RESUMO
Bacteremia (bacterial bloodstream infection) is a major cause of illness and death in sub-Saharan Africa but little is known about the role of human genetics in susceptibility. We conducted a genome-wide association study of bacteremia susceptibility in more than 5,000 Kenyan children as part of the Wellcome Trust Case Control Consortium 2 (WTCCC2). Both the blood-culture-proven bacteremia case subjects and healthy infants as controls were recruited from Kilifi, on the east coast of Kenya. Streptococcus pneumoniae is the most common cause of bacteremia in Kilifi and was thus the focus of this study. We identified an association between polymorphisms in a long intergenic non-coding RNA (lincRNA) gene (AC011288.2) and pneumococcal bacteremia and replicated the results in the same population (p combined = 1.69 × 10(-9); OR = 2.47, 95% CI = 1.84-3.31). The susceptibility allele is African specific, derived rather than ancestral, and occurs at low frequency (2.7% in control subjects and 6.4% in case subjects). Our further studies showed AC011288.2 expression only in neutrophils, a cell type that is known to play a major role in pneumococcal clearance. Identification of this novel association will further focus research on the role of lincRNAs in human infectious disease.
Assuntos
Bacteriemia/genética , Pneumonia Pneumocócica/genética , Polimorfismo Genético/genética , RNA Longo não Codificante/genética , Streptococcus pneumoniae/genética , Adolescente , Bacteriemia/microbiologia , Bacteriemia/patologia , Estudos de Casos e Controles , Criança , Pré-Escolar , Estudo de Associação Genômica Ampla , Humanos , Lactente , Recém-Nascido , Quênia/epidemiologia , Pneumonia Pneumocócica/microbiologia , Pneumonia Pneumocócica/patologia , Fatores de RiscoRESUMO
The nuclear phase of the gene expression pathway culminates in the export of mature messenger RNAs (mRNAs) to the cytoplasm through nuclear pore complexes. GANP (germinal- centre associated nuclear protein) promotes the transfer of mRNAs bound to the transport factor NXF1 to nuclear pore complexes. Here, we demonstrate that GANP, subunit of the TRanscription-EXport-2 (TREX-2) mRNA export complex, promotes selective nuclear export of a specific subset of mRNAs whose transport depends on NXF1. Genome-wide gene expression profiling showed that half of the transcripts whose nuclear export was impaired following NXF1 depletion also showed reduced export when GANP was depleted. GANP-dependent transcripts were highly expressed, yet short-lived, and were highly enriched in those encoding central components of the gene expression machinery such as RNA synthesis and processing factors. After injection into Xenopus oocyte nuclei, representative GANP-dependent transcripts showed faster nuclear export kinetics than representative transcripts that were not influenced by GANP depletion. We propose that GANP promotes the nuclear export of specific classes of mRNAs that may facilitate rapid changes in gene expression.
Assuntos
Acetiltransferases/fisiologia , Núcleo Celular/metabolismo , Peptídeos e Proteínas de Sinalização Intracelular/fisiologia , RNA Mensageiro/metabolismo , Transporte Ativo do Núcleo Celular , Animais , Linhagem Celular Tumoral , Humanos , Proteínas de Transporte Nucleocitoplasmático/fisiologia , RNA Mensageiro/classificação , Proteínas de Ligação a RNA/fisiologia , XenopusRESUMO
We have sequenced miRNA libraries from human embryonic, neural and foetal mesenchymal stem cells. We report that the majority of miRNA genes encode mature isomers that vary in size by one or more bases at the 3' and/or 5' end of the miRNA. Northern blotting for individual miRNAs showed that the proportions of isomiRs expressed by a single miRNA gene often differ between cell and tissue types. IsomiRs were readily co-immunoprecipitated with Argonaute proteins in vivo and were active in luciferase assays, indicating that they are functional. Bioinformatics analysis predicts substantial differences in targeting between miRNAs with minor 5' differences and in support of this we report that a 5' isomiR-9-1 gained the ability to inhibit the expression of DNMT3B and NCAM2 but lost the ability to inhibit CDH1 in vitro. This result was confirmed by the use of isomiR-specific sponges. Our analysis of the miRGator database indicates that a small percentage of human miRNA genes express isomiRs as the dominant transcript in certain cell types and analysis of miRBase shows that 5' isomiRs have replaced canonical miRNAs many times during evolution. This strongly indicates that isomiRs are of functional importance and have contributed to the evolution of miRNA genes.
