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1.
Eur J Hum Genet ; 2019 Sep 26.
Artigo em Inglês | MEDLINE | ID: mdl-31558840

RESUMO

Insights into individual differences in gene expression and its heritability (h2) can help in understanding pathways from DNA to phenotype. We estimated the heritability of gene expression of 52,844 genes measured in whole blood in the largest twin RNA-Seq sample to date (1497 individuals including 459 monozygotic twin pairs and 150 dizygotic twin pairs) from classical twin modeling and identity-by-state-based approaches. We estimated for each gene h2total, composed of cis-heritability (h2cis, the variance explained by single nucleotide polymorphisms in the cis-window of the gene), and trans-heritability (h2res, the residual variance explained by all other genome-wide variants). Mean h2total was 0.26, which was significantly higher than heritability estimates earlier found in a microarray-based study using largely overlapping (>60%) RNA samples (mean h2 = 0.14, p = 6.15 × 10-258). Mean h2cis was 0.06 and strongly correlated with beta of the top cis expression quantitative loci (eQTL, ρ = 0.76, p < 10-308) and with estimates from earlier RNA-Seq-based studies. Mean h2res was 0.20 and correlated with the beta of the corresponding trans-eQTL (ρ = 0.04, p < 1.89 × 10-3) and was significantly higher for genes involved in cytokine-cytokine interactions (p = 4.22 × 10-15), many other immune system pathways, and genes identified in genome-wide association studies for various traits including behavioral disorders and cancer. This study provides a thorough characterization of cis- and trans-h2 estimates of gene expression, which is of value for interpretation of GWAS and gene expression studies.

2.
Arthritis Rheumatol ; 2018 Oct 09.
Artigo em Inglês | MEDLINE | ID: mdl-30298554

RESUMO

OBJECTIVES: Multiple osteoarthritis (OA) susceptibility single nucleotide polymorphisms (SNPs) mark imbalanced expression of positional genes in articular cartilage, reflected by unequally expressed alleles among heterozygotes (allelic imbalance, AI). Here, we explored the articular cartilage transcriptome from OA patients for AI events to identify putative disease driving genetic variation. METHODS: AI was assessed in N=42 preserved and N=5 lesioned OA cartilage samples (RAAK-study) for which RNA sequencing data was available. Hereto, the count fraction of the alternative allele among the alternative and reference allele together (phi) was determined for heterozygous individuals. A meta-analysis per SNP was performed to generate a meta-phi and P-value per SNP with a multiple testing correcting false discovery rate (FDR). To further validate AI events, we explored them as function of multiple additional OA features. RESULTS: In total 2070 SNPs were observed that consistently marked AI of 1031 unique genes in articular cartilage. Of these 49 genes were found significantly differentially expressed (0.5>FC>2, FDR<0.05) between preserved and paired lesioned cartilage and 21 genes previously reported to confer susceptibility for OA and/or related phenotypes. Moreover, notable highly significant AI SNPs in the CRLF1, WWP2, and RPS3 genes were identified that were related to multiple OA features. CONCLUSION: A framework and resulting datasets for researchers in the OA research field to probe for disease relevant genetic variation that affects respective gene expression in the disease's pivotal affected tissue and likely including putative novel compelling OA risk genes such as CRLF1, WWP2, and RPS3. This article is protected by copyright. All rights reserved.

3.
Nat Commun ; 9(1): 3097, 2018 08 06.
Artigo em Inglês | MEDLINE | ID: mdl-30082726

RESUMO

Identification of causal drivers behind regulatory gene networks is crucial in understanding gene function. Here, we develop a method for the large-scale inference of gene-gene interactions in observational population genomics data that are both directed (using local genetic instruments as causal anchors, akin to Mendelian Randomization) and specific (by controlling for linkage disequilibrium and pleiotropy). Analysis of genotype and whole-blood RNA-sequencing data from 3072 individuals identified 49 genes as drivers of downstream transcriptional changes (Wald P < 7 × 10-10), among which transcription factors were overrepresented (Fisher's P = 3.3 × 10-7). Our analysis suggests new gene functions and targets, including for SENP7 (zinc-finger genes involved in retroviral repression) and BCL2A1 (target genes possibly involved in auditory dysfunction). Our work highlights the utility of population genomics data in deriving directed gene expression networks. A resource of trans-effects for all 6600 genes with a genetic instrument can be explored individually using a web-based browser.

