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1.
G3 (Bethesda) ; 9(8): 2521-2533, 2019 08 08.
Artigo em Inglês | MEDLINE | ID: mdl-31186305

RESUMO

Identifying the regulatory mechanisms of genome-wide association study (GWAS) loci affecting adipose tissue has been restricted due to limited characterization of adipose transcriptional regulatory elements. We profiled chromatin accessibility in three frozen human subcutaneous adipose tissue needle biopsies and preadipocytes and adipocytes from the Simpson Golabi-Behmel Syndrome (SGBS) cell strain using an assay for transposase-accessible chromatin (ATAC-seq). We identified 68,571 representative accessible chromatin regions (peaks) across adipose tissue samples (FDR < 5%). GWAS loci for eight cardiometabolic traits were enriched in these peaks (P < 0.005), with the strongest enrichment for waist-hip ratio. Of 110 recently described cardiometabolic GWAS loci colocalized with adipose tissue eQTLs, 59 loci had one or more variants overlapping an adipose tissue peak. Annotated variants at the SNX10 waist-hip ratio locus and the ATP2A1-SH2B1 body mass index locus showed allelic differences in regulatory assays. These adipose tissue accessible chromatin regions elucidate genetic variants that may alter adipose tissue function to impact cardiometabolic traits.

2.
Hum Mol Genet ; 2018 Nov 16.
Artigo em Inglês | MEDLINE | ID: mdl-30445632

RESUMO

Total cholesterol (TC) and low-density lipoprotein cholesterol (LDL-C) are heritable risk factors for cardiovascular disease, yet the molecular mechanisms underlying the majority of blood lipid-associated GWAS signals remain elusive. One association signal is located in intron 3 of VLDLR; rs3780181-A is a risk allele associated (P≤2x10-9) with increased TC and LDL-C. We investigated variants, genes, and mechanisms underlying this association signal. We used a functional genetic approach to show that the intronic region spanning rs3780181 exhibited 1.6 to 7.6-fold enhancer activity in human HepG2 hepatocyte, THP-1 monocyte, and SGBS preadipocyte cells and that the rs3780181-A risk allele showed significantly less enhancer activity compared to the G-allele, consistent with the direction of an eQTL in liver. In addition, rs3780181 alleles showed differential binding to multiple nuclear proteins, including stronger IRF2 binding to the rs3780181 G-allele. We used a CRISPR-cas9 approach to delete 475 and 663 bp of the putative enhancer element in HEK293T kidney cells; compared to expression of mock-edited cell lines, the homozygous enhancer deletion cell lines showed 1.2-fold significantly (P<0.04) decreased expression of VLDLR, as well as 1.5-fold decreased expression of SMARCA2, located 388 kb away. Together, these results identify an enhancer of VLDLR expression and suggest that altered binding of one or more factors bound to rs3780181 alleles decreases enhancer activity and reduce at least VLDLR expression, leading to increased TC and LDL-C.

3.
Am J Hum Genet ; 102(4): 620-635, 2018 04 05.
Artigo em Inglês | MEDLINE | ID: mdl-29625024

RESUMO

Genome-wide association studies (GWASs) and functional genomics approaches implicate enhancer disruption in islet dysfunction and type 2 diabetes (T2D) risk. We applied genetic fine-mapping and functional (epi)genomic approaches to a T2D- and proinsulin-associated 15q22.2 locus to identify a most likely causal variant, determine its direction of effect, and elucidate plausible target genes. Fine-mapping and conditional analyses of proinsulin levels of 8,635 non-diabetic individuals from the METSIM study support a single association signal represented by a cluster of 16 strongly associated (p < 10-17) variants in high linkage disequilibrium (r2 > 0.8) with the GWAS index SNP rs7172432. These variants reside in an evolutionarily and functionally conserved islet and ß cell stretch or super enhancer; the most strongly associated variant (rs7163757, p = 3 × 10-19) overlaps a conserved islet open chromatin site. DNA sequence containing the rs7163757 risk allele displayed 2-fold higher enhancer activity than the non-risk allele in reporter assays (p < 0.01) and was differentially bound by ß cell nuclear extract proteins. Transcription factor NFAT specifically potentiated risk-allele enhancer activity and altered patterns of nuclear protein binding to the risk allele in vitro, suggesting that it could be a factor mediating risk-allele effects. Finally, the rs7163757 proinsulin-raising and T2D risk allele (C) was associated with increased expression of C2CD4B, and possibly C2CD4A, both of which were induced by inflammatory cytokines, in human islets. Together, these data suggest that rs7163757 contributes to genetic risk of islet dysfunction and T2D by increasing NFAT-mediated islet enhancer activity and modulating C2CD4B, and possibly C2CD4A, expression in (patho)physiologic states.