Assuntos
MicroRNAs/metabolismo , Animais , Proteínas Argonautas/metabolismo , Linhagem Celular , Evolução Molecular , Humanos , Camundongos , MicroRNAs/química , MicroRNAs/genética , Precursores de RNA/química , RNA Mensageiro/metabolismo , Células-Tronco/metabolismoRESUMO
Smooth muscle cell (SMC) proliferation is a hallmark of vascular injury and disease. Global hypomethylation occurs during SMC proliferation in culture and in vivo during neointimal formation. Regardless of the programmed or stochastic nature of hypomethylation, identifying these changes is important in understanding vascular disease, as maintenance of a cells' epigenetic profile is essential for maintaining cellular phenotype. Global hypomethylation of proliferating aortic SMCs and concomitant decrease of DNMT1 expression were identified in culture during passage. An epigenome screen identified regions of the genome that were hypomethylated during proliferation and a region containing Collagen, type XV, alpha 1 (COL15A1) was selected by 'genomic convergence' for characterization. COL15A1 transcript and protein levels increased with passage-dependent decreases in DNA methylation and the transcript was sensitive to treatment with 5-Aza-2'-deoxycytidine, suggesting DNA methylation-mediated gene expression. Phenotypically, knockdown of COL15A1 increased SMC migration and decreased proliferation and Col15a1 expression was induced in an atherosclerotic lesion and localized to the atherosclerotic cap. A sequence variant in COL15A1 that is significantly associated with atherosclerosis (rs4142986, P = 0.017, OR = 1.434) was methylated and methylation of the risk allele correlated with decreased gene expression and increased atherosclerosis in human aorta. In summary, hypomethylation of COL15A1 occurs during SMC proliferation and the consequent increased gene expression may impact SMC phenotype and atherosclerosis formation. Hypomethylated genes, such as COL15A1, provide evidence for concomitant epigenetic regulation and genetic susceptibility, and define a class of causal targets that sit at the intersection of genetic and epigenetic predisposition in the etiology of complex disease.
Assuntos
Aterosclerose/genética , Senescência Celular/genética , Colágeno/genética , Epigênese Genética , Aterosclerose/patologia , Movimento Celular/genética , Proliferação de Células , Células Cultivadas , Metilação de DNA/genética , Regulação da Expressão Gênica , Humanos , Miócitos de Músculo Liso/citologia , Miócitos de Músculo Liso/metabolismo , Neointima/genéticaRESUMO
We performed a genome-wide association study (GWAS) in 1705 Parkinson's disease (PD) UK patients and 5175 UK controls, the largest sample size so far for a PD GWAS. Replication was attempted in an additional cohort of 1039 French PD cases and 1984 controls for the 27 regions showing the strongest evidence of association (P< 10(-4)). We replicated published associations in the 4q22/SNCA and 17q21/MAPT chromosome regions (P< 10(-10)) and found evidence for an additional independent association in 4q22/SNCA. A detailed analysis of the haplotype structure at 17q21 showed that there are three separate risk groups within this region. We found weak but consistent evidence of association for common variants located in three previously published associated regions (4p15/BST1, 4p16/GAK and 1q32/PARK16). We found no support for the previously reported SNP association in 12q12/LRRK2. We also found an association of the two SNPs in 4q22/SNCA with the age of onset of the disease.
Assuntos
Cromossomos Humanos Par 17/genética , Predisposição Genética para Doença , Doença de Parkinson/genética , alfa-Sinucleína/genética , Idade de Início , Estudos de Casos e Controles , Estudo de Associação Genômica Ampla , Haplótipos , Humanos , Polimorfismo de Nucleotídeo Único , Tamanho da Amostra , População BrancaRESUMO
We report an alternative approach to transcriptome sequencing for the Illumina Genome Analyzer, in which the reverse transcription reaction takes place on the flowcell. No amplification is performed during the library preparation, so PCR biases and duplicates are avoided, and because the template is poly(A)(+) RNA rather than cDNA, the resulting sequences are necessarily strand-specific. The method is compatible with paired- or single-end sequencing.
