RESUMEN
Bipolar disorder (BD) is a prevalent mood disorder that tends to cluster in families. Despite high heritability estimates, few genetic susceptibility factors have been identified over decades of genetic research. One possible interpretation for the shortcomings of previous studies to detect causative genes is that BD is caused by highly penetrant rare variants in many genes. We explored this hypothesis by sequencing the exomes of affected individuals from 40 well-characterized multiplex families. We identified rare variants segregating with affected status in many interesting genes, and found an enrichment of deleterious variants in G protein-coupled receptor (GPCR) family genes, which are important drug targets. Furthermore, we showed targeted downstream GPCR dysregulation for some of the variants that may contribute to disease pathology. Particularly interesting was the finding of a rare and functionally relevant nonsense mutation in the corticotropin-releasing hormone receptor 2 (CRHR2) gene that tracked with affected status in one family. By focusing on rare variants in informative families, we identified key biochemical pathways likely implicated in this complex disorder.
Asunto(s)
Trastorno Bipolar/genética , Receptores Acoplados a Proteínas G/genética , Receptores Acoplados a Proteínas G/metabolismo , Adulto , Trastorno Bipolar/metabolismo , Estudios de Casos y Controles , Familia , Femenino , Frecuencia de los Genes/genética , Ligamiento Genético , Predisposición Genética a la Enfermedad/genética , Variación Genética/genética , Estudio de Asociación del Genoma Completo , Secuenciación de Nucleótidos de Alto Rendimiento/métodos , Humanos , Masculino , Persona de Mediana Edad , Linaje , Receptores de Hormona Liberadora de Corticotropina/genética , Secuenciación del ExomaRESUMEN
Expansion of polyalanine tracts causes at least nine inherited human diseases. Among these, a polyalanine tract expansion in the poly (A)-binding protein nuclear 1 (expPABPN1) causes oculopharyngeal muscular dystrophy (OPMD). So far, there is no treatment for OPMD patients. Developing drugs that efficiently sustain muscle protection by activating key cell survival mechanisms is a major challenge in OPMD research. Proteins that belong to the Wnt family are known for their role in both human development and adult tissue homeostasis. A hallmark of the Wnt signaling pathway is the increased expression of its central effector, beta-catenin (ß-catenin) by inhibiting one of its upstream effector, glycogen synthase kinase (GSK)3ß. Here, we explored a pharmacological manipulation of a Wnt signaling pathway using lithium chloride (LiCl), a GSK-3ß inhibitor, and observed the enhanced expression of ß-catenin protein as well as the decreased cell death normally observed in an OPMD cell model of murine myoblast (C2C12) expressing the expanded and pathogenic form of the expPABPN1. Furthermore, this effect was also observed in primary cultures of mouse myoblasts expressing expPABPN1. A similar effect on ß-catenin was also observed when lymphoblastoid cells lines (LCLs) derived from OPMD patients were treated with LiCl. We believe manipulation of the Wnt/ß-catenin signaling pathway may represent an effective route for the development of future therapy for patients with OPMD.
