RESUMO
Parkinson disease (PD) is a neurodegenerative disease characterized by tremor, bradykinesia, rigidity and postural instability. Post-mortem examination shows loss of neurons and Lewy bodies, which are cytoplasmic eosinophilic inclusions, in the substantia nigra and other brain regions. A few families have PD caused by mutations (A53T or A30P) in the gene SNCA (encoding alpha-synuclein). Alpha-synuclein is present in Lewy bodies of patients with sporadic PD, suggesting that alpha-synuclein may be involved in the pathogenesis of PD. It is unknown how alpha-synuclein contributes to the cellular and biochemical mechanisms of PD, and its normal functions and biochemical properties are poorly understood. To determine the protein-interaction partners of alpha-synuclein, we performed a yeast two-hybrid screen. We identified a novel interacting protein, which we term synphilin-1 (encoded by the gene SNCAIP). We found that alpha-synuclein interacts in vivo with synphilin-1 in neurons. Co-transfection of both proteins (but not control proteins) in HEK 293 cells yields cytoplasmic eosinophilic inclusions.
Assuntos
Proteínas de Transporte/metabolismo , Corpos de Inclusão/metabolismo , Proteínas do Tecido Nervoso/metabolismo , Sequência de Aminoácidos , Animais , Química Encefálica , Proteínas de Transporte/genética , Linhagem Celular , Cromossomos Humanos Par 5/genética , Feminino , Humanos , Corpos de Lewy/metabolismo , Masculino , Dados de Sequência Molecular , Proteínas do Tecido Nervoso/genética , Doença de Parkinson/genética , Doença de Parkinson/metabolismo , Plasmídeos/genética , Ligação Proteica , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Ratos , Saccharomyces cerevisiae/genética , Homologia de Sequência de Aminoácidos , Sinucleínas , Distribuição Tecidual , Extratos de Tecidos/metabolismo , Transfecção , alfa-SinucleínaAssuntos
Repetições Minissatélites , Ataxias Espinocerebelares/genética , Repetições de Trinucleotídeos , Regiões 5' não Traduzidas , Adulto , Alelos , Sequência de Bases , Primers do DNA/genética , Feminino , Genes Dominantes , Ligação Genética , Humanos , Masculino , Pessoa de Meia-Idade , LinhagemRESUMO
Atrophin-1 contains a polyglutamine repeat, expansion of which is responsible for dentatorubral and pallidoluysian atrophy (DRPLA). The normal function of atrophin-1 is unknown. We have identified five atrophin-1 interacting proteins (AIPs) which bind to atrophin-1 in the vicinity of the polyglutamine tract using the yeast two-hybrid system. Four of the interactions were confirmed using in vitro binding assays. All five interactors contained multiple WW domains. Two are novel. The AIPs can be divided into two distinct classes. AIP1 and AIP3/WWP3 are MAGUK-like multidomain proteins containing a number of protein-protein interaction modules, namely a guanylate kinase-like region, two WW domains, and multiple PDZ domains. AIP2/WWP2, AIP4, and AIP5/WWP1 are highly homologous, each having four WW domains and a HECT domain characteristic of ubiquitin ligases. These interactors are similar to recently isolated huntingtin-interacting proteins, suggesting possible commonality of function between two proteins responsible for very similar diseases.
Assuntos
Química Encefálica/genética , Proteínas do Tecido Nervoso/química , Proteínas do Tecido Nervoso/genética , Sequências Repetitivas de Ácido Nucleico , Animais , Anticorpos , Clonagem Molecular , DNA Complementar , Feto/química , Humanos , Dados de Sequência Molecular , Proteínas do Tecido Nervoso/imunologia , Ligação Proteica/genética , Estrutura Terciária de Proteína , Coelhos , Homologia de Sequência de Aminoácidos , Leveduras/genéticaRESUMO
Expansion mutations of trinucleotide repeats and other units of unstable DNA have been proposed to account for at least some of the genetic susceptibility to a number of neuropsychiatric disorders, including bipolar affective disorder, schizophrenia, autism, and panic disorder. To generate additional candidate genes for these and other disorders, cDNA libraries from human brain were probed at high stringency for clones containing CCG, CGC, GCC, CGG, GCG, and GGC repeats (referred to collectively as CCG repeats). Some 18 cDNAs containing previously unpublished or uncharacterized repeats were characterized for chromosomal locus, repeat length polymorphism, and similarity to genes of known function. The cDNAs were also compared with the 37 human genes with eight or more consecutive CCG triplets in GenBank. The repeats were mapped to a number of loci, including 1p34, 2p11.2, 2q30-32, 3p21, 3p22, 4q35, 6q22, 7qter, 13p13, 17q24, 18p11, 19p13.3, 20q12, 20q13.3, and 22q12. Length polymorphism was detected in 50% of the repeats. The newly cloned cDNAs include a complete transcript of human neurexin-1B, a portion of BCNG-1 (a newly described brain-specific ion channel), a previously unreported polymorphic repeat located in the 5' UTR region of the guanine nucleotide-binding protein (G-protein) beta2 subunit, and a human version of the mouse proline-rich protein 7. This list of cDNAs should expedite the search for expansion mutations associated with diseases of the central nervous system.
Assuntos
Química Encefálica/genética , DNA Complementar/genética , Repetições de Trinucleotídeos , Mapeamento Cromossômico , Cromossomos Humanos , Canais de Cátion Regulados por Nucleotídeos Cíclicos , Bases de Dados Factuais , Heterozigoto , Humanos , Canais Disparados por Nucleotídeos Cíclicos Ativados por Hiperpolarização , Canais Iônicos/genética , Transtornos Mentais/etiologia , Transtornos Mentais/genética , Dados de Sequência Molecular , Proteínas do Tecido Nervoso/genética , Polimorfismo de Fragmento de Restrição , Canais de PotássioRESUMO
We have recently identified a protein we called synphilin-1, which interacts in vivo with alpha-synuclein. Mutations in alpha-synuclein cause familial Parkinson's disease (PD). Alpha-synuclein protein is present in the pathologic lesions of familial and sporadic PD, and diffuse Lewy body disease, indicating an important pathogenic role for alpha-synuclein. Here we describe the structure of the human synphilin-1 gene (SNCAIP). The open reading frame of this gene is contained within ten exons. We have designed primers to amplify each SNCAIP exon, so these primers can now be used to screen for mutations or polymorphisms in patients with Parkinson's disease or related diseases. We found a highly polymorphic GT repeat within intron 5 of SNCAIP, suitable for linkage analysis of families with PD. We have mapped SNCAIP locus to Chromosome (Chr) 5q23.1-23.3 near markers WI-4673 and AFMB352XH5. In addition, using immunohistochemistry in human postmortem brain tissue, we found that synphilin-1 protein is present in neuropil, similar to alpha-synuclein protein. Because of its association with alpha-synuclein, synphilin-1 may be a candidate for involvement in Parkinson's disease or other related disorders.