Assuntos
Mapeamento Cromossômico/métodos , Transferência Ressonante de Energia de Fluorescência/métodos , Perfilação da Expressão Gênica/métodos , Análise de Sequência de DNA/métodos , Fatores de Transcrição/genética , Técnicas de Amplificação de Ácido NucleicoRESUMO
OBJECTIVE: To explore the future potential of genetic screening to detect newborns at risk of childhood-onset hearing loss. DESIGN: An expert led discussion of current and future developments in genetic technology and the knowledge base of genetic hearing loss to determine the viability of genetic screening and the implications for screening policy. RESULTS AND DISCUSSION: Despite increasing pressure to adopt genetic technologies, a major barrier for genetic screening in hearing loss is the uncertain clinical significance of the identified mutations and their interactions. Only when a reliable estimate of the future risk of hearing loss can be made at a reasonable cost, will genetic screening become viable. Given the speed of technological advancement this may be within the next 10 years. Decision-makers should start to consider how genetic screening could augment current screening programmes as well as the associated data processing and storage requirements. CONCLUSION: In the interim, we suggest that decision makers consider the benefits of (1) genetically testing all newborns and children with hearing loss, to determine aetiology and to increase knowledge of the genetic causes of hearing loss, and (2) consider screening pregnant women for the m.1555A> G mutation to reduce the risk of aminoglycoside antibiotic-associated hearing loss.
Assuntos
Análise Mutacional de DNA , Testes Genéticos , Perda Auditiva/diagnóstico , Perda Auditiva/genética , Triagem Neonatal/métodos , Idade de Início , Difusão de Inovações , Feminino , Predisposição Genética para Doença , Perda Auditiva/epidemiologia , Hereditariedade , Humanos , Recém-Nascido , Bases de Conhecimento , Linhagem , Fenótipo , Valor Preditivo dos Testes , Gravidez , Medição de Risco , Fatores de RiscoRESUMO
Inherited mutations of specific genes have elucidated the normal roles of the proteins they encode by relating specific mutations to particular phenotypes. But many potentially informative mutations in such genes are lethal early in development. Consequently, inherited mutations may not reflect all the functional roles of such proteins. Acquired, somatic defects should reflect a wider spectrum of mutations because they are not prone to negative selection in development. It has been difficult to identify such mutations so far, but microarray analysis provides a new opportunity to do so. Using this approach, we have shown that in individuals with myelodysplasia associated with alpha-thalassemia (ATMDS), somatic mutations of the gene encoding the chromatin remodeling factor ATRX cause an unexpectedly severe hematological phenotype compared with the wide spectrum of inherited mutations affecting this gene. These findings cast new light on this pleiotropic cofactor, which appears to be an essential component rather than a mere facilitator of globin gene expression.
Assuntos
DNA Helicases/genética , Mutação , Síndromes Mielodisplásicas/genética , Proteínas Nucleares/genética , Talassemia alfa/genética , Sequência de Bases , Células da Medula Óssea/metabolismo , Estudos de Casos e Controles , DNA/genética , Feminino , Expressão Gênica , Perfilação da Expressão Gênica , Globinas/genética , Humanos , Masculino , Análise de Sequência com Séries de Oligonucleotídeos , Fenótipo , Proteína Nuclear Ligada ao XRESUMO
Within the healthy population, there is substantial, heritable, and interindividual variability in the platelet response. We explored whether a proportion of this variability could be accounted for by interindividual variation in gene expression. Through a correlative analysis of genome-wide platelet RNA expression data from 37 subjects representing the normal range of platelet responsiveness within a cohort of 500 subjects, we identified 63 genes in which transcript levels correlated with variation in the platelet response to adenosine diphosphate and/or the collagen-mimetic peptide, cross-linked collagen-related peptide. Many of these encode proteins with no reported function in platelets. An association study of 6 of the 63 genes in 4235 cases and 6379 controls showed a putative association with myocardial infarction for COMMD7 (COMM domain-containing protein 7) and a major deviation from the null hypo thesis for LRRFIP1 [leucine-rich repeat (in FLII) interacting protein 1]. Morpholino-based silencing in Danio rerio identified a modest role for commd7 and a significant effect for lrrfip1 as positive regulators of thrombus formation. Proteomic analysis of human platelet LRRFIP1-interacting proteins indicated that LRRFIP1 functions as a component of the platelet cytoskeleton, where it interacts with the actin-remodeling proteins Flightless-1 and Drebrin. Taken together, these data reveal novel proteins regulating the platelet response.