4.
Nat Commun ; 9(1): 1873, 2018 05 14.
Artigo em Inglês | MEDLINE | ID: mdl-29760424

RESUMO

In contrast to mouse, human female germ cells develop asynchronously. Germ cells transition to meiosis, erase genomic imprints, and reactivate the X chromosome. It is unknown if these events all appear asynchronously, and how they relate to each other. Here we combine exome sequencing of human fetal and maternal tissues with single-cell RNA-sequencing of five donors. We reconstruct full parental haplotypes and quantify changes in parental allele-specific expression, genome-wide. First we distinguish primordial germ cells (PGC), pre-meiotic, and meiotic transcriptional stages. Next we demonstrate that germ cells from various stages monoallelically express imprinted genes and confirm this by methylation patterns. Finally, we show that roughly 30% of the PGCs are still reactivating their inactive X chromosome and that this is related to transcriptional stage rather than fetal age. Altogether, we uncover the complexity and cell-to-cell heterogeneity of transcriptional and epigenetic remodeling in female human germ cells.


Assuntos
Cromossomos Humanos X/química , Epigênese Genética , Óvulo/metabolismo , Transcriptoma , Aborto Legal , Adulto , Cromossomos Humanos X/metabolismo , Metilação de DNA , Feminino , Feto , Heterogeneidade Genética , Impressão Genômica , Haplótipos , Humanos , Masculino , Meiose , Óvulo/crescimento & desenvolvimento , Gravidez , Trimestres da Gravidez , Análise de Célula Única/métodos , Sequenciamento Completo do Exoma , Inativação do Cromossomo X
5.
Biochem J ; 475(6): 1075-1089, 2018 03 20.
Artigo em Inglês | MEDLINE | ID: mdl-29487166

RESUMO

Oct4 and Sox2 regulate the expression of target genes such as Nanog, Fgf4, and Utf1, by binding to their respective regulatory motifs. Their functional cooperation is reflected in their ability to heterodimerize on adjacent cis regulatory motifs, the composite Sox/Oct motif. Given that Oct4 and Sox2 regulate many developmental genes, a quantitative analysis of their synergistic action on different Sox/Oct motifs would yield valuable insights into the mechanisms of early embryonic development. In the present study, we measured binding affinities of Oct4 and Sox2 to different Sox/Oct motifs using fluorescence correlation spectroscopy. We found that the synergistic binding interaction is driven mainly by the level of Sox2 in the case of the Fgf4 Sox/Oct motif. Taking into account Sox2 expression levels fluctuate more than Oct4, our finding provides an explanation on how Sox2 controls the segregation of the epiblast and primitive endoderm populations within the inner cell mass of the developing rodent blastocyst.

6.
Eur J Hum Genet ; 26(1): 94-106, 2018 01.
Artigo em Inglês | MEDLINE | ID: mdl-29162933

RESUMO

Facioscapulohumeral muscular dystrophy is caused by incomplete repression of the transcription factor DUX4 in skeletal muscle as a consequence of D4Z4 macrosatellite repeat contraction in chromosome 4q35 (FSHD1) or variants in genes encoding D4Z4 chromatin repressors (FSHD2). A clinical hallmark of FSHD is variability in onset and progression suggesting the presence of disease modifiers. A well-known cis modifier is the polymorphic DUX4 polyadenylation signal (PAS) that defines FSHD permissive alleles: D4Z4 chromatin relaxation on non-permissive alleles which lack the DUX4-PAS cannot cause disease in the absence of stable DUX4 mRNA. We have explored the nature and relevance of a common variant of the major FSHD haplotype 4A161, which is defined by 1.6 kb size difference of the most distal D4Z4 repeat unit. While the short variant (4A161S) has been extensively studied, we demonstrate that the long variant (4A161L) is relatively common in the European population, is capable of expressing DUX4, but that DUX4 mRNA processing differs from 4A161S. While we do not find evidence for a difference in disease severity between FSHD carriers of an 4A161S or 4A161L allele, our study does uncover biallelic DUX4 expression in FSHD2 patients. Compared to control individuals, we observed an increased frequency of FSHD2 patients homozygous for disease permissive alleles, and who are thus capable of biallelic DUX4 expression, while SMCHD1 variant carriers with only one permissive allele were significantly more often asymptomatic. This suggests that biallelic DUX4 expression lowers the threshold for disease presentation and is a modifier for disease severity in FSHD2.

