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1.
Am J Hum Genet ; 104(3): 422-438, 2019 Mar 07.
Artigo em Inglês | MEDLINE | ID: mdl-30773277

RESUMO

SPONASTRIME dysplasia is an autosomal-recessive spondyloepimetaphyseal dysplasia characterized by spine (spondylar) abnormalities, midface hypoplasia with a depressed nasal bridge, metaphyseal striations, and disproportionate short stature. Scoliosis, coxa vara, childhood cataracts, short dental roots, and hypogammaglobulinemia have also been reported in this disorder. Although an autosomal-recessive inheritance pattern has been hypothesized, pathogenic variants in a specific gene have not been discovered in individuals with SPONASTRIME dysplasia. Here, we identified bi-allelic variants in TONSL, which encodes the Tonsoku-like DNA repair protein, in nine subjects (from eight families) with SPONASTRIME dysplasia, and four subjects (from three families) with short stature of varied severity and spondylometaphyseal dysplasia with or without immunologic and hematologic abnormalities, but no definitive metaphyseal striations at diagnosis. The finding of early embryonic lethality in a Tonsl-/- murine model and the discovery of reduced length, spinal abnormalities, reduced numbers of neutrophils, and early lethality in a tonsl-/- zebrafish model both support the hypomorphic nature of the identified TONSL variants. Moreover, functional studies revealed increased amounts of spontaneous replication fork stalling and chromosomal aberrations, as well as fewer camptothecin (CPT)-induced RAD51 foci in subject-derived cell lines. Importantly, these cellular defects were rescued upon re-expression of wild-type (WT) TONSL; this rescue is consistent with the hypothesis that hypomorphic TONSL variants are pathogenic. Overall, our studies in humans, mice, zebrafish, and subject-derived cell lines confirm that pathogenic variants in TONSL impair DNA replication and homologous recombination-dependent repair processes, and they lead to a spectrum of skeletal dysplasia phenotypes with numerous extra-skeletal manifestations.

2.
Cell Rep ; 25(5): 1281-1291.e4, 2018 Oct 30.
Artigo em Inglês | MEDLINE | ID: mdl-30380418

RESUMO

Morphogenesis and mechanoelectrical transduction of the hair cell mechanoreceptor depend on the correct assembly of Usher syndrome (USH) proteins into highly organized macromolecular complexes. Defects in these proteins lead to deafness and vestibular areflexia in USH patients. Mutations in a non-USH protein, glutaredoxin domain-containing cysteine-rich 1 (GRXCR1), cause non-syndromic sensorineural deafness. To understand the deglutathionylating enzyme function of GRXCR1 in deafness, we generated two grxcr1 zebrafish mutant alleles. We found that hair bundles are thinner in homozygous grxcr1 mutants, similar to the USH1 mutants ush1c (Harmonin) and ush1ga (Sans). In vitro assays showed that glutathionylation promotes the interaction between Ush1c and Ush1ga and that Grxcr1 regulates mechanoreceptor development by preventing physical interaction between these proteins without affecting the assembly of another USH1 protein complex, the Ush1c-Cadherin23-Myosin7aa tripartite complex. By elucidating the molecular mechanism through which Grxcr1 functions, we also identify a mechanism that dynamically regulates the formation of Usher protein complexes.

3.
Nucleic Acids Res ; 2018 Nov 08.
Artigo em Inglês | MEDLINE | ID: mdl-30407545

RESUMO

The Zebrafish Information Network (ZFIN) (https://zfin.org/) is the database for the model organism, zebrafish (Danio rerio). ZFIN expertly curates, organizes and provides a wide array of zebrafish genetic and genomic data, including genes, alleles, transgenic lines, gene expression, gene function, mutant phenotypes, orthology, human disease models, nomenclature and reagents. New features at ZFIN include increased support for genomic regions and for non-coding genes, and support for more expressive Gene Ontology annotations. ZFIN has recently taken over maintenance of the zebrafish reference genome sequence as part of the Genome Reference Consortium. ZFIN is also a founding member of the Alliance of Genome Resources, a collaboration of six model organism databases (MODs) and the Gene Ontology Consortium (GO). The recently launched Alliance portal (https://alliancegenome.org) provides a unified, comparative view of MOD, GO, and human data, and facilitates foundational and translational biomedical research.