Assuntos
Plaquetas/metabolismo , Perfilação da Expressão Gênica , Proteínas de Ligação a RNA/metabolismo , Animais , Inativação Gênica , Genótipo , Humanos , Ativação Plaquetária , Proteoma/metabolismo , Proteínas de Ligação a RNA/genética , Proteínas Repressoras/genética , Proteínas Repressoras/metabolismo , Trombose , Peixe-Zebra , Proteínas de Peixe-Zebra/genética , Proteínas de Peixe-Zebra/metabolismoRESUMO
It is well established that all of the cis-acting sequences required for fully regulated human alpha-globin expression are contained within a region of approximately 120 kb of conserved synteny. Here, we show that activation of this cluster in erythroid cells dramatically affects expression of apparently unrelated and noncontiguous genes in the 500 kb surrounding this domain, including a gene (NME4) located 300 kb from the alpha-globin cluster. Changes in NME4 expression are mediated by physical cis-interactions between this gene and the alpha-globin regulatory elements. Polymorphic structural variation within the globin cluster, altering the number of alpha-globin genes, affects the pattern of NME4 expression by altering the competition for the shared alpha-globin regulatory elements. These findings challenge the concept that the genome is organized into discrete, insulated regulatory domains. In addition, this work has important implications for our understanding of genome evolution, the interpretation of genome-wide expression, expression-quantitative trait loci, and copy number variant analyses.
Assuntos
Expressão Gênica , Polimorfismo de Nucleotídeo Único , Regiões Promotoras Genéticas , Cromossomos Humanos Par 16 , Humanos , Sequências Reguladoras de Ácido Nucleico , Telômero , alfa-Globinas/genéticaRESUMO
Hematopoiesis is a carefully controlled process that is regulated by complex networks of transcription factors that are, in part, controlled by signals resulting from ligand binding to cell-surface receptors. To further understand hematopoiesis, we have compared gene expression profiles of human erythroblasts, megakaryocytes, B cells, cytotoxic and helper T cells, natural killer cells, granulocytes, and monocytes using whole genome microarrays. A bioinformatics analysis of these data was performed focusing on transcription factors, immunoglobulin superfamily members, and lineage-specific transcripts. We observed that the numbers of lineage-specific genes varies by 2 orders of magnitude, ranging from 5 for cytotoxic T cells to 878 for granulocytes. In addition, we have identified novel coexpression patterns for key transcription factors involved in hematopoiesis (eg, GATA3-GFI1 and GATA2-KLF1). This study represents the most comprehensive analysis of gene expression in hematopoietic cells to date and has identified genes that play key roles in lineage commitment and cell function. The data, which are freely accessible, will be invaluable for future studies on hematopoiesis and the role of specific genes and will also aid the understanding of the recent genome-wide association studies.
Assuntos
Células da Medula Óssea/fisiologia , Diferenciação Celular/genética , Expressão Gênica , Atlas como Assunto , Linhagem da Célula , Células Cultivadas , Citometria de Fluxo , Perfilação da Expressão Gênica , Hematopoese , Humanos , Análise de Sequência com Séries de Oligonucleotídeos , Fatores de Transcrição/metabolismoRESUMO
CpG islands (CGIs) are dense clusters of CpG sequences that punctuate the CpG-deficient human genome and associate with many gene promoters. As CGIs also differ from bulk chromosomal DNA by their frequent lack of cytosine methylation, we devised a CGI enrichment method based on nonmethylated CpG affinity chromatography. The resulting library was sequenced to define a novel human blood CGI set that includes many that are not detected by current algorithms. Approximately half of CGIs were associated with annotated gene transcription start sites, the remainder being intra- or intergenic. Using an array representing over 17,000 CGIs, we established that 6%-8% of CGIs are methylated in genomic DNA of human blood, brain, muscle, and spleen. Inter- and intragenic CGIs are preferentially susceptible to methylation. CGIs showing tissue-specific methylation were overrepresented at numerous genetic loci that are essential for development, including HOX and PAX family members. The findings enable a comprehensive analysis of the roles played by CGI methylation in normal and diseased human tissues.