7.
Nat Genet ; 49(1): 139-145, 2017 01.
Artigo em Inglês | MEDLINE | ID: mdl-27918533

RESUMO

Genetic risk factors often localize to noncoding regions of the genome with unknown effects on disease etiology. Expression quantitative trait loci (eQTLs) help to explain the regulatory mechanisms underlying these genetic associations. Knowledge of the context that determines the nature and strength of eQTLs may help identify cell types relevant to pathophysiology and the regulatory networks underlying disease. Here we generated peripheral blood RNA-seq data from 2,116 unrelated individuals and systematically identified context-dependent eQTLs using a hypothesis-free strategy that does not require previous knowledge of the identity of the modifiers. Of the 23,060 significant cis-regulated genes (false discovery rate (FDR) ≤ 0.05), 2,743 (12%) showed context-dependent eQTL effects. The majority of these effects were influenced by cell type composition. A set of 145 cis-eQTLs depended on type I interferon signaling. Others were modulated by specific transcription factors binding to the eQTL SNPs.


Assuntos
Proteínas Sanguíneas/genética , Linhagem da Célula/genética , Predisposição Genética para Doença , Polimorfismo de Nucleotídeo Único/genética , Locos de Características Quantitativas/genética , RNA Mensageiro/sangue , Sequências Reguladoras de Ácido Nucleico/genética , Estudos de Coortes , Feminino , Perfilação da Expressão Gênica , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Masculino , Pessoa de Meia-Idade , RNA Mensageiro/genética
8.
Nat Genet ; 49(1): 131-138, 2017 01.
Artigo em Inglês | MEDLINE | ID: mdl-27918535

RESUMO

Most disease-associated genetic variants are noncoding, making it challenging to design experiments to understand their functional consequences. Identification of expression quantitative trait loci (eQTLs) has been a powerful approach to infer the downstream effects of disease-associated variants, but most of these variants remain unexplained. The analysis of DNA methylation, a key component of the epigenome, offers highly complementary data on the regulatory potential of genomic regions. Here we show that disease-associated variants have widespread effects on DNA methylation in trans that likely reflect differential occupancy of trans binding sites by cis-regulated transcription factors. Using multiple omics data sets from 3,841 Dutch individuals, we identified 1,907 established trait-associated SNPs that affect the methylation levels of 10,141 different CpG sites in trans (false discovery rate (FDR) < 0.05). These included SNPs that affect both the expression of a nearby transcription factor (such as NFKB1, CTCF and NKX2-3) and methylation of its respective binding site across the genome. Trans methylation QTLs effectively expose the downstream effects of disease-associated variants.


Assuntos
Metilação de DNA , Doença/genética , Regulação da Expressão Gênica , Polimorfismo de Nucleotídeo Único/genética , Locos de Características Quantitativas , Fatores de Transcrição/metabolismo , Sítios de Ligação , Estudos de Coortes , Feminino , Genoma Humano , Estudo de Associação Genômica Ampla , Humanos , Masculino , Pessoa de Meia-Idade , Fenótipo
9.
FASEB J ; 31(1): 238-255, 2017 01.
Artigo em Inglês | MEDLINE | ID: mdl-27733450

RESUMO

Skeletal muscle fibrosis and impaired muscle regeneration are major contributors to muscle wasting in Duchenne muscular dystrophy (DMD). Muscle growth is negatively regulated by myostatin (MSTN) and activins. Blockage of these pathways may improve muscle quality and function in DMD. Antisense oligonucleotides (AONs) were designed specifically to block the function of ALK4, a key receptor for the MSTN/activin pathway in skeletal muscle. AON-induced exon skipping resulted in specific Alk4 down-regulation, inhibition of MSTN activity, and increased myoblast differentiation in vitro Unexpectedly, a marked decrease in muscle mass (10%) was found after Alk4 AON treatment in mdx mice. In line with in vitro results, muscle regeneration was stimulated, and muscle fiber size decreased markedly. Notably, when Alk4 was down-regulated in adult wild-type mice, muscle mass decreased even more. RNAseq analysis revealed dysregulated metabolic functions and signs of muscle atrophy. We conclude that ALK4 inhibition increases myogenesis but also regulates the tight balance of protein synthesis and degradation. Therefore, caution must be used when developing therapies that interfere with MSTN/activin pathways.-Pasteuning-Vuhman, S., Boertje-van der Meulen, J. W., van Putten, M., Overzier, M., ten Dijke, P., Kielbasa, S. M., Arindrarto, W., Wolterbeek, R., Lezhnina, K. V., Ozerov, I. V., Aliper, A. M., Hoogaars, W. M., Aartsma-Rus, A., Loomans, C. J. M. New function of the myostatin/activin type I receptor (ALK4) as a mediator of muscle atrophy and muscle regeneration.