4.
PLoS Genet ; 14(4): e1007275, 2018 04.
Artigo em Inglês | MEDLINE | ID: mdl-29621232

RESUMO

To identify genetic contributions to type 2 diabetes (T2D) and related glycemic traits (fasting glucose, fasting insulin, and HbA1c), we conducted genome-wide association analyses (GWAS) in up to 7,178 Chinese subjects from nine provinces in the China Health and Nutrition Survey (CHNS). We examined patterns of population structure within CHNS and found that allele frequencies differed across provinces, consistent with genetic drift and population substructure. We further validated 32 previously described T2D- and glycemic trait-loci, including G6PC2 and SIX3-SIX2 associated with fasting glucose. At G6PC2, we replicated a known fasting glucose-associated variant (rs34177044) and identified a second signal (rs2232326), a low-frequency (4%), probably damaging missense variant (S324P). A variant within the lead fasting glucose-associated signal at SIX3-SIX2 co-localized with pancreatic islet expression quantitative trait loci (eQTL) for SIX3, SIX2, and three noncoding transcripts. To identify variants functionally responsible for the fasting glucose association at SIX3-SIX2, we tested five candidate variants for allelic differences in regulatory function. The rs12712928-C allele, associated with higher fasting glucose and lower transcript expression level, showed lower transcriptional activity in reporter assays and increased binding to GABP compared to the rs12712928-G, suggesting that rs12712928-C contributes to elevated fasting glucose levels by disrupting an islet enhancer, resulting in reduced gene expression. Taken together, these analyses identified multiple loci associated with glycemic traits across China, and suggest a regulatory mechanism at the SIX3-SIX2 fasting glucose GWAS locus.


Assuntos
Glicemia/genética , Diabetes Mellitus Tipo 2/genética , Inquéritos Epidemiológicos , China , Jejum , Feminino , Estudo de Associação Genômica Ampla , Humanos , Ilhotas Pancreáticas/metabolismo , Masculino , Mutação de Sentido Incorreto , Inquéritos Nutricionais , Locos de Características Quantitativas
5.
Diabetes ; 66(9): 2521-2530, 2017 09.
Artigo em Inglês | MEDLINE | ID: mdl-28684635

RESUMO

Molecular mechanisms remain unknown for most type 2 diabetes genome-wide association study identified loci. Variants associated with type 2 diabetes and fasting glucose levels reside in introns of ADCY5, a gene that encodes adenylate cyclase 5. Adenylate cyclase 5 catalyzes the production of cyclic AMP, which is a second messenger molecule involved in cell signaling and pancreatic ß-cell insulin secretion. We demonstrated that type 2 diabetes risk alleles are associated with decreased ADCY5 expression in human islets and examined candidate variants for regulatory function. rs11708067 overlaps a predicted enhancer region in pancreatic islets. The type 2 diabetes risk rs11708067-A allele showed fewer H3K27ac ChIP-seq reads in human islets, lower transcriptional activity in reporter assays in rodent ß-cells (rat 832/13 and mouse MIN6), and increased nuclear protein binding compared with the rs11708067-G allele. Homozygous deletion of the orthologous enhancer region in 832/13 cells resulted in a 64% reduction in expression level of Adcy5, but not adjacent gene Sec22a, and a 39% reduction in insulin secretion. Together, these data suggest that rs11708067-A risk allele contributes to type 2 diabetes by disrupting an islet enhancer, which results in reduced ADCY5 expression and impaired insulin secretion.