Assuntos
Ilhas de CpG/fisiologia , Metilação de DNA , Regulação da Expressão Gênica no Desenvolvimento , Genes Controladores do Desenvolvimento , Mapeamento Cromossômico , Feminino , Biblioteca Gênica , Genoma Humano , Humanos , Masculino , Especificidade de Órgãos , Distribuição TecidualRESUMO
Monitoring the spread of SARS-CoV-2 and reconstructing transmission chains has become a major public health focus for many governments around the world. The modest mutation rate and rapid transmission of SARS-CoV-2 prevents the reconstruction of transmission chains from consensus genome sequences, but within-host genetic diversity could theoretically help identify close contacts. Here we describe the patterns of within-host diversity in 1181 SARS-CoV-2 samples sequenced to high depth in duplicate. 95.1% of samples show within-host mutations at detectable allele frequencies. Analyses of the mutational spectra revealed strong strand asymmetries suggestive of damage or RNA editing of the plus strand, rather than replication errors, dominating the accumulation of mutations during the SARS-CoV-2 pandemic. Within- and between-host diversity show strong purifying selection, particularly against nonsense mutations. Recurrent within-host mutations, many of which coincide with known phylogenetic homoplasies, display a spectrum and patterns of purifying selection more suggestive of mutational hotspots than recombination or convergent evolution. While allele frequencies suggest that most samples result from infection by a single lineage, we identify multiple putative examples of co-infection. Integrating these results into an epidemiological inference framework, we find that while sharing of within-host variants between samples could help the reconstruction of transmission chains, mutational hotspots and rare cases of superinfection can confound these analyses.
The COVID-19 pandemic has had major health impacts across the globe. The scientific community has focused much attention on finding ways to monitor how the virus responsible for the pandemic, SARS-CoV-2, spreads. One option is to perform genetic tests, known as sequencing, on SARS-CoV-2 samples to determine the genetic code of the virus and to find any differences or mutations in the genes between the viral samples. Viruses mutate within their hosts and can develop into variants that are able to more easily transmit between hosts. Genetic sequencing can reveal how genetically similar two SARS-CoV-2 samples are. But tracking how SARS-CoV-2 moves from one person to the next through sequencing can be tricky. Even a sample of SARS-CoV-2 viruses from the same individual can display differences in their genetic material or within-host variants. Could genetic testing of within-host variants shed light on factors driving SARS-CoV-2 to evolve in humans? To get to the bottom of this, Tonkin-Hill, Martincorena et al. probed the genetics of SARS-CoV-2 within-host variants using 1,181 samples. The analyses revealed that 95.1% of samples contained within-host variants. A number of variants occurred frequently in many samples, which were consistent with mutational hotspots in the SARS-CoV-2 genome. In addition, within-host variants displayed mutation patterns that were similar to patterns found between infected individuals. The shared within-host variants between samples can help to reconstruct transmission chains. However, the observed mutational hotspots and the detection of multiple strains within an individual can make this challenging. These findings could be used to help predict how SARS-CoV-2 evolves in response to interventions such as vaccines. They also suggest that caution is needed when using information on within-host variants to determine transmission between individuals.
