that mimic human disease. Progressive retinal atrophies within the English mastiff displayed an ambiguous mode of inheritance. By conducting outcross matings between affected English mastiffs and normal animals from other breeds, the mode of inheritance was confirmed as dominant. This directed candidate gene analysis and led to identification of two synonymous mutations and one nonsynonymous mutation within the canine rhodopsin gene. The nonsynonymous mutation (T4R) is the cause of PRA in the English mastiff, and a test was developed to investigate its presence in 17 additional breeds. Testing of PRA-affected animals from 16 breeds revealed that none carry the T4R mutation, indicating a different cause of PRA. Analysis of two affected bull mastiffs revealed one heterozygote (+/T4R) and one homozygous normal individual (+/+). These findings suggest that the genetic origin of PRA is often breed specific and underline the value of outcross mating to circumvent problems that act to mask the mode of inheritance.
Assuntos
Cruzamentos Genéticos , Cães/genética , Oftalmopatias Hereditárias/genética , Genes Dominantes , Rodopsinas Sensoriais/genética , Animais , Cães/metabolismo , Oftalmopatias Hereditárias/metabolismo , Feminino , Masculino , Modelos Genéticos , Linhagem , Rodopsinas Sensoriais/metabolismoRESUMO
An outcrossed canine pedigree was developed for quantitative trait locus (QTL) mapping of hip dysplasia by breeding dysplastic Labrador retrievers to trait-free greyhounds. Measured susceptibility traits included age at onset of femoral capital chondroepiphyseal ossification (OSS), maximum hip distraction (laxity) index (DI), and the dorsolateral subluxation (DLS) score. The pedigrees consisted of 147 dogs representing four generations. For 59 dogs genotyped with 65 microsatellites, the median heterozygosity and polymorphic information content (PIC) values of the F(1) generation were 0.82 and 0.68, respectively. Seventy-seven percent of microsatellites had a PIC greater than 0.59 in the F(1)s. Ninety-six percent of alleles showed Mendelian inheritance. Based on marker informativeness, approximately 350 randomly selected markers would be required for genome-wide screening to obtain an average interval between informative markers of 10 cM. Heritability was estimated as 0.43, 0.5, and 0.61 for OSS, DI, and the DLS score, respectively. Biometric estimates of the mean (+/- variance) effective number of segregating QTLs was 1.2 (+/- 0.05), 0.8 (+/- 0.02), and 1.0 (+/- 0.03) for OSS, DI, and the DLS score, respectively. The distributions of simulated backcross trait data suggested that the loci controlling these traits acted additively and that the DI may be controlled by a major locus. When combined with previous power and quantitative genetic analyses, these estimates indicate that this pedigree is informative for QTL mapping of hip dysplasia traits.
Assuntos
Cães/genética , Predisposição Genética para Doença , Displasia Pélvica Canina/genética , Repetições de Microssatélites , Animais , Mapeamento Cromossômico , Cruzamentos Genéticos , Feminino , Masculino , Linhagem , Locos de Características QuantitativasRESUMO
X-linked retinitis pigmentosa (XLRP) is one of the most prevalent forms of a genetically heterogeneous group of inherited retinal disorders of man; more than 70% of XLRP families map to the RP2 or RP3 loci on the human X chromosome. Canine X-linked progressive retinal atrophy (XLPRA), observed in the Siberian husky, is the locus homologue of human RP3, but the gene responsible for XLPRA has not yet been identified. To develop polymorphic markers in the RP3 interval in dogs we have isolated microsatellites from canine BAC clones. Three tightly linked microsatellite loci, CUX20001, CUX30001, and CUX40002, have been investigated in 17 dog breeds or breed varieties. Calculated parameters of variability correspond with the number of repeats at each locus. Pedigree analyses showed tight linkage between the canine t-complex-associated testis-expressed 1-like gene (TCTE1l) and the gene ornithine carbamoyltransferase (OTC). Each microsatellite shows conservation within Canidae, and CUX20001 also amplified in Mustelidae and URSIDAE: These markers represent an important tool in the fine mapping process for the canine region homologous to the RP3 disease interval and are valuable for evaluation of conservation and homology of this region among related species.