Assuntos
Receptores de Ativinas Tipo I/metabolismo , Músculo Esquelético/fisiologia , Mioblastos/fisiologia , Regeneração/fisiologia , Receptores de Ativinas Tipo I/genética , Animais , Sequência de Bases , Linhagem Celular , Dano ao DNA , Regulação para Baixo , Regulação da Expressão Gênica/fisiologia , Camundongos , Camundongos Endogâmicos mdx , Desenvolvimento Muscular/fisiologia , Oligonucleotídeos Antissenso/farmacologia , RNA/genética , RNA/metabolismo , Regeneração/genética , Transdução de Sinais
10.
Elife ; 52016 12 20.
Artigo em Inglês | MEDLINE | ID: mdl-27997336

RESUMO

The regeneration-capable flatworm Macrostomum lignano is a powerful model organism to study the biology of stem cells in vivo. As a flatworm amenable to transgenesis, it complements the historically used planarian flatworm models, such as Schmidtea mediterranea. However, information on the transcriptome and markers of stem cells in M. lignano is limited. We generated a de novo transcriptome assembly and performed the first comprehensive characterization of gene expression in the proliferating cells of M. lignano, represented by somatic stem cells, called neoblasts, and germline cells. Knockdown of a selected set of neoblast genes, including Mlig-ddx39, Mlig-rrm1, Mlig-rpa3, Mlig-cdk1, and Mlig-h2a, confirmed their crucial role for the functionality of somatic neoblasts during homeostasis and regeneration. The generated M. lignano transcriptome assembly and gene expression signatures of somatic neoblasts and germline cells will be a valuable resource for future molecular studies in M. lignano.


Assuntos
Células Germinativas/fisiologia , Platelmintos/citologia , Platelmintos/genética , Células-Tronco/fisiologia , Transcriptoma , Animais , Perfilação da Expressão Gênica
11.
Oncotarget ; 7(8): 8513-23, 2016 Feb 23.
Artigo em Inglês | MEDLINE | ID: mdl-26885755

RESUMO

Chronic muscle diseases are highly prevalent in the elderly causing severe mobility limitations, pain and frailty. The intrinsic molecular mechanisms are poorly understood due to multifactorial causes, slow progression with age and variations between individuals. Understanding the underlying molecular mechanisms could lead to new treatment options which are currently limited. Shoulder complaints are highly common in the elderly, and therefore, muscles of the shoulder's rotator cuff could be considered as a model for chronic age-associated muscle degeneration. Diseased shoulder muscles were characterized by muscle atrophy and fatty infiltration compared with unaffected shoulder muscles. We confirmed fatty infiltration using histochemical analysis. Additionally, fibrosis and loss of contractile myosin expression were found in diseased muscles. Most cellular features, including proliferation rate, apoptosis and cell senescence, remained unchanged and genome-wide molecular signatures were predominantly similar between diseased and intact muscles. However, we found down-regulation of a small subset of muscle function genes, and up-regulation of extracellular region genes. Myogenesis was defected in muscle cell culture from diseased muscles but was restored by elevating MyoD levels. We suggest that impaired muscle functionality in a specific environment of thickened extra-cellular matrix is crucial for the development of chronic age-associated muscle degeneration.


Assuntos
Envelhecimento/fisiologia , Biomarcadores/metabolismo , Músculo Esquelético/patologia , Atrofia Muscular/patologia , Manguito Rotador/patologia , Ombro/patologia , Tecido Adiposo/patologia , Estudos de Casos e Controles , Células Cultivadas , Feminino , Perfilação da Expressão Gênica , Humanos , Masculino , Pessoa de Meia-Idade , Músculo Esquelético/metabolismo , Atrofia Muscular/metabolismo , Atrofia Muscular/cirurgia , Mioblastos/citologia , Mioblastos/metabolismo , RNA Mensageiro/genética , Reação em Cadeia da Polimerase em Tempo Real , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Manguito Rotador/metabolismo , Manguito Rotador/cirurgia , Ombro/cirurgia
12.
Stem Cell Reports ; 4(6): 1112-24, 2015 Jun 09.
Artigo em Inglês | MEDLINE | ID: mdl-26028532