Assuntos
COVID-19/genética , COVID-19/fisiopatologia , Variação Genética , Genoma Viral , Interações Hospedeiro-Patógeno/genética , Mutação , SARS-CoV-2/genética , Sequência de Bases , Humanos , Pandemias , FilogeniaRESUMO
COVID-19 poses a major challenge to care homes, as SARS-CoV-2 is readily transmitted and causes disproportionately severe disease in older people. Here, 1167 residents from 337 care homes were identified from a dataset of 6600 COVID-19 cases from the East of England. Older age and being a care home resident were associated with increased mortality. SARS-CoV-2 genomes were available for 700 residents from 292 care homes. By integrating genomic and temporal data, 409 viral clusters within the 292 homes were identified, indicating two different patterns - outbreaks among care home residents and independent introductions with limited onward transmission. Approximately 70% of residents in the genomic analysis were admitted to hospital during the study, providing extensive opportunities for transmission between care homes and hospitals. Limiting viral transmission within care homes should be a key target for infection control to reduce COVID-19 mortality in this population.
Assuntos
COVID-19/epidemiologia , COVID-19/transmissão , Casas de Saúde , SARS-CoV-2/genética , Idoso de 80 Anos ou mais , COVID-19/virologia , Surtos de Doenças , Inglaterra/epidemiologia , Feminino , Humanos , Transmissão de Doença Infecciosa do Paciente para o Profissional , Transmissão de Doença Infecciosa do Profissional para o Paciente , Masculino , Polimorfismo de Nucleotídeo Único , Análise de Sequência , Fatores de TempoRESUMO
Acknowledgement of the breadth of T-cell pleiotropy has provoked increasing interest in the degree to which functional responsiveness is elicited by environmental cues versus differentiation. This is particularly relevant for young animals requiring rapid responses to acute environmental exposure. In young mice, gammadelta T cells are disproportionately important for immuno-protection. To examine the situation in humans, we compared populations and clones of T cells from term and preterm babies, and adults. By comparison with alphabeta T cells, neonate-derived gammadelta cells show stronger, pleiotropic functional responsiveness, and lack signatory deficits in IFN-gamma production. Emphasising the acquisition of functional competence in utero, IFN-gamma was produced by gammadelta cells sampled from premature births, and, although one month's post-partum environmental exposure invariably increased their TNF-alpha production, it had no consistent effect on IFN-gamma or IL-2. In sum, gammadelta cells seem well positioned at birth to contribute to immuno-protection and immuno-regulation, possibly compensating for selective immaturity in the alphabeta compartment. With regard to the susceptibilities of preterm babies to viral infection, gammadelta cells from preterm neonates were commonly impaired in Toll-like receptor-3 and -7 expression and compared with cells from term babies failed to optimise cytokine production in response to coincident TCR and TLR agonists.
Assuntos
Recém-Nascido Prematuro/imunologia , Receptores de Antígenos de Linfócitos T gama-delta/metabolismo , Linfócitos T/imunologia , Adulto , Antígenos CD/genética , Antígenos de Diferenciação de Linfócitos T/genética , Fatores de Transcrição Forkhead/genética , Perfilação da Expressão Gênica , Regulação da Expressão Gênica no Desenvolvimento , Humanos , Recém-Nascido , Recém-Nascido Prematuro/sangue , Recém-Nascido Prematuro/metabolismo , Interferon gama/metabolismo , Interleucina-2/metabolismo , Lectinas Tipo C , Análise de Sequência com Séries de Oligonucleotídeos , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Receptores de Antígenos de Linfócitos T alfa-beta/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Linfócitos T/citologia , Linfócitos T/metabolismo , Fatores de Tempo , Receptor 3 Toll-Like/genética , Receptor 7 Toll-Like/genética , Fator de Necrose Tumoral alfa/metabolismoRESUMO
Histone H3 methylation at R17 and R26 recently emerged as a novel epigenetic mechanism regulating pluripotency in mouse embryos. Blastomeres of four-cell embryos with high H3 methylation at these sites show unrestricted potential, whereas those with lower levels cannot support development when aggregated in chimeras of like cells. Increasing histone H3 methylation, through expression of coactivator-associated-protein-arginine-methyltransferase 1 (CARM1) in embryos, elevates expression of key pluripotency genes and directs cells to the pluripotent inner cell mass. We demonstrate CARM1 is also required for the self-renewal and pluripotency of embryonic stem (ES) cells. In ES cells, CARM1 depletion downregulates pluripotency genes leading to their differentiation. CARM1 associates with Oct4/Pou5f1 and Sox2 promoters that display detectable levels of R17/26 histone H3 methylation. In CARM1 overexpressing ES cells, histone H3 arginine methylation is also at the Nanog promoter to which CARM1 now associates. Such cells express Nanog at elevated levels and delay their response to differentiation signals. Thus, like in four-cell embryo blastomeres, histone H3 arginine methylation by CARM1 in ES cells allows epigenetic modulation of pluripotency.
