Your browser doesn't support javascript.
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 25
Filtrar
Filtros adicionais











Intervalo de ano
1.
Sci Rep ; 9(1): 3574, 2019 Mar 05.
Artigo em Inglês | MEDLINE | ID: mdl-30837603

RESUMO

Myeloid-derived suppressor cells (MDSCs) are key players in immune evasion, tumor progression and metastasis. MDSCs accumulate under various pathological states and fall into two functionally and phenotypically distinct subsets that have been identified in humans and mice: polymorphonuclear (PMN)-MDSCs and monocytic (M)-MDSCs. As dogs are an excellent model for human tumor development and progression, we set out to identify PMN-MDSCs and M-MDSCs in clinical canine oncology patients. Canine hypodense MHC class II-CD5-CD21-CD11b+ cells can be subdivided into polymorphonuclear (CADO48A+CD14-) and monocytic (CADO48A-CD14+) MDSC subsets. The transcriptomic signatures of PMN-MDSCs and M-MDSCs are distinct, and moreover reveal a statistically significant similarity between canine and previously published human PMN-MDSC gene expression patterns. As in humans, peripheral blood frequencies of canine PMN-MDSCs and M-MDSCs are significantly higher in dogs with cancer compared to healthy control dogs (PMN-MDSCs: p < 0.001; M-MDSCs: p < 0.01). By leveraging the power of evolution, we also identified additional conserved genes in PMN-MDSCs of multiple species that may play a role in MDSC function. Our findings therefore validate the dog as a model for studying MDSCs in the context of cancer.

2.
Blood ; 132(12): 1225-1240, 2018 Sep 20.
Artigo em Inglês | MEDLINE | ID: mdl-29930011

RESUMO

SF3B1, SRSF2, and U2AF1 are the most frequently mutated splicing factor genes in the myelodysplastic syndromes (MDS). We have performed a comprehensive and systematic analysis to determine the effect of these commonly mutated splicing factors on pre-mRNA splicing in the bone marrow stem/progenitor cells and in the erythroid and myeloid precursors in splicing factor mutant MDS. Using RNA-seq, we determined the aberrantly spliced genes and dysregulated pathways in CD34+ cells of 84 patients with MDS. Splicing factor mutations result in different alterations in splicing and largely affect different genes, but these converge in common dysregulated pathways and cellular processes, focused on RNA splicing, protein synthesis, and mitochondrial dysfunction, suggesting common mechanisms of action in MDS. Many of these dysregulated pathways and cellular processes can be linked to the known disease pathophysiology associated with splicing factor mutations in MDS, whereas several others have not been previously associated with MDS, such as sirtuin signaling. We identified aberrantly spliced events associated with clinical variables, and isoforms that independently predict survival in MDS and implicate dysregulation of focal adhesion and extracellular exosomes as drivers of poor survival. Aberrantly spliced genes and dysregulated pathways were identified in the MDS-affected lineages in splicing factor mutant MDS. Functional studies demonstrated that knockdown of the mitosis regulators SEPT2 and AKAP8, aberrantly spliced target genes of SF3B1 and SRSF2 mutations, respectively, led to impaired erythroid cell growth and differentiation. This study illuminates the effect of the common spliceosome mutations on the MDS phenotype and provides novel insights into disease pathophysiology.

3.
Sci Rep ; 8(1): 2151, 2018 02 01.
Artigo em Inglês | MEDLINE | ID: mdl-29391536

RESUMO

The design and implementation of single-cell experiments is often limited by their requirement for fresh starting material. We have adapted a method for histological tissue fixation using dithio-bis(succinimidyl propionate) (DSP), or Lomant's Reagent, to stabilise cell samples for single-cell transcriptomic applications. DSP is a reversible cross-linker of free amine groups that has previously been shown to preserve tissue integrity for histology while maintaining RNA integrity and yield in bulk RNA extractions. Although RNA-seq data from DSP-fixed single cells appears to be prone to characteristic artefacts, such as slightly reduced yield of cDNA and a detectable 3' bias in comparison with fresh cells, cell preservation using DSP does not appear to substantially reduce RNA complexity at the gene level. In addition, there is evidence that instantaneous fixation of cells can reduce inter-cell technical variability. The ability of DSP-fixed cells to retain commonly used dyes, such as propidium iodide, enables the tracking of experimental sub-populations and the recording of cell viability at the point of fixation. Preserving cells using DSP will remove several barriers in the staging of single-cell experiments, including the transport of samples and the scheduling of shared equipment for downstream single-cell isolation and processing.