Assuntos
Cães/genética , Proteínas do Olho , Repetições de Microssatélites , Proteínas/genética , Animais , Cromossomos Artificiais Bacterianos , Variação Genética , Linhagem , Lobos/genética , Cromossomo XRESUMO
PURPOSE: Canine X-linked progressive retinal atrophy (XLPRA) is a hereditary, progressive retinal degeneration that has been mapped previously to the canine X chromosome in a region flanked by the dystrophin (DMD) and tissue inhibitor of metalloproteinase 1 (TIMP1) genes, and is tightly linked to the gene RPGR. The comparable region of the human X chromosome includes the disease locus for RP3, an X-linked form of retinitis pigmentosa, although the current canine disease interval is much larger. METHODS: To refine the map of the canine XLPRA disease interval, 11 X-linked markers were mapped, both meiotically, in two extensive canine pedigrees informative for XLPRA, and on a 3000-rad canine-hamster radiation hybrid (RH) panel. A 12th marker was mapped on the RH panel alone. RESULTS: The integrated map of this region of CFAX now covers approximately 47.3 centimorgans (cM) and 194 centirays (cR)(3000), and demonstrates strong conservation of synteny between humans and dogs. Genes defining the human RP3 zero-recombination interval (human homologue of mouse t complex [TCTE1L], sushi repeat-containing protein, X chromosome [SRPX], and retinitis pigmentosa guanosine triphosphatase [GTPase] regulator [RPGR]) are tightly linked to each other, to the XLPRA locus, and to the gene ornithine transcarbamylase (OTC) in dogs. CONCLUSIONS: Strong conservation of gene order was demonstrated in the short arm of the X chromosome between dogs and humans as was homology of the canine XLPRA and human RP3 intervals. These results create a valuable tool for investigating canine XLPRA and other X-linked eye diseases in dogs.
Assuntos
Mapeamento Cromossômico , Doenças do Cão/genética , Ligação Genética , Retina/patologia , Retinose Pigmentar/veterinária , Cromossomo X , Animais , Atrofia , Cromossomos Artificiais Bacterianos/genética , Primers do DNA/química , Progressão da Doença , Doenças do Cão/patologia , Cães , Feminino , Biblioteca Gênica , Genótipo , Humanos , Masculino , Repetições de Microssatélites , Linhagem , Reação em Cadeia da Polimerase , Retinose Pigmentar/genética , Retinose Pigmentar/patologiaRESUMO
The relationship between the neurosensory photoreceptors and the adjacent retinal pigment epithelium (RPE) controls not only normal retinal function, but also the pathogenesis of hereditary retinal degenerations. The molecular bases for both primary photoreceptor and RPE diseases that cause blindness have been identified. Gene therapy has been used successfully to slow degeneration in rodent models of primary photoreceptor diseases, but efficacy of gene therapy directed at photoreceptors and RPE in a large-animal model of human disease has not been reported. Here we study one of the most clinically severe retinal degenerations, Leber congenital amaurosis (LCA). LCA causes near total blindness in infancy and can result from mutations in RPE65 (LCA, type II; MIM 180069 and 204100). A naturally occurring animal model, the RPE65-/- dog, suffers from early and severe visual impairment similar to that seen in human LCA. We used a recombinant adeno-associated virus (AAV) carrying wild-type RPE65 (AAV-RPE65) to test the efficacy of gene therapy in this model. Our results indicate that visual function was restored in this large animal model of childhood blindness.
Assuntos
Cegueira/terapia , Modelos Animais de Doenças , Doenças do Cão/genética , Proteínas do Olho/genética , Terapia Genética/métodos , Atrofias Ópticas Hereditárias/terapia , Proteínas/genética , Animais , Animais Geneticamente Modificados , Proteínas de Transporte , Dependovirus/genética , Cães , cis-trans-IsomerasesRESUMO
PURPOSE: D-cis-diltiazem, a calcium channel blocker, has been reported to enhance photoreceptor survival in the rd mouse, a model of retinitis pigmentosa (RP) resulting from mutation of the PDE6B gene. We tested the hypothesis that diltiazem treatment would similarly rescue the canine rcd1 model of RP, which is also caused by a null mutation in the PDE6B gene. METHODS: D-cis-diltiazem was delivered orally twice daily to rcd1 affected dogs beginning at 4 weeks of age; untreated age-matched rcd1 dogs served as controls. At 14 weeks, electroretinograms (ERG) were performed on all animals; 14 dogs were euthanized at this age, and 2 dogs at 25 weeks of age. Eyes were enucleated, fixed, and processed for routine histological examination. RESULTS: No significant differences were found in ERG or histopathologic parameters between diltiazem-treated and untreated rcd1 dogs. Neither rcd1 group showed a rod b-wave; ERGs evoked by single white flashes (dark- or light-adapted) and flicker were also identical between groups. Similarly, treated and untreated animals did not differ in the degree of preservation of the photoreceptor layer, confirmed in cell counts within the outer nuclear layer. CONCLUSIONS: Treatment of rcd1 affected dogs with D-cis-diltiazem did not modify the photoreceptor disease when results were assessed using either ERG or histopathologic criteria. The positive photoreceptor-rescue effect of calcium channel blockers reported in the rd mouse was thus not generalizable to another species with retinal degeneration due to mutation in the PDE6B gene. Caution needs to be exerted in extrapolation to the comparable human forms of RP.