Assuntos
Diferenciação Celular/fisiologia , Células-Tronco Embrionárias/citologia , Células-Tronco Embrionárias/metabolismo , Células-Tronco Pluripotentes/citologia , Células-Tronco Pluripotentes/metabolismo , Proteína-Arginina N-Metiltransferases/metabolismo , Animais , Arginina/metabolismo , Western Blotting , Diferenciação Celular/genética , Linhagem Celular , Imunoprecipitação da Cromatina , Histonas/química , Histonas/metabolismo , Proteínas de Homeodomínio/genética , Metilação , Camundongos , Proteína Homeobox Nanog , Fator 3 de Transcrição de Octâmero/genética , Análise de Sequência com Séries de Oligonucleotídeos , Regiões Promotoras Genéticas/genética , Regiões Promotoras Genéticas/fisiologia , Interferência de RNA , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Fatores de Transcrição SOXB1/genéticaRESUMO
Recurrent chromosomal aberrations in solid tumors can reveal the genetic pathways involved in the evolution of a malignancy and in some cases predict biological behavior. However, the role of individual genetic backgrounds in shaping karyotypes of sporadic tumors is unknown. The genetic structure of purebred dog breeds, coupled with their susceptibility to spontaneous cancers, provides a robust model with which to address this question. We tested the hypothesis that there is an association between breed and the distribution of genomic copy number imbalances in naturally occurring canine tumors through assessment of a cohort of Golden Retrievers and Rottweilers diagnosed with spontaneous appendicular osteosarcoma. Our findings reveal significant correlations between breed and tumor karyotypes that are independent of gender, age at diagnosis, and histological classification. These data indicate for the first time that individual genetic backgrounds, as defined by breed in dogs, influence tumor karyotypes in a cancer with extensive genomic instability.
Assuntos
Aberrações Cromossômicas/veterinária , Doenças do Cão/genética , Predisposição Genética para Doença/genética , Osteossarcoma/veterinária , Animais , Hibridização Genômica Comparativa , Cães , Cariotipagem/veterinária , Osteossarcoma/genética , Especificidade da EspécieRESUMO
Injection-site-associated sarcomas (ISAS), commonly arising at the site of routine vaccine administration, afflict as many as 22,000 domestic cats annually in the USA. These tumors are typically more aggressive and prone to recurrence than spontaneous sarcomas (non-ISAS), generally receiving a poorer long-term prognosis and warranting a more aggressive therapeutic approach. Although certain clinical and histological factors are highly suggestive of ISAS, timely diagnosis and optimal clinical management may be hindered by the absence of definitive markers that can distinguish between tumors with underlying injection-related etiology and their spontaneous counterpart. Specific nonrandom chromosome copy number aberrations (CNAs) have been associated with the clinical behavior of a vast spectrum of human tumors, providing an extensive resource of potential diagnostic and prognostic biomarkers. Although similar principles are now being applied with great success in other species, their relevance to feline molecular oncology has not yet been investigated in any detail. We report the construction of a genomic microarray platform for detection of recurrent CNAs in feline tumors through cytogenetic assignment of 210 large-insert DNA clones selected at intervals of approximately 15 Mb from the feline genome sequence assembly. Microarray-based profiling of 19 ISAS and 27 non-ISAS cases identified an extensive range of genomic imbalances that were highly recurrent throughout the combined panel of 46 sarcomas. Deletions of two specific regions were significantly associated with the non-ISAS phenotype. Further characterization of these regions may ultimately permit molecular distinction between ISAS and non-ISAS, as a tool for predicting tumor behavior and prognosis, as well as refining means for therapeutic intervention.