4.
Mech Dev ; 2018 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-30287385

RESUMO

The semicircular canals in the inner ear sense angular acceleration. In zebrafish, the semicircular canals develop from epithelial projections that grow toward each other and fuse to form pillars. The growth of the epithelial projections is driven by the production and secretion of extracellular matrix components by the epithelium. The conserved oligomeric Golgi 4 protein, Cog4, functions in retrograde vesicle transport within the Golgi and mutations can lead to sensory neural hearing loss. In zebrafish cog4 mutants, the inner ear is smaller and the number of hair cells is reduced. Here, we show that formation of the pillars is delayed and that secretion of extracellular matrix components (ECM) is impaired in cog4-/- mutants. These results show that Cog4 is required for secretion of ECM molecules essential to drive the growth of the epithelial projections and thus regulates morphogenesis of the semicircular canals.

5.
N Engl J Med ; 379(22): 2131-2139, 2018 11 29.
Artigo em Inglês | MEDLINE | ID: mdl-30304647

RESUMO

BACKGROUND: Many patients remain without a diagnosis despite extensive medical evaluation. The Undiagnosed Diseases Network (UDN) was established to apply a multidisciplinary model in the evaluation of the most challenging cases and to identify the biologic characteristics of newly discovered diseases. The UDN, which is funded by the National Institutes of Health, was formed in 2014 as a network of seven clinical sites, two sequencing cores, and a coordinating center. Later, a central biorepository, a metabolomics core, and a model organisms screening center were added. METHODS: We evaluated patients who were referred to the UDN over a period of 20 months. The patients were required to have an undiagnosed condition despite thorough evaluation by a health care provider. We determined the rate of diagnosis among patients who subsequently had a complete evaluation, and we observed the effect of diagnosis on medical care. RESULTS: A total of 1519 patients (53% female) were referred to the UDN, of whom 601 (40%) were accepted for evaluation. Of the accepted patients, 192 (32%) had previously undergone exome sequencing. Symptoms were neurologic in 40% of the applicants, musculoskeletal in 10%, immunologic in 7%, gastrointestinal in 7%, and rheumatologic in 6%. Of the 382 patients who had a complete evaluation, 132 received a diagnosis, yielding a rate of diagnosis of 35%. A total of 15 diagnoses (11%) were made by clinical review alone, and 98 (74%) were made by exome or genome sequencing. Of the diagnoses, 21% led to recommendations regarding changes in therapy, 37% led to changes in diagnostic testing, and 36% led to variant-specific genetic counseling. We defined 31 new syndromes. CONCLUSIONS: The UDN established a diagnosis in 132 of the 382 patients who had a complete evaluation, yielding a rate of diagnosis of 35%. (Funded by the National Institutes of Health Common Fund.).

6.
Exp Eye Res ; 173: 148-159, 2018 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-29777677

RESUMO

Mutations in USH2A are the most frequent cause of Usher syndrome and autosomal recessive nonsyndromic retinitis pigmentosa. To unravel the pathogenic mechanisms underlying USH2A-associated retinal degeneration and to evaluate future therapeutic strategies that could potentially halt the progression of this devastating disorder, an animal model is needed. The available Ush2a knock-out mouse model does not mimic the human phenotype, because it presents with only a mild and late-onset retinal degeneration. Using CRISPR/Cas9-technology, we introduced protein-truncating germline lesions into the zebrafish ush2a gene (ush2armc1: c.2337_2342delinsAC; p.Cys780GlnfsTer32 and ush2ab1245: c.15520_15523delinsTG; p.Ala5174fsTer). Homozygous mutants were viable and displayed no obvious morphological or developmental defects. Immunohistochemical analyses with antibodies recognizing the N- or C-terminal region of the ush2a-encoded protein, usherin, demonstrated complete absence of usherin in photoreceptors of ush2armc1, but presence of the ectodomain of usherin at the periciliary membrane of ush2ab1245-derived photoreceptors. Furthermore, defects of usherin led to a reduction in localization of USH2 complex members, whirlin and Adgrv1, at the photoreceptor periciliary membrane of both mutants. Significantly elevated levels of apoptotic photoreceptors could be observed in both mutants when kept under constant bright illumination for three days. Electroretinogram (ERG) recordings revealed a significant and similar decrease in both a- and b-wave amplitudes in ush2armc1 as well as ush2ab1245 larvae as compared to strain- and age-matched wild-type larvae. In conclusion, this study shows that mutant ush2a zebrafish models present with early-onset retinal dysfunction that is exacerbated by light exposure. These models provide a better understanding of the pathophysiology underlying USH2A-associated RP and a unique opportunity to evaluate future therapeutic strategies.