Assuntos
3',5'-GMP Cíclico Fosfodiesterases/genética , Bloqueadores dos Canais de Cálcio/uso terapêutico , Diltiazem/uso terapêutico , Proteínas do Olho/genética , Células Fotorreceptoras de Vertebrados/efeitos dos fármacos , Retinose Pigmentar/tratamento farmacológico , Animais , Nucleotídeo Cíclico Fosfodiesterase do Tipo 6 , Adaptação à Escuridão , Modelos Animais de Doenças , Cães , Eletrorretinografia , Feminino , Masculino , Estimulação Luminosa , Células Fotorreceptoras de Vertebrados/patologia , Células Fotorreceptoras de Vertebrados/fisiologia , Retinose Pigmentar/genética , Retinose Pigmentar/fisiopatologiaRESUMO
One hundred and sixteen interspersed repetitive DNA sequence (IRS)-PCR markers have been developed and characterized from Canis familiaris for high-throughput filter-based genotyping. We present a detailed analysis of markers produced by amplification using primers directed to the conserved regions of the C. familiaris short interspersed nuclear element (Can-SINE). The majority of IRS-PCR markers developed were moderately to highly polymorphic with mean heterozygosity (HET) and polymorphism information content (PIC) values of approximately 0.6. The HET value for 22.3% of the markers exceeded 0.7. We also demonstrate that sequence variation of Can-SINEs between breeds is significant and also represents a rich source of polymorphisms. Mapping of 73 of the markers to the existing integrated linkage-radiation hybrid map enriches the map as well as establishes the utility of the markers. The significance and utility of this new class of IRS-PCR Can-SINE-based markers for high-throughput genotyping is discussed. This method can also be extended to other species that are currently map-poor but have a sufficiently high density of SINEs to allow IRS-PCR.
Assuntos
Cães/genética , Elementos Nucleotídeos Curtos e Dispersos/genética , Animais , Sequência Conservada , Ligação Genética , Marcadores Genéticos/genética , Genótipo , Reação em Cadeia da Polimerase/veterinária , Polimorfismo Genético/genética , Polimorfismo Conformacional de Fita Simples , Mapeamento de Híbridos Radioativos/veterináriaRESUMO
The disease phenotype of oculo-skeletal dysplasia (OSD) detected in Labrador retrievers and Samoyeds shows a large degree of similarity with human Stickler and Kniest dysplasia. Type II collagen (COL2A1) mRNA, which is defective in a larger number of Stickler and Kniest patients, has been cloned and characterized from normal dog. The amino acid sequence of the canine type II procollagen is predicted to contain 1487 residues, with high degree of homology with its human homologue, and maintains all the characteristic structural domains. In addition to cartilage, expression of COL2A1 has also been detected in canine retina and testes. In testes, the N-propeptide region of COL2A1 displayed differential splicing and expressed both splice variants, IIA (with exon 2) and IIB (without exon 2), suggesting the importance of both forms in testis maturation and maintenance. Despite a severe decrease of type II collagen protein in the vitreous of OSD affected Labrador retrievers, COL2A1 gene has been excluded from having any causal association with the disease locus by linkage analysis. Using an intragenic RFLP marker, COL2A1 gene has also been tested as a candidate gene for the non-allelic form of the other canine OSD identified in Samoyeds, and excluded by linkage analysis. Oculo-skeletal dysplastic Labrador retriever and Samoyed provide two animal models for chondrodysplasia with genetic heterogeneity.