4.
BMC Genomics ; 18(1): 53, 2017 01 07.
Artigo em Inglês | MEDLINE | ID: mdl-28061811

RESUMO

BACKGROUND: Single-cell RNA-Seq can be a valuable and unbiased tool to dissect cellular heterogeneity, despite the transcriptome's limitations in describing higher functional phenotypes and protein events. Perhaps the most important shortfall with transcriptomic 'snapshots' of cell populations is that they risk being descriptive, only cataloging heterogeneity at one point in time, and without microenvironmental context. Studying the genetic ('nature') and environmental ('nurture') modifiers of heterogeneity, and how cell population dynamics unfold over time in response to these modifiers is key when studying highly plastic cells such as macrophages. RESULTS: We introduce the programmable Polaris™ microfluidic lab-on-chip for single-cell sequencing, which performs live-cell imaging while controlling for the culture microenvironment of each cell. Using gene-edited macrophages we demonstrate how previously unappreciated knockout effects of SAMHD1, such as an altered oxidative stress response, have a large paracrine signaling component. Furthermore, we demonstrate single-cell pathway enrichments for cell cycle arrest and APOBEC3G degradation, both associated with the oxidative stress response and altered proteostasis. Interestingly, SAMHD1 and APOBEC3G are both HIV-1 inhibitors ('restriction factors'), with no known co-regulation. CONCLUSION: As single-cell methods continue to mature, so will the ability to move beyond simple 'snapshots' of cell populations towards studying the determinants of population dynamics. By combining single-cell culture, live-cell imaging, and single-cell sequencing, we have demonstrated the ability to study cell phenotypes and microenvironmental influences. It's these microenvironmental components - ignored by standard single-cell workflows - that likely determine how macrophages, for example, react to inflammation and form treatment resistant HIV reservoirs.


Assuntos
Interação Gene-Ambiente , Macrófagos/citologia , Análise de Sequência de RNA , Análise de Célula Única , Técnicas de Inativação de Genes , Humanos , Macrófagos/metabolismo , Fenótipo , Proteína 1 com Domínio SAM e Domínio HD/deficiência , Proteína 1 com Domínio SAM e Domínio HD/genética
5.
Nat Genet ; 47(7): 717-726, 2015 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-25985138

RESUMO

To assess factors influencing the success of whole-genome sequencing for mainstream clinical diagnosis, we sequenced 217 individuals from 156 independent cases or families across a broad spectrum of disorders in whom previous screening had identified no pathogenic variants. We quantified the number of candidate variants identified using different strategies for variant calling, filtering, annotation and prioritization. We found that jointly calling variants across samples, filtering against both local and external databases, deploying multiple annotation tools and using familial transmission above biological plausibility contributed to accuracy. Overall, we identified disease-causing variants in 21% of cases, with the proportion increasing to 34% (23/68) for mendelian disorders and 57% (8/14) in family trios. We also discovered 32 potentially clinically actionable variants in 18 genes unrelated to the referral disorder, although only 4 were ultimately considered reportable. Our results demonstrate the value of genome sequencing for routine clinical diagnosis but also highlight many outstanding challenges.


Assuntos
Doenças Genéticas Inatas/diagnóstico , Sequenciamento de Nucleotídeos em Larga Escala , Técnicas de Diagnóstico Molecular , Sequência de Bases , Análise Mutacional de DNA , Doenças Genéticas Inatas/genética , Genoma Humano , Humanos , Anotação de Sequência Molecular , Polimorfismo de Nucleotídeo Único , Sensibilidade e Especificidade
6.
Hum Mol Genet ; 23(12): 3200-11, 2014 Jun 15.
Artigo em Inglês | MEDLINE | ID: mdl-24463883