Assuntos
Adenilil Ciclases/metabolismo , Diabetes Mellitus Tipo 2/metabolismo , Regulação da Expressão Gênica/fisiologia , Variação Genética , Estudo de Associação Genômica Ampla , Ilhotas Pancreáticas/metabolismo , Adenilil Ciclases/genética , Diabetes Mellitus Tipo 2/genética , Humanos , Insulina/metabolismo
6.
Proc Natl Acad Sci U S A ; 114(9): 2301-2306, 2017 02 28.
Artigo em Inglês | MEDLINE | ID: mdl-28193859

RESUMO

Genome-wide association studies (GWAS) have identified >100 independent SNPs that modulate the risk of type 2 diabetes (T2D) and related traits. However, the pathogenic mechanisms of most of these SNPs remain elusive. Here, we examined genomic, epigenomic, and transcriptomic profiles in human pancreatic islets to understand the links between genetic variation, chromatin landscape, and gene expression in the context of T2D. We first integrated genome and transcriptome variation across 112 islet samples to produce dense cis-expression quantitative trait loci (cis-eQTL) maps. Additional integration with chromatin-state maps for islets and other diverse tissue types revealed that cis-eQTLs for islet-specific genes are specifically and significantly enriched in islet stretch enhancers. High-resolution chromatin accessibility profiling using assay for transposase-accessible chromatin sequencing (ATAC-seq) in two islet samples enabled us to identify specific transcription factor (TF) footprints embedded in active regulatory elements, which are highly enriched for islet cis-eQTL. Aggregate allelic bias signatures in TF footprints enabled us de novo to reconstruct TF binding affinities genetically, which support the high-quality nature of the TF footprint predictions. Interestingly, we found that T2D GWAS loci were strikingly and specifically enriched in islet Regulatory Factor X (RFX) footprints. Remarkably, within and across independent loci, T2D risk alleles that overlap with RFX footprints uniformly disrupt the RFX motifs at high-information content positions. Together, these results suggest that common regulatory variations have shaped islet TF footprints and the transcriptome and that a confluent RFX regulatory grammar plays a significant role in the genetic component of T2D predisposition.


Assuntos
Diabetes Mellitus Tipo 2/genética , Predisposição Genética para Doença , Genoma Humano , Ilhotas Pancreáticas/metabolismo , Locos de Características Quantitativas , Transcriptoma , Alelos , Sequência de Bases , Sítios de Ligação , Cromatina/química , Cromatina/metabolismo , Diabetes Mellitus Tipo 2/metabolismo , Diabetes Mellitus Tipo 2/patologia , Epigênese Genética , Perfilação da Expressão Gênica , Variação Genética , Estudo de Associação Genômica Ampla , Impressão Genômica , Humanos , Ilhotas Pancreáticas/patologia , Polimorfismo de Nucleotídeo Único , Ligação Proteica , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Fatores de Transcrição de Fator Regulador X/genética , Fatores de Transcrição de Fator Regulador X/metabolismo
7.
Nat Commun ; 7: 11764, 2016 06 29.
Artigo em Inglês | MEDLINE | ID: mdl-27353450

RESUMO

Type 2 diabetes (T2D) results from the combined effects of genetic and environmental factors on multiple tissues over time. Of the >100 variants associated with T2D and related traits in genome-wide association studies (GWAS), >90% occur in non-coding regions, suggesting a strong regulatory component to T2D risk. Here to understand how T2D status, metabolic traits and genetic variation influence gene expression, we analyse skeletal muscle biopsies from 271 well-phenotyped Finnish participants with glucose tolerance ranging from normal to newly diagnosed T2D. We perform high-depth strand-specific mRNA-sequencing and dense genotyping. Computational integration of these data with epigenome data, including ATAC-seq on skeletal muscle, and transcriptome data across diverse tissues reveals that the tissue-specific genetic regulatory architecture of skeletal muscle is highly enriched in muscle stretch/super enhancers, including some that overlap T2D GWAS variants. In one such example, T2D risk alleles residing in a muscle stretch/super enhancer are linked to increased expression and alternative splicing of muscle-specific isoforms of ANK1.