Assuntos
Variações do Número de Cópias de DNA , Sarcoma/genética , Animais , Doenças do Gato/diagnóstico , Gatos , Análise Citogenética , Dosagem de Genes , Perfilação da Expressão Gênica , Injeções/efeitos adversos , Análise de Sequência com Séries de Oligonucleotídeos , Sarcoma/diagnóstico , Sarcoma/veterináriaRESUMO
BACKGROUND: Autism comprises a spectrum of behavioral and cognitive disturbances of childhood development and is known to be highly heritable. Although numerous approaches have been used to identify genes implicated in the development of autism, less than 10% of autism cases have been attributed to single gene disorders. METHODS: We describe the use of high-resolution genome-wide tilepath microarrays and comparative genomic hybridization to identify copy number variants within 119 probands from multiplex autism families. We next carried out DNA methylation analysis by bisulfite sequencing in a proband and his family, expanding this analysis to methylation analysis of peripheral blood and temporal cortex DNA of autism cases and matched controls from independent datasets. We also assessed oxytocin receptor (OXTR) gene expression within the temporal cortex tissue by quantitative real-time polymerase chain reaction (PCR). RESULTS: Our analysis revealed a genomic deletion containing the oxytocin receptor gene, OXTR (MIM accession no.: 167055), previously implicated in autism, was present in an autism proband and his mother who exhibits symptoms of obsessive-compulsive disorder. The proband's affected sibling did not harbor this deletion but instead may exhibit epigenetic misregulation of this gene through aberrant gene silencing by DNA methylation. Further DNA methylation analysis of the CpG island known to regulate OXTR expression identified several CpG dinucleotides that show independent statistically significant increases in the DNA methylation status in the peripheral blood cells and temporal cortex in independent datasets of individuals with autism as compared to control samples. Associated with the increase in methylation of these CpG dinucleotides is our finding that OXTR mRNA showed decreased expression in the temporal cortex tissue of autism cases matched for age and sex compared to controls. CONCLUSION: Together, these data provide further evidence for the role of OXTR and the oxytocin signaling pathway in the etiology of autism and, for the first time, implicate the epigenetic regulation of OXTR in the development of the disorder.See the related commentary by Gurrieri and Neri: http://www.biomedcentral.com/1741-7015/7/63.
Assuntos
Transtorno Autístico/genética , DNA/genética , DNA/metabolismo , Receptores de Ocitocina/deficiência , Adolescente , Adulto , Criança , Pré-Escolar , Hibridização Genômica Comparativa , Metilação de DNA , Feminino , Humanos , Masculino , Análise em Microsséries , Deleção de Sequência , Adulto JovemRESUMO
Numerous attributes render the domestic dog a highly pertinent model for cancer-associated gene discovery. We performed microarray-based comparative genomic hybridization analysis of 60 spontaneous canine intracranial tumors to examine the degree to which dog and human patients exhibit aberrations of ancestrally related chromosome regions, consistent with a shared pathogenesis. Canine gliomas and meningiomas both demonstrated chromosome copy number aberrations (CNAs) that share evolutionarily conserved synteny with those previously reported in their human counterpart. Interestingly, however, genomic imbalances orthologous to some of the hallmark aberrations of human intracranial tumors, including chromosome 22/NF2 deletions in meningiomas and chromosome 1p/19q deletions in oligodendrogliomas, were not major events in the dog. Furthermore, and perhaps most significantly, we identified highly recurrent CNAs in canine intracranial tumors for which the human orthologue has been reported previously at low frequency but which have not, thus far, been associated intimately with the pathogenesis of the tumor. The presence of orthologous CNAs in canine and human intracranial cancers is strongly suggestive of their biological significance in tumor development and/or progression. Moreover, the limited genetic heterogenity within purebred dog populations, coupled with the contrasting organization of the dog and human karyotypes, offers tremendous opportunities for refining evolutionarily conserved regions of tumor-associated genomic imbalance that may harbor novel candidate genes involved in their pathogenesis. A comparative approach to the study of canine and human intracranial tumors may therefore provide new insights into their genetic etiology, towards development of more sophisticated molecular subclassification and tailored therapies in both species.