RESUMO

In severe early-onset epilepsy, precise clinical and molecular genetic diagnosis is complex, as many metabolic and electro-physiological processes have been implicated in disease causation. The clinical phenotypes share many features such as complex seizure types and developmental delay. Molecular diagnosis has historically been confined to sequential testing of candidate genes known to be associated with specific sub-phenotypes, but the diagnostic yield of this approach can be low. We conducted whole-genome sequencing (WGS) on six patients with severe early-onset epilepsy who had previously been refractory to molecular diagnosis, and their parents. Four of these patients had a clinical diagnosis of Ohtahara Syndrome (OS) and two patients had severe non-syndromic early-onset epilepsy (NSEOE). In two OS cases, we found de novo non-synonymous mutations in the genes KCNQ2 and SCN2A. In a third OS case, WGS revealed paternal isodisomy for chromosome 9, leading to identification of the causal homozygous missense variant in KCNT1, which produced a substantial increase in potassium channel current. The fourth OS patient had a recessive mutation in PIGQ that led to exon skipping and defective glycophosphatidyl inositol biosynthesis. The two patients with NSEOE had likely pathogenic de novo mutations in CBL and CSNK1G1, respectively. Mutations in these genes were not found among 500 additional individuals with epilepsy. This work reveals two novel genes for OS, KCNT1 and PIGQ. It also uncovers unexpected genetic mechanisms and emphasizes the power of WGS as a clinical tool for making molecular diagnoses, particularly for highly heterogeneous disorders.


Assuntos
Epilepsia/genética , Epilepsia/patologia , Proteínas de Membrana/genética , Proteínas do Tecido Nervoso/genética , Canais de Potássio/genética , Criança , Pré-Escolar , Cromossomos Humanos Par 9 , Epilepsia/diagnóstico , Predisposição Genética para Doença , Estudo de Associação Genômica Ampla , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Canal de Potássio KCNQ2/genética , Masculino , Mutação , Canal de Sódio Disparado por Voltagem NAV1.2/genética , Patologia Molecular , Proteínas Proto-Oncogênicas c-cbl/genética , Dissomia Uniparental , Adulto Jovem
7.
Nat Genet ; 45(3): 304-7, 2013 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-23354436

RESUMO

Craniosynostosis, the premature fusion of the cranial sutures, is a heterogeneous disorder with a prevalence of ∼1 in 2,200 (refs. 1,2). A specific genetic etiology can be identified in ∼21% of cases, including mutations of TWIST1, which encodes a class II basic helix-loop-helix (bHLH) transcription factor, and causes Saethre-Chotzen syndrome, typically associated with coronal synostosis. Using exome sequencing, we identified 38 heterozygous TCF12 mutations in 347 samples from unrelated individuals with craniosynostosis. The mutations predominantly occurred in individuals with coronal synostosis and accounted for 32% and 10% of subjects with bilateral and unilateral pathology, respectively. TCF12 encodes one of three class I E proteins that heterodimerize with class II bHLH proteins such as TWIST1. We show that TCF12 and TWIST1 act synergistically in a transactivation assay and that mice doubly heterozygous for loss-of-function mutations in Tcf12 and Twist1 have severe coronal synostosis. Hence, the dosage of TCF12-TWIST1 heterodimers is critical for normal coronal suture development.


Assuntos
Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Craniossinostoses , Proteínas Nucleares/genética , Proteína 1 Relacionada a Twist/genética , Acrocefalossindactilia/complicações , Acrocefalossindactilia/genética , Acrocefalossindactilia/patologia , Animais , Suturas Cranianas/crescimento & desenvolvimento , Suturas Cranianas/patologia , Craniossinostoses/complicações , Craniossinostoses/genética , Craniossinostoses/patologia , Dimerização , Exoma , Regulação da Expressão Gênica no Desenvolvimento , Heterozigoto , Humanos , Camundongos , Camundongos Transgênicos , Dados de Sequência Molecular , Mutação , Análise de Sequência de DNA , Ativação Transcricional
8.
Eur J Hum Genet ; 21(3): 274-80, 2013 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-22968130

RESUMO

Inherited retinal degeneration (IRD) is a common cause of visual impairment (prevalence ∼1/3500). There is considerable phenotype and genotype heterogeneity, making a specific diagnosis very difficult without molecular testing. We investigated targeted capture combined with next-generation sequencing using Nimblegen 12plex arrays and the Roche 454 sequencing platform to explore its potential for clinical diagnostics in two common types of IRD, retinitis pigmentosa and cone-rod dystrophy. 50 patients (36 unknowns and 14 positive controls) were screened, and pathogenic mutations were identified in 25% of patients in the unknown, with 53% in the early-onset cases. All patients with new mutations detected had an age of onset <21 years and 44% had a family history. Thirty-one percent of mutations detected were novel. A de novo mutation in rhodopsin was identified in one early-onset case without a family history. Bioinformatic pipelines were developed to identify likely pathogenic mutations and stringent criteria were used for assignment of pathogenicity. Analysis of sequencing metrics revealed significant variability in capture efficiency and depth of coverage. We conclude that targeted capture and next-generation sequencing are likely to be very useful in a diagnostic setting, but patients with earlier onset of disease are more likely to benefit from using this strategy. The mutation-detection rate suggests that many patients are likely to have mutations in novel genes.