Assuntos
Diabetes Mellitus Tipo 2/genética , Diabetes Mellitus Tipo 2/metabolismo , Estudo de Associação Genômica Ampla , Músculo Esquelético/metabolismo , Alelos , Epigenômica , Feminino , Regulação da Expressão Gênica , Predisposição Genética para Doença , Humanos , Masculino , Polimorfismo de Nucleotídeo Único , RNA Mensageiro , Análise de Sequência de RNA
8.
Am J Hum Genet ; 98(2): 229-42, 2016 Feb 04.
Artigo em Inglês | MEDLINE | ID: mdl-26805783

RESUMO

Platelets play an essential role in hemostasis and thrombosis. We performed a genome-wide association study of platelet count in 12,491 participants of the Hispanic Community Health Study/Study of Latinos by using a mixed-model method that accounts for admixture and family relationships. We discovered and replicated associations with five genes (ACTN1, ETV7, GABBR1-MOG, MEF2C, and ZBTB9-BAK1). Our strongest association was with Amerindian-specific variant rs117672662 (p value = 1.16 × 10(-28)) in ACTN1, a gene implicated in congenital macrothrombocytopenia. rs117672662 exhibited allelic differences in transcriptional activity and protein binding in hematopoietic cells. Our results underscore the value of diverse populations to extend insights into the allelic architecture of complex traits.


Assuntos
Estudos de Associação Genética/métodos , Loci Gênicos , Hispano-Americanos/genética , Contagem de Plaquetas , Actinina/genética , Adolescente , Adulto , Idoso , Alelos , Frequência do Gene , Genótipo , Técnicas de Genotipagem , Humanos , Fatores de Transcrição MEF2/genética , Proteínas de Membrana/genética , Pessoa de Meia-Idade , Fenótipo , Polimorfismo de Nucleotídeo Único , Receptores de GABA-B/genética , Adulto Jovem
9.
Am J Hum Genet ; 97(6): 801-15, 2015 Dec 03.
Artigo em Inglês | MEDLINE | ID: mdl-26637976

RESUMO

Genome-wide association studies (GWASs) have identified more than 150 loci associated with blood lipid and cholesterol levels; however, the functional and molecular mechanisms for many associations are unknown. We examined the functional regulatory effects of candidate variants at the GALNT2 locus associated with high-density lipoprotein cholesterol (HDL-C). Fine-mapping and conditional analyses in the METSIM study identified a single locus harboring 25 noncoding variants (r(2) > 0.7 with the lead GWAS variants) strongly associated with total cholesterol in medium-sized HDL (e.g., rs17315646, p = 3.5 × 10(-12)). We used luciferase reporter assays in HepG2 cells to test all 25 variants for allelic differences in regulatory enhancer activity. rs2281721 showed allelic differences in transcriptional activity (75-fold [T] versus 27-fold [C] more than the empty-vector control), as did a separate 780-bp segment containing rs4846913, rs2144300, and rs6143660 (49-fold [AT(-) haplotype] versus 16-fold [CC(+) haplotype] more). Using electrophoretic mobility shift assays, we observed differential CEBPB binding to rs4846913, and we confirmed this binding in a native chromatin context by performing chromatin-immunoprecipitation (ChIP) assays in HepG2 and Huh-7 cell lines of differing genotypes. Additionally, sequence reads in HepG2 DNase-I-hypersensitivity and CEBPB ChIP-seq signals spanning rs4846913 showed significant allelic imbalance. Allelic-expression-imbalance assays performed with RNA from primary human hepatocyte samples and expression-quantitative-trait-locus (eQTL) data in human subcutaneous adipose tissue samples confirmed that alleles associated with increased HDL-C are associated with a modest increase in GALNT2 expression. Together, these data suggest that at least rs4846913 and rs2281721 play key roles in influencing GALNT2 expression at this HDL-C locus.