RESUMO

Differentiated derivatives of human pluripotent stem cells in culture are generally phenotypically immature compared to their adult counterparts. Their identity is often difficult to determine with certainty because little is known about their human fetal equivalents in vivo. Cellular identity and signaling pathways directing differentiation are usually determined by extrapolating information from either human adult tissue or model organisms, assuming conservation with humans. To resolve this, we generated a collection of human fetal transcriptional profiles at different developmental stages. Moreover, we developed an algorithm, KeyGenes, which uses this dataset to quantify the extent to which next-generation sequencing or microarray data resemble specific cell or tissue types in the human fetus. Using KeyGenes combined with the human fetal atlas, we identified multiple cell and tissue samples unambiguously on a limited set of features. We thus provide a flexible and expandable platform to monitor and evaluate the efficiency of differentiation in vitro.


Assuntos
Algoritmos , Feto/metabolismo , Diferenciação Celular , Bases de Dados Factuais , Feminino , Feto/citologia , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Análise de Sequência com Séries de Oligonucleotídeos , Células-Tronco Pluripotentes/citologia , Células-Tronco Pluripotentes/metabolismo , Gravidez , Primeiro Trimestre da Gravidez/genética , Primeiro Trimestre da Gravidez/metabolismo , Segundo Trimestre da Gravidez/genética , Segundo Trimestre da Gravidez/metabolismo , Análise de Sequência de DNA , Transcriptoma
13.
Cardiovasc Diabetol ; 11: 41, 2012 Apr 25.
Artigo em Inglês | MEDLINE | ID: mdl-22533685

RESUMO

BACKGROUND: Uncoupling protein 2 (UCP2) gene polymorphisms have been reported as genetic risk factors for obesity and type 2 diabetes mellitus (T2DM). We examined the association of commonly observed UCP2 G(-866)A (rs659366) and Ala55Val (C > T) (rs660339) single nucleotide polymorphisms (SNPs) with obesity, high fasting plasma glucose, and serum lipids in a Balinese population. METHODS: A total of 603 participants (278 urban and 325 rural subjects) were recruited from Bali Island, Indonesia. Fasting plasma glucose (FPG), triglyceride (TG), high density lipoprotein cholesterol (HDL-C), low density lipoprotein cholesterol (LDL-C) and total cholesterol (TC) were measured. Obesity was determined based on WHO classifications for adult Asians. Participants were genotyped for G(-866)A and Ala55Val polymorphisms of the UCP2 gene. RESULTS: Obesity prevalence was higher in urban subjects (51%) as compared to rural subjects (23%). The genotype, minor allele (MAF), and heterozygosity frequencies were similar between urban and rural subjects for both SNPs. All genotype frequencies were in Hardy-Weinberg equilibrium. A combined analysis of genotypes and environment revealed that the urban subjects carrying the A/A genotype of the G(-866)A SNP have higher BMI than the rural subjects with the same genotype. Since the two SNPs showed strong linkage disequilibrium (D' = 0.946, r2 = 0.657), a haplotype analysis was performed. We found that the AT haplotype was associated with high BMI only when the urban environment was taken into account. CONCLUSIONS: We have demonstrated the importance of environmental settings in studying the influence of the common UCP2 gene polymorphisms in the development of obesity in a Balinese population.


Assuntos
Canais Iônicos/genética , Proteínas Mitocondriais/genética , Obesidade/genética , Polimorfismo de Nucleotídeo Único , Adulto , Idoso , Grupo com Ancestrais do Continente Asiático/genética , Biomarcadores/sangue , Glicemia/análise , Estudos Transversais , Feminino , Frequência do Gene , Interação Gene-Ambiente , Predisposição Genética para Doença , Heterozigoto , Humanos , Indonésia/epidemiologia , Desequilíbrio de Ligação , Lipídeos/sangue , Modelos Logísticos , Masculino , Pessoa de Meia-Idade , Obesidade/sangue , Obesidade/etnologia , Fenótipo , Prevalência , Medição de Risco , Fatores de Risco , Saúde da População Rural/etnologia , Proteína Desacopladora 2 , Saúde da População Urbana/etnologia
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