Assuntos
Mutação , Degeneração Retiniana/diagnóstico , Degeneração Retiniana/genética , Análise de Sequência de DNA/métodos , Idade de Início , Humanos , Degeneração Retiniana/epidemiologia , Retinite Pigmentosa/diagnóstico , Retinite Pigmentosa/genética , Rodopsina/genética
9.
Bioinformatics ; 28(22): 2981-2, 2012 Nov 15.
Artigo em Inglês | MEDLINE | ID: mdl-22962342

RESUMO

SUMMARY: GREVE has been developed to assist with the identification of recurrent genomic aberrations across cancer samples. The exact characterization of such aberrations remains a challenge despite the availability of increasing amount of data, from SNParray to next-generation sequencing. Furthermore, genomic aberrations in cancer are especially difficult to handle because they are, by nature, unique to the patients. However, their recurrence in specific regions of the genome has been shown to reflect their relevance in the development of tumors. GREVE makes use of previously characterized events to identify such regions and focus any further analysis. AVAILABILITY: GREVE is available through a web interface and open-source application (http://www.well.ox.ac.uk/GREVE).


Assuntos
Aberrações Cromossômicas , Genoma Humano , Neoplasias/genética , Software , Pontos de Quebra do Cromossomo , Humanos
10.
PLoS Genet ; 8(2): e1002505, 2012.
Artigo em Inglês | MEDLINE | ID: mdl-22383892

RESUMO

Metabolic Syndrome (MetS) is highly prevalent and has considerable public health impact, but its underlying genetic factors remain elusive. To identify gene networks involved in MetS, we conducted whole-genome expression and genotype profiling on abdominal (ABD) and gluteal (GLU) adipose tissue, and whole blood (WB), from 29 MetS cases and 44 controls. Co-expression network analysis for each tissue independently identified nine, six, and zero MetS-associated modules of coexpressed genes in ABD, GLU, and WB, respectively. Of 8,992 probesets expressed in ABD or GLU, 685 (7.6%) were expressed in ABD and 51 (0.6%) in GLU only. Differential eigengene network analysis of 8,256 shared probesets detected 22 shared modules with high preservation across adipose depots (D(ABD-GLU) = 0.89), seven of which were associated with MetS (FDR P<0.01). The strongest associated module, significantly enriched for immune response-related processes, contained 94/620 (15%) genes with inter-depot differences. In an independent cohort of 145/141 twins with ABD and WB longitudinal expression data, median variability in ABD due to familiality was greater for MetS-associated versus un-associated modules (ABD: 0.48 versus 0.18, P = 0.08; GLU: 0.54 versus 0.20, P = 7.8×10(-4)). Cis-eQTL analysis of probesets associated with MetS (FDR P<0.01) and/or inter-depot differences (FDR P<0.01) provided evidence for 32 eQTLs. Corresponding eSNPs were tested for association with MetS-related phenotypes in two GWAS of >100,000 individuals; rs10282458, affecting expression of RARRES2 (encoding chemerin), was associated with body mass index (BMI) (P = 6.0×10(-4)); and rs2395185, affecting inter-depot differences of HLA-DRB1 expression, was associated with high-density lipoprotein (P = 8.7×10(-4)) and BMI-adjusted waist-to-hip ratio (P = 2.4×10(-4)). Since many genes and their interactions influence complex traits such as MetS, integrated analysis of genotypes and coexpression networks across multiple tissues relevant to clinical traits is an efficient strategy to identify novel associations.