Assuntos
Proteína beta Intensificadora de Ligação a CCAAT/genética , HDL-Colesterol/genética , Genoma Humano , N-Acetilgalactosaminiltransferases/genética , Locos de Características Quantitativas , Tecido Adiposo/citologia , Tecido Adiposo/metabolismo , Alelos , Proteína beta Intensificadora de Ligação a CCAAT/metabolismo , HDL-Colesterol/metabolismo , Cromatina/química , Cromatina/metabolismo , Imunoprecipitação da Cromatina , Mapeamento Cromossômico , Ensaio de Desvio de Mobilidade Eletroforética , Frequência do Gene , Genes Reporter , Estudo de Associação Genômica Ampla , Haplótipos , Células Hep G2 , Humanos , Luciferases/genética , Luciferases/metabolismo , N-Acetilgalactosaminiltransferases/metabolismo , Cultura Primária de Células , Ligação Proteica
10.
PLoS Genet ; 10(9): e1004633, 2014 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-25211022

RESUMO

Many of the type 2 diabetes loci identified through genome-wide association studies localize to non-protein-coding intronic and intergenic regions and likely contain variants that regulate gene transcription. The CDC123/CAMK1D type 2 diabetes association signal on chromosome 10 spans an intergenic region between CDC123 and CAMK1D and also overlaps the CDC123 3'UTR. To gain insight into the molecular mechanisms underlying the association signal, we used open chromatin, histone modifications and transcription factor ChIP-seq data sets from type 2 diabetes-relevant cell types to identify SNPs overlapping predicted regulatory regions. Two regions containing type 2 diabetes-associated variants were tested for enhancer activity using luciferase reporter assays. One SNP, rs11257655, displayed allelic differences in transcriptional enhancer activity in 832/13 and MIN6 insulinoma cells as well as in human HepG2 hepatocellular carcinoma cells. The rs11257655 risk allele T showed greater transcriptional activity than the non-risk allele C in all cell types tested. Using electromobility shift and supershift assays we demonstrated that the rs11257655 risk allele showed allele-specific binding to FOXA1 and FOXA2. We validated FOXA1 and FOXA2 enrichment at the rs11257655 risk allele using allele-specific ChIP in human islets. These results suggest that rs11257655 affects transcriptional activity through altered binding of a protein complex that includes FOXA1 and FOXA2, providing a potential molecular mechanism at this GWAS locus.


Assuntos
Proteína Quinase Tipo 1 Dependente de Cálcio-Calmodulina/genética , Proteínas de Ciclo Celular/genética , Diabetes Mellitus Tipo 2/genética , Diabetes Mellitus Tipo 2/metabolismo , Variação Genética , Fator 3-alfa Nuclear de Hepatócito/metabolismo , Fator 3-beta Nuclear de Hepatócito/metabolismo , Sequências Reguladoras de Ácido Nucleico , Alelos , Animais , Linhagem Celular , Elementos Facilitadores Genéticos , Regulação da Expressão Gênica , Loci Gênicos , Estudo de Associação Genômica Ampla , Hepatócitos/metabolismo , Humanos , Ilhotas Pancreáticas/metabolismo , Camundongos , Polimorfismo de Nucleotídeo Único , Ligação Proteica , Transcrição Genética
11.
Genomics ; 104(2): 105-12, 2014 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-24997396

RESUMO

Characterization of the epigenome promises to yield the functional elements buried in the human genome sequence, thus helping to annotate non-coding DNA polymorphisms with regulatory functions. Here, we develop two novel strategies to combine epigenomic data with transcriptomic profiles in humans or mice to prioritize potential candidate SNPs associated with lipid levels by genome-wide association study (GWAS). First, after confirming that lipid-associated loci that are also expression quantitative trait loci (eQTL) in human livers are enriched for ENCODE regulatory marks in the human hepatocellular HepG2 cell line, we prioritize candidate SNPs based on the number of these marks that overlap the variant position. This method recognized the known SORT1 rs12740374 regulatory SNP associated with LDL-cholesterol, and highlighted candidate functional SNPs at 15 additional lipid loci. In the second strategy, we combine ENCODE chromatin immunoprecipitation followed by high-throughput DNA sequencing (ChIP-seq) data and liver expression datasets from knockout mice lacking specific transcription factors. This approach identified SNPs in specific transcription factor binding sites that are located near target genes of these transcription factors. We show that FOXA2 transcription factor binding sites are enriched at lipid-associated loci and experimentally validate that alleles of one such proxy SNP located near the FOXA2 target gene BIRC5 show allelic differences in FOXA2-DNA binding and enhancer activity. These methods can be used to generate testable hypotheses for many non-coding SNPs associated with complex diseases or traits.