Assuntos
Colágeno/genética , RNA Mensageiro/metabolismo , Sequência de Aminoácidos , Animais , Sequência de Bases , Cartilagem/anormalidades , Cartilagem/metabolismo , Clonagem Molecular , DNA Complementar/química , DNA Complementar/genética , Cães , Feminino , Expressão Gênica , Genótipo , Masculino , Dados de Sequência Molecular , Linhagem , RNA/genética , RNA/metabolismo , RNA Mensageiro/genética , Retina/anormalidades , Retina/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Análise de Sequência de DNA , Distribuição Tecidual , Corpo Vítreo/metabolismoRESUMO
X-linked progressive retinal atrophy (XLPRA) in the Siberian husky dog is a naturally occurring X-linked retinopathy closely resembling X-linked retinitis pigmentosa (XLRP) in humans. In affected males, initial degeneration of rods is followed by cone degeneration and complete retinal atrophy; carrier females have random patches of rod degeneration consistent with random X chromosome inactivation. By typing the XLPRA pedigree with five intragenic markers [dystrophin, retinitis pigmentosa GTPase regulator ( RPGR ), tissue inhibitor of metalloproteinases 1, androgen receptor and factor IX], we established a linkage map of the canine X chromosome, and confirmed that the order of these five genes is identical to that on the human X. XLPRA was tightly linked to an intragenic RPGR polymorphism (LOD 11.7, zero recombination), thus confirming locus homology with RP3. We cloned the full-length canine RPGR cDNA and three additional splice variants. No disease-causing mutation was found in the RPGR-coding sequence of the four splice variants characterized, a finding similar to approximately 80% of human XLRP patients whose disease maps to the RP3 locus. In addition, there were no significant differences in the proportional expression of each splice variant in normal and pre-degenerate XLPRA-affected retina. Expression of all RPGR splice variants increased later in the disease, when retinas were undergoing active degeneration. The results provide further evidence of cross-species retention of a complex splicing pattern in the 3' portion of RPGR, the functional significance of which is unknown. In addition, the possibility of another disease locus in the RP3 region is supported.
Assuntos
Mapeamento Cromossômico/veterinária , Doenças do Cão/genética , Proteínas do Olho , Ligação Genética , Retinose Pigmentar/veterinária , Cromossomo X/genética , Processamento Alternativo , Animais , Proteínas de Transporte/genética , Doenças do Cão/patologia , Cães , Feminino , Marcadores Genéticos , Masculino , Dados de Sequência Molecular , Especificidade de Órgãos/genética , Proteínas/genética , Retina/patologia , Retinose Pigmentar/genética , Retinose Pigmentar/patologia , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Análise de Sequência de DNARESUMO
A novel gene expressed predominantly in retina, but detected at a conspicuously lower level in retina of canine progressive rod cone degeneration (prcd), has been identified by suppression subtractive hybridization and retinal cDNA library screening. The characterized region of cDNA of the novel gene includes 1017 nucleotides of coding sequence predicted to encode a protein of 338 amino acids (M(r) 39389), 791 nucleotides of 5'-untranslated region (UTR), and 300 nucleotides of 3'-UTR including the poly(A)(+) tail. Multiple transcripts were detected in retina by Northern blot analysis, and a lower level of expression was observed in brain and liver by RT-PCR. The transcript appears to be developmentally regulated with a burst in gene expression at a time period (34 postnatal days) that coincides with the photoreceptor differentiation phase of retinal development. The deduced amino acid sequence from the cDNA of the novel gene has 24% identity and 48% similarity with the multifunctional glycoprotein clusterin. Hence, the putative gene product from the novel transcript has been named clusterin-like protein 1 (CLUL1). The human homologue of CLUL1 cDNA has 84 and 70% identity at the level of nucleotides and amino acids, respectively, with the characterized canine cDNA. The presence of a stretch of 128 amino acids in the putative human CLUL1, not detected in canine CLUL1, suggests alternate splicing events. An STS database search revealed that the human homologue of CLUL1 maps to chromosome 18p, a location not yet reported to harbor an RP locus. Tissue-specific expression of CLUL1 in retina, and its lower abundance in different forms of PRA suggest that this novel gene may represent an as-yet unidentified locus for a retinal disorder.