Assuntos
Tecido Adiposo/metabolismo , Perfilação da Expressão Gênica , Redes Reguladoras de Genes , Síndrome Metabólica/genética , Índice de Massa Corporal , Quimiocinas/genética , Feminino , Loci Gênicos , Estudo de Associação Genômica Ampla , Cadeias HLA-DRB1/genética , Humanos , Peptídeos e Proteínas de Sinalização Intercelular , Síndrome Metabólica/patologia , Especificidade de Órgãos , Fenótipo , Locos de Características Quantitativas
11.
Science ; 336(6078): 193-8, 2012 Apr 13.
Artigo em Inglês | MEDLINE | ID: mdl-22422862

RESUMO

To study the evolution of recombination rates in apes, we developed methodology to construct a fine-scale genetic map from high-throughput sequence data from 10 Western chimpanzees, Pan troglodytes verus. Compared to the human genetic map, broad-scale recombination rates tend to be conserved, but with exceptions, particularly in regions of chromosomal rearrangements and around the site of ancestral fusion in human chromosome 2. At fine scales, chimpanzee recombination is dominated by hotspots, which show no overlap with those of humans even though rates are similarly elevated around CpG islands and decreased within genes. The hotspot-specifying protein PRDM9 shows extensive variation among Western chimpanzees, and there is little evidence that any sequence motifs are enriched in hotspots. The contrasting locations of hotspots provide a natural experiment, which demonstrates the impact of recombination on base composition.


Assuntos
Cromossomos de Mamíferos/genética , Pan troglodytes/genética , Recombinação Genética , Animais , Sequência de Bases , Mapeamento Cromossômico , Cromossomos Humanos Par 2/genética , Ilhas de CpG , Evolução Molecular , Feminino , Variação Genética , Haplótipos , Sequenciamento de Nucleotídeos em Larga Escala , Histona-Lisina N-Metiltransferase/genética , Humanos , Masculino , Polimorfismo de Nucleotídeo Único , Análise de Sequência de DNA , Especificidade da Espécie
12.
PLoS One ; 6(7): e22070, 2011.
Artigo em Inglês | MEDLINE | ID: mdl-21789213

RESUMO

The integrated analysis of genotypic and expression data for association with complex traits could identify novel genetic pathways involved in complex traits. We profiled 19,573 expression probes in Epstein-Barr virus-transformed lymphoblastoid cell lines (LCLs) from 299 twins and correlated these with 44 quantitative traits (QTs). For 939 expressed probes correlating with more than one QT, we investigated the presence of eQTL associations in three datasets of 57 CEU HapMap founders and 86 unrelated twins. Genome-wide association analysis of these probes with 2.2 m SNPs revealed 131 potential eQTLs (1,989 eQTL SNPs) overlapping between the HapMap datasets, five of which were in cis (58 eQTL SNPs). We then tested 535 SNPs tagging the eQTL SNPs, for association with the relevant QT in 2,905 twins. We identified nine potential SNP-QT associations (P<0.01) but none significantly replicated in five large consortia of 1,097-16,129 subjects. We also failed to replicate previous reported eQTL associations with body mass index, plasma low-density lipoprotein cholesterol, high-density lipoprotein cholesterol and triglycerides levels derived from lymphocytes, adipose and liver tissue. Our results and additional power calculations suggest that proponents may have been overoptimistic in the power of LCLs in eQTL approaches to elucidate regulatory genetic effects on complex traits using the small datasets generated to date. Nevertheless, larger tissue-specific expression data sets relevant to specific traits are becoming available, and should enable the adoption of similar integrated analyses in the near future.


Assuntos
Redes Reguladoras de Genes/genética , Genoma Humano/genética , Estudo de Associação Genômica Ampla , Linfócitos/metabolismo , Locos de Características Quantitativas/genética , Característica Quantitativa Herdável , Adulto , Idoso , Idoso de 80 Anos ou mais , Linhagem Celular , Estudos de Coortes , Bases de Dados Genéticas , Feminino , Regulação da Expressão Gênica , Haplótipos/genética , Humanos , Padrões de Herança/genética , Pessoa de Meia-Idade , Polimorfismo de Nucleotídeo Único/genética , Análise de Componente Principal , Reprodutibilidade dos Testes , Tamanho da Amostra , Adulto Jovem
13.
Genome Res ; 21(7): 1042-54, 2011 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-21628452