Assuntos
Epigenômica/métodos , Estudos de Associação Genética/métodos , Lipídeos/química , Fígado/metabolismo , Fatores de Transcrição/genética , Alelos , Animais , Sítios de Ligação , Imunoprecipitação da Cromatina/métodos , Mapeamento Cromossômico , Genoma Humano , Células Hep G2 , Fator 3-beta Nuclear de Hepatócito/genética , Fator 3-beta Nuclear de Hepatócito/metabolismo , Fator 4 Nuclear de Hepatócito/genética , Fator 4 Nuclear de Hepatócito/metabolismo , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Humanos , Células K562 , Camundongos , Camundongos Knockout , Fenótipo , Polimorfismo de Nucleotídeo Único , Ligação Proteica/genética , Locos de Características Quantitativas , Fatores de Transcrição/metabolismo , Transcriptoma/genética
12.
Hum Mol Genet ; 23(4): 1108-19, 2014 Feb 15.
Artigo em Inglês | MEDLINE | ID: mdl-24105470

RESUMO

Blood levels of adiponectin, an adipocyte-secreted protein correlated with metabolic and cardiovascular risks, are highly heritable. Genome-wide association (GWA) studies for adiponectin levels have identified 14 loci harboring variants associated with blood levels of adiponectin. To identify novel adiponectin-associated loci, particularly those of importance in East Asians, we conducted a meta-analysis of GWA studies for adiponectin in 7827 individuals, followed by two stages of replications in 4298 and 5954 additional individuals. We identified a novel adiponectin-associated locus on chromosome 10 near WDR11-FGFR2 (P = 3.0 × 10(-14)) and provided suggestive evidence for a locus on chromosome 12 near OR8S1-LALBA (P = 1.2 × 10(-7)). Of the adiponectin-associated loci previously described, we confirmed the association at CDH13 (P = 6.8 × 10(-165)), ADIPOQ (P = 1.8 × 10(-22)), PEPD (P = 3.6 × 10(-12)), CMIP (P = 2.1 × 10(-10)), ZNF664 (P = 2.3 × 10(-7)) and GPR109A (P = 7.4 × 10(-6)). Conditional analysis at ADIPOQ revealed a second signal with suggestive evidence of association only after conditioning on the lead SNP (Pinitial = 0.020; Pconditional = 7.0 × 10(-7)). We further confirmed the independence of two pairs of closely located loci (<2 Mb) on chromosome 16 at CMIP and CDH13, and on chromosome 12 at GPR109A and ZNF664. In addition, the newly identified signal near WDR11-FGFR2 exhibited evidence of association with triglycerides (P = 3.3 × 10(-4)), high density lipoprotein cholesterol (HDL-C, P = 4.9 × 10(-4)) and body mass index (BMI)-adjusted waist-hip ratio (P = 9.8 × 10(-3)). These findings improve our knowledge of the genetic basis of adiponectin variation, demonstrate the shared allelic architecture for adiponectin with lipids and central obesity and motivate further studies of underlying mechanisms.


Assuntos
Adiponectina/sangue , Doenças Cardiovasculares/genética , Proteínas de Membrana/genética , Proteínas Proto-Oncogênicas/genética , Receptor Tipo 2 de Fator de Crescimento de Fibroblastos/genética , Grupo com Ancestrais do Continente Asiático , Doenças Cardiovasculares/sangue , Estudos de Coortes , Loci Gênicos , Predisposição Genética para Doença , Estudo de Associação Genômica Ampla , Humanos , Polimorfismo de Nucleotídeo Único
13.
Diabetes ; 62(5): 1756-62, 2013 May.
Artigo em Inglês | MEDLINE | ID: mdl-23328127