Asunto(s)
Cloruro de Litio/farmacología , Cloruro de Litio/uso terapéutico , Distrofia Muscular Oculofaríngea/tratamiento farmacológico , Distrofia Muscular Oculofaríngea/patología , Vía de Señalización Wnt/efectos de los fármacos , Animales , Muerte Celular/efectos de los fármacos , Diferenciación Celular/efectos de los fármacos , Línea Celular , Núcleo Celular/efectos de los fármacos , Núcleo Celular/metabolismo , Proliferación Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Relación Dosis-Respuesta a Droga , Proteínas Fluorescentes Verdes/metabolismo , Humanos , Linfocitos/efectos de los fármacos , Linfocitos/metabolismo , Ratones , Distrofia Muscular Oculofaríngea/metabolismo , Proteínas Mutantes/metabolismo , Mioblastos/efectos de los fármacos , Mioblastos/metabolismo , Mioblastos/patología , Proteína I de Unión a Poli(A)/genética , Proteína I de Unión a Poli(A)/metabolismo , Transporte de Proteínas/efectos de los fármacos , Expansión de Repetición de Trinucleótido/genética , beta Catenina/metabolismoRESUMEN
Complex genetic traits can be dissected in mice, using well-defined sets of recombinant inbred strains, congenic strains, and recombinant congenic strains (RCS). We report the creation of a series of 37 independent RCS derived from the commonly used inbred strains of laboratory mouse A/J (A) and C57BL/6J (B6). These RCS were derived by systematic inbreeding of independent pairs of animals from a (F1 x A) x A and a (F1 x B) x B double backcross (N3), to create AcB and BcA strains, respectively. Fifteen AcB strains and 22 BcA strains at between 18 and 30 generations of inbreeding have been generated, are healthy, and show stable breeding performance. These strains have been genotyped for a total of 625 informative microsatellite DNA markers covering the entire genome, with an average spacing of 2.6 cM. Haplotype analyses indicate that on average, AcB and BcA strains contain 13.25% of the donor genome, a value close to the 12.5% expected from the breeding scheme used in their creation. In the AcB set, approximately 79% of the B6 genome has been transferred in independent strains, while in the BcA set approximately 84% of the A genome is represented on the B6 background. This represents an excellent coverage of congenic segments from both parental genomes in the two sets of strains, which can now be used to map simple and complex traits in a genome-wide fashion. As an example of the power of AcB/BcA strains as a mapping tool, the 37 strains were typed for susceptibility to infection with Legionella pneumophila, a monogenic trait controlled by the Lgn1 locus on Chromosome 13. Analysis of the strain distribution pattern of L. pneumophila susceptibility allowed direct mapping of Lgn1 to a 3-cM interval. The AcB/BcA set should prove a useful tool with which to investigate the complex genetic basis of known interstrain differences between A and B6 for many important diseases.
Asunto(s)
Ratones Congénicos/genética , Carácter Cuantitativo Heredable , Animales , Cromosomas/genética , Cruzamientos Genéticos , ADN/genética , Femenino , Frecuencia de los Genes , Genoma , Genotipo , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Endogámicos , Repeticiones de Microsatélite , Recombinación GenéticaAsunto(s)
Actitud Frente a la Salud , Servicios Comunitarios de Salud Mental/economía , Servicios Comunitarios de Salud Mental/legislación & jurisprudencia , Seguro Psiquiátrico/legislación & jurisprudencia , Opinión Pública , Humanos , Seguro Psiquiátrico/estadística & datos numéricos , Rhode IslandRESUMEN
Oculopharyngeal muscular dystrophy is caused by expansion of a (GCG)n trinucleotide repeat in the poly(A) binding protein 2 (PABP2) gene. The pathological hallmark of oculopharyngeal muscular dystrophy is the accumulation of intranuclear inclusions in muscle fibers. To test whether the polyalanine expansion of PABP2 directly leads to the formation of the nuclear aggregates, both normal and expanded PABP2 cDNAs were expressed in COS-7 cells. We find that expression of mutated PABP2 protein is sufficient for its accumulation as intranuclear inclusions.
Asunto(s)
Distrofias Musculares/genética , Proteínas de Unión al ARN/genética , Expansión de Repetición de Trinucleótido/genética , Humanos , Inmunohistoquímica , Modelos Neurológicos , Distrofias Musculares/patología , Proteínas de Unión a Poli(A)Asunto(s)
Servicios de Salud Mental , Niño , Servicios de Salud del Niño , Desinstitucionalización/historia , Desinstitucionalización/tendencias , Política de Salud , Historia del Siglo XX , Personas con Mala Vivienda , Humanos , Cobertura del Seguro , Seguro de Salud/historia , Seguro de Salud/tendencias , Programas Controlados de Atención en Salud/historia , Programas Controlados de Atención en Salud/legislación & jurisprudencia , Programas Controlados de Atención en Salud/tendencias , Massachusetts , Medicaid , Servicios de Salud Mental/historia , Servicios de Salud Mental/legislación & jurisprudencia , Servicios de Salud Mental/organización & administración , Servicios de Salud Mental/tendencias , Gobierno Estatal , Estados UnidosRESUMEN
As part of our effort to clone positionally the oculopharyngeal muscular dystrophy (OPMD) gene, we constructed a YAC contig, a cosmid contig, and an EcoRI restriction map of the OPMD candidate region. The YAC contig spans more than 2 Mb and encompasses the loci D14S283 and D14S990 and the cardiac alpha and beta myosin heavy chain genes (MYH6 and MYH7). A 700-kb cosmid contig containing the D14S990 and the myosin genes and a long-range restriction map covering the region between D14S990 and the MYH6 and MYH7 gene cluster were established. A detailed EcoRI restriction map of the cosmid contig was determined, and five putative CpG islands were identified. Based on these data, the four loci were mapped within an approximately 600-kb region with the following centromere to telomere order: D14S283, D14S990, MYH6, and MYH7. The YAC and cosmid contigs will facilitate the identification of genes lying within the OPMD candidate interval.