RESUMO

The human major histocompatibility complex (MHC) on chromosome 6p21 is a paradigm for genomics, showing remarkable polymorphism and striking association with immune and non-immune diseases. The complex genomic landscape of the MHC, notably strong linkage disequilibrium, has made resolving causal variants very challenging. A promising approach is to investigate gene expression levels considered as tractable intermediate phenotypes in mapping complex diseases. However, how transcription varies across the MHC, notably relative to specific haplotypes, remains unknown. Here, using an original hybrid tiling and splice junction microarray that includes alternate allele probes, we draw the first high-resolution strand-specific transcription map for three common MHC haplotypes (HLA-A1-B8-Cw7-DR3, HLA-A3-B7-Cw7-DR15, and HLA-A26-B18-Cw5-DR3-DQ2) strongly associated with autoimmune diseases including type 1 diabetes, systemic lupus erythematosus, and multiple sclerosis. We find that haplotype-specific differences in gene expression are common across the MHC, affecting 96 genes (46.4%), most significantly the zing finger protein gene ZFP57. Differentially expressed probes are correlated with polymorphisms between haplotypes, consistent with cis effects that we directly demonstrate for ZFP57 in a cohort of healthy volunteers (P = 1.2 × 10(-14)). We establish that alternative splicing is significantly more frequent in the MHC than genome-wide (72.5% vs. 62.1% of genes, P ≤ 1 × 10(-4)) and shows marked haplotypic differences. We also unmask novel and abundant intergenic transcription involving 31% of transcribed blocks identified. Our study reveals that the renowned MHC polymorphism also manifests as transcript diversity, and our novel haplotype-based approach marks a new step toward identification of regulatory variants involved in the control of MHC-associated phenotypes and diseases.


Assuntos
Perfilação da Expressão Gênica/métodos , Variação Genética , Haplótipos , Complexo Principal de Histocompatibilidade , Alelos , Processamento Alternativo , Células Cultivadas , Mapeamento Cromossômico , Diabetes Mellitus Tipo 1/genética , Predisposição Genética para Doença , Antígenos HLA-A/genética , Antígenos HLA-A/metabolismo , Humanos , Desequilíbrio de Ligação , Lúpus Eritematoso Sistêmico/genética , Esclerose Múltipla/genética , Análise de Sequência com Séries de Oligonucleotídeos , Locos de Características Quantitativas , Análise de Sequência de DNA , Transcrição Genética
14.
BMC Bioinformatics ; 10: 367, 2009 Oct 31.
Artigo em Inglês | MEDLINE | ID: mdl-19878600

RESUMO

BACKGROUND: A number of tools for the examination of linkage disequilibrium (LD) patterns between nearby alleles exist, but none are available for quickly and easily investigating LD at longer ranges (>500 kb). We have developed a web-based query tool (GLIDERS: Genome-wide LInkage DisEquilibrium Repository and Search engine) that enables the retrieval of pairwise associations with r2 >or= 0.3 across the human genome for any SNP genotyped within HapMap phase 2 and 3, regardless of distance between the markers. DESCRIPTION: GLIDERS is an easy to use web tool that only requires the user to enter rs numbers of SNPs they want to retrieve genome-wide LD for (both nearby and long-range). The intuitive web interface handles both manual entry of SNP IDs as well as allowing users to upload files of SNP IDs. The user can limit the resulting inter SNP associations with easy to use menu options. These include MAF limit (5-45%), distance limits between SNPs (minimum and maximum), r2 (0.3 to 1), HapMap population sample (CEU, YRI and JPT+CHB combined) and HapMap build/release. All resulting genome-wide inter-SNP associations are displayed on a single output page, which has a link to a downloadable tab delimited text file. CONCLUSION: GLIDERS is a quick and easy way to retrieve genome-wide inter-SNP associations and to explore LD patterns for any number of SNPs of interest. GLIDERS can be useful in identifying SNPs with long-range LD. This can highlight mis-mapping or other potential association signal localisation problems.


Assuntos
Estudo de Associação Genômica Ampla/métodos , Genoma , Desequilíbrio de Ligação/genética , Polimorfismo de Nucleotídeo Único , Ferramenta de Busca/métodos , Software , Bases de Dados Genéticas , Humanos , Internet , Interface Usuário-Computador
15.
PLoS One ; 3(3): e1898, 2008 Mar 26.
Artigo em Inglês | MEDLINE | ID: mdl-18365030