RESUMO

Translation of noncoding common variant association signals into meaningful molecular and biological mechanisms explaining disease susceptibility remains challenging. For the type 2 diabetes association signal in JAZF1 intron 1, we hypothesized that the underlying risk variants have cis-regulatory effects in islets or other type 2 diabetes-relevant cell types. We used maps of experimentally predicted open chromatin regions to prioritize variants for functional follow-up studies of transcriptional activity. Twelve regions containing type 2 diabetes-associated variants were tested for enhancer activity in 832/13 and MIN6 insulinoma cells. Three regions exhibited enhancer activity and only rs1635852 displayed allelic differences in enhancer activity; the type 2 diabetes risk allele T showed lower transcriptional activity than the nonrisk allele C. This risk allele showed increased binding to protein complexes, suggesting that it functions as part of a transcriptional repressor complex. We applied DNA affinity capture to identify factors in the complex and determined that the risk allele preferentially binds the pancreatic master regulator PDX1. These data suggest that the rs1635852 region in JAZF1 intron 1 is part of a cis-regulatory complex and that maps of open chromatin are useful to guide identification of variants with allelic differences in regulatory activity at type 2 diabetes loci.


Assuntos
Cromatina/metabolismo , Diabetes Mellitus Tipo 2/genética , Ilhotas Pancreáticas/metabolismo , Proteínas de Neoplasias/genética , Polimorfismo de Nucleotídeo Único , Transcrição Genética , Alelos , Animais , Linhagem Celular , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Diabetes Mellitus Tipo 2/metabolismo , Genes Reporter , Estudos de Associação Genética , Proteínas de Homeodomínio/metabolismo , Humanos , Íntrons , Desequilíbrio de Ligação , Camundongos , Proteínas de Neoplasias/metabolismo , Ratos , Proteínas Recombinantes/metabolismo , Transativadores/metabolismo , Utah
14.
Cereb Cortex ; 18(11): 2560-73, 2008 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-18296432

RESUMO

Subunit composition of N-methyl-D-aspartate-type glutamate receptors (NMDARs) dictates their function, yet the ontogenic profiles of human NMDAR subunits from gestation to adulthood have not been determined. We examined NMDAR mRNA and protein development in human dorsolateral prefrontal cortex (DLPFC), an area in which NMDARs are critical for higher cognitive processing and NMDAR hypofunction is hypothesized in schizophrenia. Using quantitative reverse transcriptase-polymerase chain reaction and western blotting, we found NR1 expression begins low prenatally, peaks in adolescence, yet remains high throughout life, suggesting lifelong importance of NMDAR function. In contrast, NR3A levels are low during gestation, surge soon after birth, and decline progressively through adolescence and into adulthood. Because NR3A subunits uniquely attenuate NMDAR-mediated currents, limit calcium influx, and suppress dendritic spine formation, high levels during early childhood may be important for regulating neuroprotection and activity-dependent sculpting of synapses. We also examined whether subunit changes underlie reduced NMDAR activity in schizophrenia. Our results reveal normal NR1 and NR3A protein levels in DLPFC from schizophrenic patients, indicating that NMDAR hypofunction is unlikely to be maintained by gross changes in NR3A-containing NMDARs or overall NMDAR numbers. These data provide insights into NMDAR functions in the developing CNS and will contribute to designing pharmacotherapies for neurological disorders.


Assuntos
Regulação da Expressão Gênica no Desenvolvimento , Córtex Pré-Frontal/crescimento & desenvolvimento , Córtex Pré-Frontal/fisiologia , Receptores de N-Metil-D-Aspartato/genética , Esquizofrenia/fisiopatologia , Adolescente , Adulto , Idoso , Animais , Antipsicóticos/farmacologia , Antipsicóticos/uso terapêutico , Criança , Pré-Escolar , Feto , Humanos , Lactente , Recém-Nascido , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Pessoa de Meia-Idade , Córtex Pré-Frontal/embriologia , Ratos , Ratos Sprague-Dawley , Receptores de N-Metil-D-Aspartato/metabolismo , Esquizofrenia/tratamento farmacológico , Adulto Jovem
15.
J Neuroimmunol ; 159(1-2): 106-12, 2005 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-15652408

RESUMO

Maternal infection during pregnancy is associated with increased risk for neurodevelopmental disorders. Polyriboinosinic-polyribocytidilic acid (poly I:C) or saline was administered to rats to model maternal infection; levels of TNFalpha, brain-derived neurotrophic factor (BDNF), and nerve growth factor (NGF) were determined by ELISA. TNFalpha was significantly increased in maternal plasma, placenta, and amniotic fluid, while it was significantly decreased in fetal liver/spleen and neonatal brain. NGF and BDNF were significantly decreased in the placenta and fetal liver/spleen. There was no change in BDNF or NGF in the fetal or neonatal brain. Changes in TNFalpha, BDNF, and NGF after maternal exposure to poly I:C represent a potential mechanism through which maternal infection increases risk for neurodevelopmental disorders.