Asunto(s)
Cromosomas Humanos Par 14/genética , Genes/genética , Distrofias Musculares/genética , Paseo de Cromosoma , Mapeo Contig , Electroforesis en Gel de Campo Pulsado , Marcadores Genéticos , Humanos , Músculos Oculomotores , Músculos Faríngeos , Mapeo RestrictivoRESUMEN
The Canadian health care system has been frequently discussed but firmly rejected as a model for guiding the transformation of American medicine. In mental health care, however, one can find a growing convergence between the two systems. This convergence is largely coming about as a result of Canadian borrowing from American programs and strategies. In this paper we provide examples of planning service, and policy elements in U.S. mental health care that Canadians have drawn upon. We analyze this phenomenon in terms of cross-national policy learning, identifying the factors and forces that facilitate this trend for Canada while impeding it for the United States.
Asunto(s)
Comparación Transcultural , Política de Salud , Servicios de Salud Mental/legislación & jurisprudencia , Canadá , Redes Comunitarias , Humanos , Programas Controlados de Atención en Salud , Servicios de Salud Mental/organización & administración , Modelos Organizacionales , Estados UnidosRESUMEN
Autosomal dominant oculopharyngeal muscular dystrophy (OPMD) is an adult-onset disease with a world-wide distribution. It usually presents in the sixth decade with progressive swallowing difficulties (dysphagia), eyelid drooping (ptosis) and proximal limb weakness. Unique nuclear filament inclusions in skeletal muscle fibres are its pathological hallmark. We isolated the poly(A) binding protein 2 gene (PABP2) from a 217-kb candidate interval on chromosome 14q11 (B.B. et al., manuscript submitted). A (GCG)6 repeat encoding a polyalanine tract located at the N terminus of the protein was expanded to (GCG)8-13 in the 144 OPMD families screened. More severe phenotypes were observed in compound heterozygotes for the (GCG)9 mutation and a (GCG)7 allele that is found in 2% of the population, whereas homozygosity for the (GCG)7 allele leads to autosomal recessive OPMD. Thus the (GCG)7 allele is an example of a polymorphism which can act either as a modifier of a dominant phenotype or as a recessive mutation. Pathological expansions of the polyalanine tract may cause mutated PABP2 oligomers to accumulate as filament inclusions in nuclei.