RESUMO

BACKGROUND: Vaccination against hepatitis B virus infection (HBV) is safe and effective; however, vaccine-induced antibody level wanes over time. Peak vaccine-induced anti-HBs level is directly related to antibody decay, as well as risk of infection and persistent carriage despite vaccination. We investigated the role of host genetic factors in long-term immunity against HBV infection based on peak anti-HBs level and seroconversion to anti-HBc. METHODS: We analyzed 715 SNP across 133 candidate genes in 662 infant vaccinees from The Gambia, assessing peak vaccine-induced anti-HBs level and core antibody (anti-HBc) status, whilst adjusting for covariates. A replication study comprised 43 SNPs in a further 393 individuals. RESULTS: In our initial screen we found variation in IFNG, MAPK8, and IL10RA to affect peak anti-HBs level (GMTratio of < 0.6 or > 1.5 and P < or = 0.001) and lesser associations in other genes. Odds of core-conversion was associated with variation in CD163. A coding change in ITGAL (R719V) with likely functional relevance showed evidence of association with increased peak anti-HBs level in both screens (1st screen: s595_22 GMTratio 1.71, P = 0.013; 2nd screen: s595_22 GMTratio 2.15, P = 0.011). CONCLUSION: This is to our knowledge the largest study to date assessing genetic determinants of HBV vaccine-induced immunity. We report on associations with anti-HBs level, which is directly related to durability of antibody level and predictive of vaccine efficacy long-term. A coding change in ITGAL, which plays a central role in immune cell interaction, was shown to exert beneficial effects on induction of peak antibody level in response to HBV vaccination. Variation in this gene does not appear to have been studied in relation to immune responses to viral or vaccine challenges previously. Our findings suggest that genetic variation in loci other than the HLA region affect immunity induced by HBV vaccination.


Assuntos
Anticorpos Anti-Hepatite B/biossíntese , Vacinas contra Hepatite B/administração & dosagem , Hepatite B/imunologia , Hepatite B/genética , Vacinas contra Hepatite B/imunologia , Humanos
16.
Mamm Genome ; 18(2): 123-4, 2007 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-17347895

RESUMO

We recently described methods for estimating the number of N-ethyl-N-nitrosourea (ENU)-induced coding mutations in phenotypic and genotypic screens. In this article we revisit these methods, clarifying their application. In particular, we focus on the difference between unconditional and conditional probabilities. We also introduce a website to assist investigators in the application of these equations ( http://www.well.ox.ac.uk/enuMutRat ).


Assuntos
Etilnitrosoureia/farmacologia , Testes Genéticos/métodos , Mutação , Genótipo , Mutagênese , Fenótipo , Probabilidade
17.
J Am Soc Nephrol ; 16(12): 3517-26, 2005 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-16207829

RESUMO

Autosomal dominant polycystic kidney disease (PKD) is the most common genetic disease that leads to kidney failure in humans. In addition to the known causative genes PKD1 and PKD2, there are mutations that result in cystic changes in the kidney, such as nephronophthisis, autosomal recessive polycystic kidney disease, or medullary cystic kidney disease. Recent efforts to improve the understanding of renal cystogenesis have been greatly enhanced by studies in rodent models of PKD. Genetic studies in the (cy/+) rat showed that PKD spontaneously develops as a consequence of a mutation in a gene different from the rat orthologs of PKD1 and PKD2 or other genes that are known to be involved in human cystic kidney diseases. This article reports the positional cloning and mutation analysis of the rat PKD gene, which revealed a C to T transition that replaces an arginine by a tryptophan at amino acid 823 in the protein sequence. It was determined that Pkdr1 is specifically expressed in renal proximal tubules and encodes a novel protein, SamCystin, that contains ankyrin repeats and a sterile alpha motif. The characterization of this protein, which does not share structural homologies with known polycystins, may give new insights into the pathophysiology of renal cyst development in patients.


Assuntos
Proteínas de Membrana/genética , Mutação de Sentido Incorreto , Rim Policístico Autossômico Dominante/genética , Rim Policístico Autossômico Dominante/patologia , Animais , Sequência de Bases , DNA Complementar/análise , Modelos Animais de Doenças , Regulação da Expressão Gênica , Predisposição Genética para Doença , Hibridização In Situ , Dados de Sequência Molecular , Proteínas Nucleares/agonistas , RNA/análise , Ratos , Ratos Endogâmicos BN , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Sensibilidade e Especificidade , Canais de Cátion TRPP
18.
Hum Mol Genet ; 14(18): 2757-67, 2005 Sep 15.
Artigo em Inglês | MEDLINE | ID: mdl-16103130