7.
Zebrafish ; 15(3): 279-290, 2018 06.
Artigo em Inglês | MEDLINE | ID: mdl-29369744

RESUMO

Sperm cryopreservation is a highly efficient method for preserving genetic resources. It extends the reproductive period of males and significantly reduces costs normally associated with maintenance of live animal colonies. However, previous zebrafish (Danio rerio) cryopreservation methods have produced variable outcomes and low post-thaw fertilization rates. To improve post-thaw fertilization rates after cryopreservation, we developed a new extender and cryoprotective medium (CPM), introduced quality assessment (QA), determined the optimal cooling rate, and improved the post-thaw in vitro fertilization process. We found that the hypertonic extender E400 preserved motility of sperm held on ice for at least 6 h. We implemented QA by measuring sperm cell densities with a NanoDrop spectrophotometer and sperm motility with computer-assisted sperm analysis (CASA). We developed a CPM, RMMB, which contains raffinose, skim milk, methanol, and bicine buffer. Post-thaw motility indicated that the optimal cooling rate in two types of cryogenic vials was between 10 and 15°C/min. Test thaws from this method produced average motility of 20% ± 13% and an average post-thaw fertilization rate of 68% ± 16%.


Assuntos
Criopreservação/métodos , Crioprotetores/química , Técnicas de Cultura/métodos , Fertilização In Vitro/métodos , Preservação do Sêmen/métodos , Espermatozoides/fisiologia , Peixe-Zebra/fisiologia , Animais , Masculino , Motilidade Espermática
8.
Genetics ; 207(1): 9-27, 2017 09.
Artigo em Inglês | MEDLINE | ID: mdl-28874452

RESUMO

Efforts to identify the genetic underpinnings of rare undiagnosed diseases increasingly involve the use of next-generation sequencing and comparative genomic hybridization methods. These efforts are limited by a lack of knowledge regarding gene function, and an inability to predict the impact of genetic variation on the encoded protein function. Diagnostic challenges posed by undiagnosed diseases have solutions in model organism research, which provides a wealth of detailed biological information. Model organism geneticists are by necessity experts in particular genes, gene families, specific organs, and biological functions. Here, we review the current state of research into undiagnosed diseases, highlighting large efforts in North America and internationally, including the Undiagnosed Diseases Network (UDN) (Supplemental Material, File S1) and UDN International (UDNI), the Centers for Mendelian Genomics (CMG), and the Canadian Rare Diseases Models and Mechanisms Network (RDMM). We discuss how merging human genetics with model organism research guides experimental studies to solve these medical mysteries, gain new insights into disease pathogenesis, and uncover new therapeutic strategies.


Assuntos
Modelos Animais de Doenças , Doenças Genéticas Inatas/genética , Doenças Raras/genética , Animais , Drosophila/genética , Exoma , Doenças Genéticas Inatas/diagnóstico , Doenças Genéticas Inatas/terapia , Testes Genéticos/métodos , Humanos , Doenças Raras/diagnóstico , Doenças Raras/terapia , Peixe-Zebra/genética
9.
ILAR J ; 58(1): 4-16, 2017 07 01.
Artigo em Inglês | MEDLINE | ID: mdl-28838067

RESUMO

The Zebrafish Model Organism Database (ZFIN; https://zfin.org) is the central resource for genetic, genomic, and phenotypic data for zebrafish (Danio rerio) research. ZFIN continuously assesses trends in zebrafish research, adding new data types and providing data repositories and tools that members of the research community can use to navigate data. The many research advantages and flexibility of manipulation of zebrafish have made them an increasingly attractive animal to model and study human disease.To facilitate disease-related research, ZFIN developed support to provide human disease information as well as annotation of zebrafish models of human disease. Human disease term pages at ZFIN provide information about disease names, synonyms, and references to other databases as well as a list of publications reporting studies of human diseases in which zebrafish were used. Zebrafish orthologs of human genes that are implicated in human disease etiology are routinely studied to provide an understanding of the molecular basis of disease. Therefore, a list of human genes involved in the disease with their corresponding zebrafish ortholog is displayed on the disease page, with links to additional information regarding the genes and existing mutations. Studying human disease often requires the use of models that recapitulate some or all of the pathologies observed in human diseases. Access to information regarding existing and published models can be critical, because they provide a tractable way to gain insight into the phenotypic outcomes of the disease. ZFIN annotates zebrafish models of human disease and supports retrieval of these published models by listing zebrafish models on the disease term page as well as by providing search interfaces and data download files to access the data. The improvements ZFIN has made to annotate, display, and search data related to human disease, especially zebrafish models for disease and disease-associated gene information, should be helpful to researchers and clinicians considering the use of zebrafish to study human disease.


Assuntos
Bases de Dados Genéticas , Modelos Animais de Doenças , Proteínas de Peixe-Zebra/genética , Peixe-Zebra/genética , Animais , Biologia Computacional/métodos , Curadoria de Dados/métodos , Estudos de Associação Genética , Genoma , Genômica , Humanos , Modelos Animais , Mutação
10.
Cell ; 169(1): 6-12, 2017 03 23.
Artigo em Inglês | MEDLINE | ID: mdl-28340351

RESUMO

Genome sequencing has revolutionized the diagnosis of genetic diseases. Close collaborations between basic scientists and clinical genomicists are now needed to link genetic variants with disease causation. To facilitate such collaborations, we recommend prioritizing clinically relevant genes for functional studies, developing reference variant-phenotype databases, adopting phenotype description standards, and promoting data sharing.


Assuntos
Pesquisa Biomédica , Genômica , Animais , Análise Mutacional de DNA , Bases de Dados Genéticas , Doença/genética , Projeto Genoma Humano , Humanos , Disseminação de Informação , Modelos Animais
11.
Nucleic Acids Res ; 45(D1): D758-D768, 2017 01 04.
Artigo em Inglês | MEDLINE | ID: mdl-27899582

RESUMO

The Zebrafish Model Organism Database (ZFIN; http://zfin.org) is the central resource for zebrafish (Danio rerio) genetic, genomic, phenotypic and developmental data. ZFIN curators provide expert manual curation and integration of comprehensive data involving zebrafish genes, mutants, transgenic constructs and lines, phenotypes, genotypes, gene expressions, morpholinos, TALENs, CRISPRs, antibodies, anatomical structures, models of human disease and publications. We integrate curated, directly submitted, and collaboratively generated data, making these available to zebrafish research community. Among the vertebrate model organisms, zebrafish are superbly suited for rapid generation of sequence-targeted mutant lines, characterization of phenotypes including gene expression patterns, and generation of human disease models. The recent rapid adoption of zebrafish as human disease models is making management of these data particularly important to both the research and clinical communities. Here, we describe recent enhancements to ZFIN including use of the zebrafish experimental conditions ontology, 'Fish' records in the ZFIN database, support for gene expression phenotypes, models of human disease, mutation details at the DNA, RNA and protein levels, and updates to the ZFIN single box search.


Assuntos
Bases de Dados Genéticas , Estudos de Associação Genética/métodos , Genômica/métodos , Ferramenta de Busca , Peixe-Zebra/genética , Animais , Biologia Computacional/métodos , Curadoria de Dados , Modelos Animais de Doenças , Expressão Gênica , Predisposição Genética para Doença , Genótipo , Humanos , Mutação , Fenótipo
12.
PLoS Genet ; 12(5): e1006054, 2016 05.
Artigo em Inglês | MEDLINE | ID: mdl-27195754

RESUMO

Hedgehog (Hh) signaling regulates multiple aspects of metazoan development and tissue homeostasis, and is constitutively active in numerous cancers. We identified Ubr3, an E3 ubiquitin ligase, as a novel, positive regulator of Hh signaling in Drosophila and vertebrates. Hh signaling regulates the Ubr3-mediated poly-ubiquitination and degradation of Cos2, a central component of Hh signaling. In developing Drosophila eye discs, loss of ubr3 leads to a delayed differentiation of photoreceptors and a reduction in Hh signaling. In zebrafish, loss of Ubr3 causes a decrease in Shh signaling in the developing eyes, somites, and sensory neurons. However, not all tissues that require Hh signaling are affected in zebrafish. Mouse UBR3 poly-ubiquitinates Kif7, the mammalian homologue of Cos2. Finally, loss of UBR3 up-regulates Kif7 protein levels and decreases Hh signaling in cultured cells. In summary, our work identifies Ubr3 as a novel, evolutionarily conserved modulator of Hh signaling that boosts Hh in some tissues.


Assuntos
Proteínas de Drosophila/genética , Olho/metabolismo , Cinesina/genética , Ubiquitina-Proteína Ligases/genética , Animais , Proteínas de Drosophila/metabolismo , Drosophila melanogaster/genética , Drosophila melanogaster/crescimento & desenvolvimento , Olho/crescimento & desenvolvimento , Proteínas Hedgehog/genética , Cinesina/metabolismo , Camundongos , Células Fotorreceptoras/metabolismo , Poliubiquitina , Proteólise , RNA Interferente Pequeno , Transdução de Sinais , Ubiquitina-Proteína Ligases/metabolismo , Ubiquitinação , Peixe-Zebra/genética
13.
Mol Biol Evol ; 33(1): 13-24, 2016 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-26500251

RESUMO

Phenotypes resulting from mutations in genetic model organisms can help reveal candidate genes for evolutionarily important phenotypic changes in related taxa. Although testing candidate gene hypotheses experimentally in nonmodel organisms is typically difficult, ontology-driven information systems can help generate testable hypotheses about developmental processes in experimentally tractable organisms. Here, we tested candidate gene hypotheses suggested by expert use of the Phenoscape Knowledgebase, specifically looking for genes that are candidates responsible for evolutionarily interesting phenotypes in the ostariophysan fishes that bear resemblance to mutant phenotypes in zebrafish. For this, we searched ZFIN for genetic perturbations that result in either loss of basihyal element or loss of scales phenotypes, because these are the ancestral phenotypes observed in catfishes (Siluriformes). We tested the identified candidate genes by examining their endogenous expression patterns in the channel catfish, Ictalurus punctatus. The experimental results were consistent with the hypotheses that these features evolved through disruption in developmental pathways at, or upstream of, brpf1 and eda/edar for the ancestral losses of basihyal element and scales, respectively. These results demonstrate that ontological annotations of the phenotypic effects of genetic alterations in model organisms, when aggregated within a knowledgebase, can be used effectively to generate testable, and useful, hypotheses about evolutionary changes in morphology.


Assuntos
Peixes-Gato/genética , Evolução Molecular , Expressão Gênica , Modelos Genéticos , Fenótipo , Animais , Biologia Computacional , Expressão Gênica/genética , Expressão Gênica/fisiologia , Software
14.
Genesis ; 53(8): 449, 2015 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-26249149
15.
Genesis ; 53(8): 498-509, 2015 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-26097180

RESUMO

The Zebrafish Model Organism Database (ZFIN; http://zfin.org) is the central resource for genetic and genomic data from zebrafish (Danio rerio) research. ZFIN staff curate detailed information about genes, mutants, genotypes, reporter lines, sequences, constructs, antibodies, knockdown reagents, expression patterns, phenotypes, gene product function, and orthology from publications. Researchers can submit mutant, transgenic, expression, and phenotype data directly to ZFIN and use the ZFIN Community Wiki to share antibody and protocol information. Data can be accessed through topic-specific searches, a new site-wide search, and the data-mining resource ZebrafishMine (http://zebrafishmine.org). Data download and web service options are also available. ZFIN collaborates with major bioinformatics organizations to verify and integrate genomic sequence data, provide nomenclature support, establish reciprocal links, and participate in the development of standardized structured vocabularies (ontologies) used for data annotation and searching. ZFIN-curated gene, function, expression, and phenotype data are available for comparative exploration at several multi-species resources. The use of zebrafish as a model for human disease is increasing. ZFIN is supporting this growing area with three major projects: adding easy access to computed orthology data from gene pages, curating details of the gene expression pattern changes in mutant fish, and curating zebrafish models of human diseases.


Assuntos
Bases de Dados Genéticas , Proteínas de Peixe-Zebra/genética , Peixe-Zebra/genética , Animais , Biologia Computacional/métodos , Curadoria de Dados/métodos , Estudos de Associação Genética , Genômica/métodos , Internet , Modelos Animais
16.
Genesis ; 53(8): 547-60, 2015 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-26097192

RESUMO

InterMine is a data integration warehouse and analysis software system developed for large and complex biological data sets. Designed for integrative analysis, it can be accessed through a user-friendly web interface. For bioinformaticians, extensive web services as well as programming interfaces for most common scripting languages support access to all features. The web interface includes a useful identifier look-up system, and both simple and sophisticated search options. Interactive results tables enable exploration, and data can be filtered, summarized, and browsed. A set of graphical analysis tools provide a rich environment for data exploration including statistical enrichment of sets of genes or other entities. InterMine databases have been developed for the major model organisms, budding yeast, nematode worm, fruit fly, zebrafish, mouse, and rat together with a newly developed human database. Here, we describe how this has facilitated interoperation and development of cross-organism analysis tools and reports. InterMine as a data exploration and analysis tool is also described. All the InterMine-based systems described in this article are resources freely available to the scientific community.


Assuntos
Bases de Dados Factuais , Software , Animais , Biologia Computacional/métodos , Bases de Dados Genéticas , Genômica , Humanos , Internet , Integração de Sistemas , Interface Usuário-Computador
17.
Hum Mol Genet ; 24(9): 2594-603, 2015 May 01.
Artigo em Inglês | MEDLINE | ID: mdl-25616960

RESUMO

Determination of variant pathogenicity represents a major challenge in the era of high-throughput sequencing. Erroneous categorization may result if variants affect genes that are in fact dispensable. We demonstrate that this also applies to rare, apparently unambiguous truncating mutations of an established disease gene. By whole-exome sequencing (WES) in a consanguineous family with congenital non-syndromic deafness, we unexpectedly identified a homozygous nonsense variant, p.Arg1066*, in AHI1, a gene associated with Joubert syndrome (JBTS), a severe recessive ciliopathy. None of four homozygotes expressed any signs of JBTS, and one of them had normal hearing, which also ruled out p.Arg1066* as the cause of deafness. Homozygosity mapping and WES in the only other reported JBTS family with a homozygous C-terminal truncation (p.Trp1088Leufs*16) confirmed AHI1 as disease gene, but based on a more N-terminal missense mutation impairing WD40-repeat formation. Morpholinos against N-terminal zebrafish Ahi1, orthologous to where human mutations cluster, produced a ciliopathy, but targeting near human p.Arg1066 and p.Trp1088 did not. Most AHI1 mutations in JBTS patients result in truncated protein lacking WD40-repeats and the SH3 domain; disease was hitherto attributed to loss of these protein interaction modules. Our findings indicate that normal development does not require the C-terminal SH3 domain. This has far-reaching implications, considering that variants like p.Glu984* identified by preconception screening ('Kingsmore panel') do not necessarily indicate JBTS carriership. Genomes of individuals with consanguineous background are enriched for homozygous variants that may unmask dispensable regions of disease genes and unrecognized false positives in diagnostic large-scale sequencing and preconception carrier screening.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/genética , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Estudos de Associação Genética , Mutação , Anormalidades Múltiplas/diagnóstico , Anormalidades Múltiplas/genética , Proteínas Adaptadoras de Transdução de Sinal/química , Animais , Encéfalo/patologia , Cerebelo/anormalidades , Mapeamento Cromossômico , Consanguinidade , Análise Mutacional de DNA , Modelos Animais de Doenças , Evolução Molecular , Exoma , Anormalidades do Olho/diagnóstico , Anormalidades do Olho/genética , Feminino , Ordem dos Genes , Genes Recessivos , Loci Gênicos , Heterozigoto , Sequenciamento de Nucleotídeos em Larga Escala , Homozigoto , Humanos , Doenças Renais Císticas/diagnóstico , Doenças Renais Císticas/genética , Imagem por Ressonância Magnética , Masculino , Modelos Moleculares , Linhagem , Conformação Proteica , Retina/anormalidades , Peixe-Zebra/genética
18.
Hum Mutat ; 35(10): 1153-62, 2014 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-25044745

RESUMO

We describe a consanguineous Iraqi family with Leber congenital amaurosis (LCA), Joubert syndrome (JBTS), and polycystic kidney disease (PKD). Targeted next-generation sequencing for excluding mutations in known LCA and JBTS genes, homozygosity mapping, and whole-exome sequencing identified a homozygous missense variant, c.317G>C (p.Arg106Pro), in POC1B, a gene essential for ciliogenesis, basal body, and centrosome integrity. In silico modeling suggested a requirement of p.Arg106 for the formation of the third WD40 repeat and a protein interaction interface. In human and mouse retina, POC1B localized to the basal body and centriole adjacent to the connecting cilium of photoreceptors and in synapses of the outer plexiform layer. Knockdown of Poc1b in zebrafish caused cystic kidneys and retinal degeneration with shortened and reduced photoreceptor connecting cilia, compatible with the human syndromic ciliopathy. A recent study describes homozygosity for p.Arg106ProPOC1B in a family with nonsyndromic cone-rod dystrophy. The phenotype associated with homozygous p.Arg106ProPOC1B may thus be highly variable, analogous to homozygous p.Leu710Ser in WDR19 causing either isolated retinitis pigmentosa or Jeune syndrome. Our study indicates that POC1B is required for retinal integrity, and we propose POC1B mutations as a probable cause for JBTS with severe PKD.


Assuntos
Proteínas de Ciclo Celular/genética , Doenças Cerebelares/genética , Anormalidades do Olho/genética , Doenças Renais Císticas/genética , Mutação , Retina/anormalidades , Anormalidades Múltiplas , Motivos de Aminoácidos , Sequência de Aminoácidos , Animais , Proteínas de Ciclo Celular/metabolismo , Doenças Cerebelares/metabolismo , Doenças Cerebelares/patologia , Cerebelo/anormalidades , Criança , Cílios/metabolismo , Cílios/ultraestrutura , Anormalidades do Olho/metabolismo , Anormalidades do Olho/patologia , Técnicas de Silenciamento de Genes , Humanos , Iraque , Rim/patologia , Doenças Renais Císticas/metabolismo , Doenças Renais Císticas/patologia , Amaurose Congênita de Leber/genética , Amaurose Congênita de Leber/metabolismo , Masculino , Camundongos , Dados de Sequência Molecular , Linhagem , Retina/metabolismo , Retina/patologia , Peixe-Zebra
19.
Dis Model Mech ; 7(7): 739-43, 2014 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-24973743

RESUMO

Advances in genomics and next-generation sequencing have provided clinical researchers with unprecedented opportunities to understand the molecular basis of human genetic disorders. This abundance of information places new requirements on traditional disease models, which have the potential to be used to confirm newly identified pathogenic mutations and test the efficacy of emerging therapies. The unique attributes of zebrafish are being increasingly leveraged to create functional disease models, facilitate drug discovery, and provide critical scientific bases for the development of new clinical tools for the diagnosis and treatment of human disease. In this short review and the accompanying poster, we highlight a few illustrative examples of the applications of the zebrafish model to the study of human health and disease.


Assuntos
Saúde , Pesquisa Médica Translacional , Peixe-Zebra/metabolismo , Animais , Modelos Animais de Doenças , Estudos de Associação Genética , Humanos , Modelos Genéticos
20.
Dis Model Mech ; 7(5): 547-59, 2014 May.
Artigo em Inglês | MEDLINE | ID: mdl-24626987

RESUMO

Usher syndrome (USH), the leading cause of hereditary combined hearing and vision loss, is characterized by sensorineural deafness and progressive retinal degeneration. Mutations in several different genes produce USH, but the proximal cause of sensory cell death remains mysterious. We adapted a proximity ligation assay to analyze associations among three of the USH proteins, Cdh23, Harmonin and Myo7aa, and the microtubule-based transporter Ift88 in zebrafish inner ear mechanosensory hair cells. We found that the proteins are in close enough proximity to form complexes and that these complexes preassemble at the endoplasmic reticulum (ER). Defects in any one of the three USH proteins disrupt formation and trafficking of the complex and result in diminished levels of the other proteins, generalized trafficking defects and ER stress that triggers apoptosis. ER stress, thus, contributes to sensory hair cell loss and provides a new target to explore for protective therapies for USH.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Retículo Endoplasmático/metabolismo , Homeostase , Complexos Multiproteicos/metabolismo , Proteínas de Peixe-Zebra/metabolismo , Peixe-Zebra/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/genética , Animais , Estresse do Retículo Endoplasmático , Células Ciliadas Auditivas Internas , Mecanorreceptores/metabolismo , Mutação/genética , Ligação Proteica , Transporte Proteico , Frações Subcelulares/metabolismo , Vesículas Transportadoras/metabolismo , Proteínas de Peixe-Zebra/genética
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