Asunto(s)
Cromosomas Humanos Par 14 , Distrofias Musculares/genética , Proteínas de Unión al ARN/genética , Repeticiones de Trinucleótidos , Adulto , Anciano , Secuencia de Bases , Canadá , Mapeo Cromosómico , Clonación Molecular , Femenino , Francia/etnología , Genes Dominantes , Humanos , Masculino , Persona de Mediana Edad , Datos de Secuencia Molecular , Linaje , Proteínas de Unión a Poli(A) , Población BlancaAsunto(s)
Anécdotas como Asunto , Programas Controlados de Atención en Salud/legislación & jurisprudencia , Opinión Pública , Actitud Frente a la Salud , Accesibilidad a los Servicios de Salud/legislación & jurisprudencia , Programas Controlados de Atención en Salud/organización & administración , Defensa del Paciente/legislación & jurisprudencia , Política , Estados UnidosRESUMEN
Several inherited diseases have been mapped to the distal tip of human chromosome 21. In our recent efforts to clone candidate genes for some of these disorders, we have assembled a cosmid and BAC contig spanning 770 kb. We have identified expressed sequences from this contig by means of a cDNA hybrid selection scheme. We present here the isolation, cDNA sequence, genomic organization, and polymorphisms analysis of one such expressed sequence, GT334, which had been identified independently and designated EHOC-1. GT334 is split into 23 exons, and spans an estimated 95 kb of genomic DNA. A pseudogene of the histone H2AZ gene has been identified, and maps within the third intron. We have identified an ORF potentially encoding a protein 1259 amino acids in length, longer than that described in the EHOC-1 gene. The GT334 gene was screened for single base pair changes using single-strand conformation polymorphism (SSCP) analysis and we have identified seven sequence variations within this gene. These polymorphisms can be used as markers in the genetic mapping of other diseases localized to this region.
Asunto(s)
Cromosomas Humanos Par 21 , Genes , Proteínas de la Membrana/genética , Secuencia de Aminoácidos , Secuencia de Bases , Cósmidos , ADN Complementario/genética , Exones , Expresión Génica , Humanos , Intrones , Datos de Secuencia Molecular , Polimorfismo Genético , Polimorfismo Conformacional Retorcido-Simple , Mapeo Restrictivo , Proteínas de Transporte VesicularRESUMEN
Progressive myoclonus epilepsy type 1 (EPM1, also known as Unverricht-Lundborg disease) is an autosomal recessive disorder characterized by progressively worsening myoclonic jerks, frequent generalized tonic-clonic seizures, and a slowly progressive decline in cognition. Recently, two mutations in the cystatin B gene (also known as stefin B, STFB) mapping to 21q22.3 have been implicated in the EPM1 phenotype: a G-->C substitution in the last nucleotide of intron 1 that was predicted to cause a splicing defect in one family, and a C-->T substitution that would change an Arg codon (CGA) to a stop codon (TGA) at amino acid position 68, resulting in a truncated cystatin B protein in two other families. A fourth family showed undetectable amounts of STFB mRNA by northern blot analysis in an affected individual. We present haplotype and mutational analyses of our collection of 20 unrelated EPM1 patients and families from different ethnic groups. We identify four different mutations, the most common of which consists of an unstable approximately 600-900 bp insertion which is resistant to PCR amplification. This insertion maps to a 12-bp polymorphic tandem repeat located in the 5' flanking region of the STFB gene, in the region of the promoter. The size of the insertion varies between different EPM1 chromosomes sharing a common haplotype and a common origin, suggesting some level of meiotic instability over the course of many generations. This dynamic mutation, which appears distinct from conventional trinucleotide repeat expansions, may arise via a novel mechanism related to the instability of tandemly repeated sequences.
Asunto(s)
Cistatinas/genética , Elementos Transponibles de ADN , Epilepsias Mioclónicas/genética , Mutación , Secuencia de Bases , Cromosomas Humanos Par 21 , Cistatina B , Inhibidores de Cisteína Proteinasa/genética , Cartilla de ADN , Femenino , Haplotipos , Humanos , Masculino , Datos de Secuencia Molecular , Linaje , Reacción en Cadena de la Polimerasa , Polimorfismo Genético , Secuencias Reguladoras de Ácidos Nucleicos , Secuencias Repetitivas de Ácidos NucleicosRESUMEN
As part of efforts to identify candidate genes for disorders mapped to 21q22.3, we have constructed a 405-kb cosmid contig encompassing five tightly linked markers mapping to this region. A subset of these cosmids was used to identify cDNA fragments by the method of hybrid selection. We present here the cDNA sequence of one such gene (GT335) mapping to this region. The gene is expressed as a 1.7-kb transcript predominantly in heart and skeletal muscle, potentially displays alternate splicing, and is predicted to encode a protein 268 amino acids in length. GT335 spans an estimated 13 kb of genomic DNA and is split into seven exons. Five of the six introns conform to the GT . . . AG consensus for intronic splice junctions; the sixth contains nonconventional (AT . . . AC) intronic junctions. We screened this gene for single-basepair mutations using single-strand conformation polymorphism and sequence analysis of both cDNA and genomic DNA from a number of unrelated individuals and have identified several sequence variations, two of which cause conservative amino acid substitutions. This gene is well conserved evolutionarily, with homologs identified in zebrafish and Escherichia coli, suggesting that it plays an important role in basic cellular metabolism.