RESUMO

Single-nucleotide polymorphism (SNP) tagging is widely used as a way of saving genotyping costs in association studies. A number of different tagging methods have been developed to reduce the number of markers to be genotyped while maintaining power for detecting effects on non-assayed SNPs. How the different methods perform in different settings, the degree to which they overlap and share common tags and how they differ are important questions. We investigated these questions by comparing three widely used tagging methods/algorithms--one haplotype r2-based method, one pair-wise r2-based method and one method which was based on haplotype diversity but focused on major haplotypes. Tagging efficiency was defined as the number of genotyped markers divided by the number of tagging SNPs. Tagging effectiveness was defined as the proportion of un-genotyped or 'hidden' SNPs being detected (having a pair-wise or haplotype r2 with a set of tagging SNPs over a threshold, e.g. haplotype r2> or =0.80). The ENCODE regions genotyped on the HapMap CEPH individuals were examined in this study. Tagging effectiveness was generally poor for rare SNPs than for common SNPs, for all three tagging methods. Inclusion of rare SNPs into initial HapMap scheme could enhance the performance of tags on rare hidden SNPs at the expense of increased genotyping cost. At a moderate tagging efficiency, more than 90% of hidden SNPs detected by tagging SNPs selected by one method were also detected by tagging SNPs selected by another method, and this figure could be increased to 100% if tagging efficiency was allowed to drop. These results indicate that the tagging space is highly concordant between different tagging methods, despite the fact that they often involve different sets of tagging SNPs.


Assuntos
Técnicas Genéticas , Polimorfismo de Nucleotídeo Único/genética , Sitios de Sequências Rotuladas , Algoritmos , Bases de Dados Genéticas , Frequência do Gene , Marcadores Genéticos/genética , Genótipo , Haplótipos/genética
19.
Nucleic Acids Res ; 33(11): 3455-64, 2005.
Artigo em Inglês | MEDLINE | ID: mdl-15961730

RESUMO

Comparative genome hybridization (CGH) to DNA microarrays (array CGH) is a technique capable of detecting deletions and duplications in genomes at high resolution. However, array CGH studies of the human genome noting false negative and false positive results using large insert clones as probes have raised important concerns regarding the suitability of this approach for clinical diagnostic applications. Here, we adapt the Smith-Waterman dynamic-programming algorithm to provide a sensitive and robust analytic approach (SW-ARRAY) for detecting copy-number changes in array CGH data. In a blind series of hybridizations to arrays consisting of the entire tiling path for the terminal 2 Mb of human chromosome 16p, the method identified all monosomies between 267 and 1567 kb with a high degree of statistical significance and accurately located the boundaries of deletions in the range 267-1052 kb. The approach is unique in offering both a nonparametric segmentation procedure and a nonparametric test of significance. It is scalable and well-suited to high resolution whole genome array CGH studies that use array probes derived from large insert clones as well as PCR products and oligonucleotides.


Assuntos
Algoritmos , Aneuploidia , Biologia Computacional/métodos , Genômica/métodos , Análise de Sequência com Séries de Oligonucleotídeos/métodos , Software , Deleção Cromossômica , Sondas de DNA , Genoma Humano , Humanos
20.
Nat Genet ; 35(3): 258-63, 2003 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-14566338

RESUMO

Asthma is a common disease in children and young adults. Four separate reports have linked asthma and related phenotypes to an ill-defined interval between 2q14 and 2q32 (refs. 1-4), and two mouse genome screens have linked bronchial hyper-responsiveness to the region homologous to 2q14 (refs. 5,6). We found and replicated association between asthma and the D2S308 microsatellite, 800 kb distal to the IL1 cluster on 2q14. We sequenced the surrounding region and constructed a comprehensive, high-density, single-nucleotide polymorphism (SNP) linkage disequilibrium (LD) map. SNP association was limited to the initial exons of a solitary gene of 3.6 kb (DPP10), which extends over 1 Mb of genomic DNA. DPP10 encodes a homolog of dipeptidyl peptidases (DPPs) that cleave terminal dipeptides from cytokines and chemokines, and it presents a potential new target for asthma therapy.


Assuntos
Asma/genética , Cromossomos Humanos Par 2 , Sequência de Aminoácidos , Clonagem Molecular , Genótipo , Humanos , Repetições de Microssatélites/genética , Homologia de Sequência de Aminoácidos
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA