RESUMO
Hereditary neuralgic amyotrophy (HNA) is an autosomal dominant recurrent neuropathy affecting the brachial plexus. HNA is triggered by environmental factors such as infection or parturition. We report three mutations in the gene septin 9 (SEPT9) in six families with HNA linked to chromosome 17q25. HNA is the first monogenetic disease caused by mutations in a gene of the septin family. Septins are implicated in formation of the cytoskeleton, cell division and tumorigenesis.
Assuntos
Neurite do Plexo Braquial/genética , Cromossomos Humanos Par 17/genética , GTP Fosfo-Hidrolases/genética , Mutação , Sequência de Aminoácidos , Animais , Sequência de Bases , Cães , Humanos , Camundongos , Dados de Sequência Molecular , Ratos , SeptinasRESUMO
Different mutations in the human Crumbs homolog-1 (CRB1) gene cause a variety of retinal dystrophies, such as Leber congenital amaurosis, early onset retinitis pigmentosa (e.g., RP12), RP with Coats-like exudative vasculopathy, and pigmented paravenous retinochoroidal atrophy. Loss of Crb1 leads to displaced photoreceptors and focal degeneration of all neural layers attributable to loss of adhesion between photoreceptors and Müller glia cells. To gain insight into genotype-phenotype relationship, we generated Crb1(C249W) mice that harbor an amino acid substitution (Cys249Trp) in the extracellular sixth calcium-binding epidermal growth factor domain of Crb1. Our analysis showed that Crb1(C249W) as wild-type protein trafficked to the subapical region adjacent to adherens junctions at the outer limiting membrane (OLM). Hence, these data suggest correct trafficking of the corresponding mutant CRB1 in RP12 patients. Crb1(C249W) mice showed loss of photoreceptors in the retina, relatively late compared with mice lacking Crb1. Scanning laser ophthalmoscopy revealed autofluorescent dots that presumably represent layer abnormalities after OLM disturbance. Gene expression analyses revealed lower levels of pituitary tumor transforming gene 1 (Pttg1) transcripts in Crb1(C249W/-) knock-in and Crb1(-/-) knock-out compared with control retinas. Exposure to white light decreased levels of Pttg1 in Crb1 mutant retinas. We hypothesize deregulation of Pttg1 expression attributable to a C249W substitution in the extracellular domain of Crb1.
Assuntos
Substituição de Aminoácidos/genética , Proteínas do Olho/genética , Regulação da Expressão Gênica/fisiologia , Proteínas de Membrana/genética , Proteínas de Neoplasias/antagonistas & inibidores , Proteínas de Neoplasias/genética , Proteínas do Tecido Nervoso/genética , Degeneração Retiniana/genética , Sequência de Aminoácidos , Animais , Cisteína/genética , Proteínas do Olho/fisiologia , Humanos , Proteínas de Membrana/deficiência , Proteínas de Membrana/fisiologia , Camundongos , Camundongos Knockout , Dados de Sequência Molecular , Proteínas de Neoplasias/biossíntese , Proteínas do Tecido Nervoso/deficiência , Proteínas do Tecido Nervoso/fisiologia , Estrutura Terciária de Proteína/genética , Degeneração Retiniana/metabolismo , Degeneração Retiniana/patologia , Retinose Pigmentar/genética , Retinose Pigmentar/metabolismo , Retinose Pigmentar/patologia , Securina , Triptofano/genéticaRESUMO
Mutations in the human Crumbs homologue-1 (CRB1) gene cause retinal diseases including Leber's congenital amaurosis (LCA) and retinitis pigmentosa type 12. The CRB1 transmembrane protein localizes at a subapical region (SAR) above intercellular adherens junctions between photoreceptor and Müller glia (MG) cells. We demonstrate that the Crb1-/- phenotype, as shown in Crb1-/- mice, is accelerated and intensified in primary retina cultures. Immuno-electron microscopy showed strong Crb1 immunoreactivity at the SAR in MG cells but barely in photoreceptor cells, whereas Crb2, Crb3, Patj, Pals1 and Mupp1 were present in both cell types. Human CRB1, introduced in MG cells in Crb1-/- primary retinas, was targeted to the SAR. RNA interference-induced silencing of the Crb1-interacting-protein Pals1 (protein associated with Lin7; Mpp5) in MG cells resulted in loss of Crb1, Crb2, Mupp1 and Veli3 protein localization and partial loss of Crb3. We conclude that Pals1 is required for correct localization of Crb family members and its interactors at the SAR of polarized MG cells.
Assuntos
Proteínas de Membrana/metabolismo , Proteínas do Tecido Nervoso/metabolismo , Neuroglia/metabolismo , Núcleosídeo-Fosfato Quinase/metabolismo , Animais , Sequência de Bases , Polaridade Celular , DNA/genética , Proteínas do Olho/genética , Proteínas do Olho/metabolismo , Humanos , Proteínas de Membrana/deficiência , Proteínas de Membrana/genética , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Microscopia Imunoeletrônica , Mutação , Proteínas do Tecido Nervoso/deficiência , Proteínas do Tecido Nervoso/genética , Neuroglia/ultraestrutura , Núcleosídeo-Fosfato Quinase/genética , Técnicas de Cultura de Órgãos , Fenótipo , Células Fotorreceptoras de Vertebrados/metabolismo , Células Fotorreceptoras de Vertebrados/ultraestrutura , Interferência de RNA , Retina/crescimento & desenvolvimento , Retina/metabolismoRESUMO
Hereditary neuralgic amyotrophy (HNA) is an autosomal-dominant inherited recurrent focal neuropathy affecting mainly the brachial plexus. In this study we report the genomic structure and mutation analysis of three candidate genes: sphingosine kinase 1 (SPHK1); tissue inhibitor of metalloproteinase 2 (TIMP2); and cytoglobin (CYGB). We did not find any disease-associated mutations, indicating that HNA is not caused by point mutations in these genes. However, we identified several sequencing errors in the cDNA of SPHK1 as well as seven novel single-nucleotide polymorphisms.
Assuntos
Neurite do Plexo Braquial/genética , Análise Mutacional de DNA , Globinas/genética , Fosfotransferases (Aceptor do Grupo Álcool)/genética , Inibidor Tecidual de Metaloproteinase-2/genética , Substituição de Aminoácidos , Citoglobina , DNA Complementar/genética , Éxons/genética , Testes Genéticos , Biblioteca Genômica , Humanos , Dados de Sequência Molecular , Mutação Puntual/genética , Polimorfismo de Nucleotídeo Único/genéticaRESUMO
Loss of Crumbs homologue 1 (CRB1) function causes either the eye disease Leber congenital amaurosis or progressive retinitis pigmentosa, depending on the amount of residual CRB1 activity and the genetic background. Crb1 localizes specifically to the sub-apical region adjacent to the adherens junction complex at the outer limiting membrane in the retina. We show that it is associated here with multiple PDZ protein 1 (Mupp1), protein associated with Lin-7 (Pals1 or Mpp5) and Mpp4. We have produced Crb1(-/-) mice completely lacking any functional Crb1. Although the retinas are initially normal, by 3-9 months the Crb1(-/-) retinas develop localized lesions where the integrity of the outer limiting membrane is lost and giant half rosettes are formed. After delamination of the photoreceptor layer, neuronal cell death occurs in the inner and outer nuclear layers of the retina. On moderate exposure to light for 3 days at 3 months of age, the number of severe focal retinal lesions significantly increases in the Crb1(-/-) retina. Crb2, Crb3 and Crb1 interacting proteins remain localized to the sub-apical region and therefore are not sufficient to maintain cell adhesion during light exposure in Crb1(-/-) retinas. Thus we propose that during light exposure Crb1 is essential to maintain, but not assemble, adherens junctions between photoreceptors and Müller glia cells and prevents retinal disorganization and dystrophy. Hence, light may be an influential factor in the development of the corresponding human diseases.