Asunto(s)
Cromosomas Humanos Par 21/genética , Escherichia coli/genética , Genes , Proteínas de Homeodominio , Proteínas Musculares/genética , Proteínas , Secuencia de Aminoácidos , Animales , Secuencia de Bases , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico , Southern Blotting , Mapeo Cromosómico , Secuencia de Consenso , Cósmidos/genética , ADN Complementario/genética , Expresión Génica , Humanos , Proteínas Mitocondriales , Datos de Secuencia Molecular , Reacción en Cadena de la Polimerasa , Polimorfismo Conformacional Retorcido-Simple , Empalme del ARN , Alineación de Secuencia , Homología de Secuencia de Aminoácido , Factor de Transcripción HES-1 , Pez Cebra/genéticaRESUMEN
As part of efforts to identify candidate genes for disease mapping to the 21q22.3 region, we have assembled a 770-kb cosmid and BAC contig containing eight tightly linked markers. These cosmids and BACs were restriction mapped using eight rare cutting enzymes, with the goal of identifying CpG-rich islands. One such island was identified by the clustering of NotI, EagI, SstII, and BssHII sites, and corresponded to the NotI linking clone LJ104 described previously. A 7.6-kb HindIII fragment containing this CpG-rich island was subcloned and partially sequenced. A homology search using the sequence obtained from either side of the NotI site identified an expressed sequence tag with homology to the yeast periodic tryptophan protein 2 (PWP2). Several cDNAs corresponding to the human PWP2 gene were identified and partially sequenced. Northern blot analysis revealed a 3.3-kb transcript that was well expressed in all tissues tested. A cDNA consensus of 3157 bp was obtained, and an open reading frame potentially encoding 919 amino acid residues was identified. The predicted protein shows 42% identity and 57% similarity at the amino acid level to the yeast PWP2 protein, which is a member of the WD-repeat containing superfamily, and potentially encodes a G-protein beta subunit. The PWP2 gene is split into 21 exons, ranging in size from 53 to 516 bp, and spans an estimated 25 kb. The gene is transcribed in a 21cen-->21qter direction, with its 5' end mapping approximately 195 kb proximal to the 5' end of the phosphofructokinase-liver isoform gene. Four single base-pair polymorphisms were identified using single-stranded conformation polymorphism analysis. Possible functions of the protein based on homology to other members of the WD-repeat-containing family are discussed.
Asunto(s)
Cromosomas Humanos Par 21 , Proteínas/genética , Proteínas de Saccharomyces cerevisiae , Secuencia de Aminoácidos , Secuencia de Bases , Mapeo Cromosómico , ADN Complementario , Expresión Génica , Genoma , Humanos , Datos de Secuencia Molecular , Polimorfismo Genético , Polimorfismo Conformacional Retorcido-Simple , ARN Mensajero , Proteínas Ribosómicas , Análisis de Secuencia de ADN , Homología de Secuencia de Aminoácido , LevadurasAsunto(s)
Política de Salud/legislación & jurisprudencia , Trastornos Mentales/terapia , Servicios de Salud Mental/legislación & jurisprudencia , Formulación de Políticas , Reforma de la Atención de Salud , Sistemas Prepagos de Salud , Política de Salud/tendencias , Humanos , Defensa del Paciente , Bienestar Social , Estados UnidosRESUMEN
Pretreatment of LPS-induced RAW 264.7 cells with three PKC inhibitors suggests that induction of TNF-alpha, nitric oxide (NO), gelatinase B (Gel B) and IL-6 involves at least three distinct signalling pathways. We confirmed the PKC dependence of TNF-alpha and NO productions and found that Gel B was inhibited by Calphostin C (CAL), but potentiated by staurosporine (STAR) and CGP 41 251. IL-6 production was stimulated by the three inhibitors. Our results indicate that up-regulation of Gel B, TNF-alpha and NO seems to involve PKC at different levels, whereas up-regulation of IL-6 production appears to be PKC-independent. However, IL-6 production in RAW 264.7 cells seems to be down-regulated by a PKC-dependent feedback mechanism.
Asunto(s)
Alcaloides/farmacología , Colagenasas/biosíntesis , Expresión Génica/efectos de los fármacos , Interleucina-6/biosíntesis , Macrófagos/inmunología , Naftalenos , Compuestos Policíclicos/farmacología , Proteína Quinasa C/antagonistas & inhibidores , Animales , Bioensayo , Northern Blotting , Línea Celular Transformada , Colagenasas/análisis , Interleucina-6/análisis , Macrófagos/efectos de los fármacos , Macrófagos/metabolismo , Metaloproteinasa 9 de la Matriz , Ratones , Óxido Nítrico/biosíntesis , ARN Mensajero/análisis , ARN Mensajero/biosíntesis , Estaurosporina , Factor de Necrosis Tumoral alfa/biosíntesisRESUMEN
Frankia alni CpI1 has two glutamine synthetases (GSs), GSI and GSII. The GSI gene (glnA) was isolated from a cosmid library of F. alni CpI1 DNA by heterologous probing with glnA from Streptomyces coelicolor. The glnA gene was shown to be located upstream of the GSII gene (glnII) by DNA-DNA hybridization. The nucleotide sequences of the 1,422-bp CpI1 glnA gene and of the 449-bp intervening region between glnA and glnII were determined, and the glnA amino acid sequence was deduced. In common with GSIs from other organisms, CpI1 GSI contains five conserved regions near the active site and a conserved tyrosine at the adenylylation site. F. alni CpI1 glnA complemented the glutamine growth requirement of the Escherichia coli glnA deletion strain YMC11 but only when expressed from an E. coli lac promoter. While the functional significance of maintaining two GSs adjacent to one another remains unclear, this arrangement in F. alni provides support for the recently proposed origin of GSI and GSII as resulting from a gene duplication early in the evolution of life.
Asunto(s)
Actinomycetales/genética , Genes Bacterianos/genética , Ligamiento Genético , Glutamato-Amoníaco Ligasa/genética , Isoenzimas/genética , Actinomycetales/enzimología , Secuencia de Aminoácidos , Secuencia de Bases , Clonación Molecular , Estabilidad de Enzimas , Escherichia coli/genética , Eliminación de Gen , Calor , Datos de Secuencia Molecular , Familia de Multigenes/genética , Proteínas Recombinantes/biosíntesis , Secuencias Reguladoras de Ácidos Nucleicos/genética , Análisis de Secuencia de ADN , Homología de Secuencia de Aminoácido , Streptomyces/enzimología , Streptomyces/genéticaRESUMEN
We performed molecular phylogenetic analyses of glutamine synthetase (GS) genes in order to investigate their evolutionary history. The analyses were done on 30 DNA sequences of the GS gene which included both prokaryotes and eukaryotes. Two types of GS genes are known at present: the GSI gene found so far only in prokaryotes and the GSII gene found in both prokaryotes and eukaryotes. Our study has shown that the two types of GS gene were produced by a gene duplication which preceded, perhaps by > 1000 million years, the divergence of eukaryotes and prokaryotes. The results are consistent with the facts that (i) GS is a key enzyme of nitrogen metabolism found in all extant life forms and (ii) the oldest biological fossils date back 3800 million years. Thus, we suggest that GS genes are one of the oldest existing and functioning genes in the history of gene evolution and that GSI genes should also exist in eukaryotes. Furthermore, our study may stimulate investigation on the evolution of "preprokaryotes," by which we mean the organisms that existed during the era between the origin of life and the divergence of prokaryotes and eukaryotes.