Assuntos
Fator Neurotrófico Derivado do Encéfalo/biossíntese , Exposição Materna , Troca Materno-Fetal/efeitos dos fármacos , Fator de Crescimento Neural/biossíntese , Poli I-C/farmacologia , Efeitos Tardios da Exposição Pré-Natal , Fator de Necrose Tumoral alfa/biossíntese , Líquido Amniótico/imunologia , Líquido Amniótico/metabolismo , Animais , Animais Recém-Nascidos , Encéfalo/efeitos dos fármacos , Encéfalo/metabolismo , Fator Neurotrófico Derivado do Encéfalo/antagonistas & inibidores , Fator Neurotrófico Derivado do Encéfalo/sangue , Regulação para Baixo/efeitos dos fármacos , Regulação para Baixo/imunologia , Feminino , Injeções Intraperitoneais , Fígado/efeitos dos fármacos , Fígado/embriologia , Fígado/metabolismo , Troca Materno-Fetal/imunologia , Fator de Crescimento Neural/antagonistas & inibidores , Fator de Crescimento Neural/sangue , Poli I-C/administração & dosagem , Gravidez , Proteínas da Gravidez/antagonistas & inibidores , Proteínas da Gravidez/biossíntese , Ratos , Ratos Sprague-Dawley , Baço/efeitos dos fármacos , Baço/embriologia , Baço/metabolismo , Fator de Necrose Tumoral alfa/antagonistas & inibidores , Fator de Necrose Tumoral alfa/fisiologia , Regulação para Cima/efeitos dos fármacos , Regulação para Cima/imunologia
16.
Neuropsychopharmacology ; 29(7): 1221-9, 2004 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-15085088

RESUMO

Prenatal exposure to infection increases risk for schizophrenia, and we have hypothesized that inflammatory cytokines, generated in response to maternal infection, alter neuron development and increase risk for schizophrenia. We sought to study the effect of cytokines generated in response to infection-interleukin-1beta (IL-1beta), tumor necrosis factor-alpha (TNFalpha), and interleukin-6 (IL-6)-on the dendritic development of cortical neurons. Primary mixed neuronal cultures were obtained from E18 rats and exposed to 0, 100, or 1000 units (U)/ml of IL-1beta, TNFalpha, IL-6, or IL-1beta+TNFalpha for 44 h. MAP-2-positive neurons were randomly identified for each condition and the number of primary dendrites, nodes, and total dendrite length was determined. We found that 100 U of TNFalpha significantly reduced the number of nodes (27%, p=0.02) and total dendritic length (14%, p=0.04), but did not affect overall neuron survival. A measure of 100 U IL-1beta+TNFalpha significantly reduced the number of primary dendrites (17%, p=0.006), nodes (32%, p=0.001), and total dendritic length (30%, p<0.0001), although it did not affect overall neuron survival. At 1000 U, each cytokine significantly reduced the number of primary dendrites (14-24%), nodes (28-37%), as well as total dendritic length (25-30%); neuron survival was reduced by 14-21%. These results indicate that inflammatory cytokines can significantly reduce dendrite development and complexity of developing cortical neurons, consistent with the neuropathology of schizophrenia. These findings also support the hypothesis that cytokines play a key mechanistic role in the link between prenatal exposure to infection and risk for schizophrenia.


Assuntos
Interleucina-1beta/fisiologia , Interleucina-6/fisiologia , Efeitos Tardios da Exposição Pré-Natal , Esquizofrenia/etiologia , Fator de Necrose Tumoral alfa/fisiologia , Animais , Proliferação de Células , Córtex Cerebral/citologia , Córtex Cerebral/crescimento & desenvolvimento , Dendritos , Feminino , Infecção/complicações , Gravidez , Ratos , Ratos Sprague-Dawley , Esquizofrenia/imunologia
17.
J Neuroimmunol ; 138(1-2): 49-55, 2003 May.
Artigo em Inglês | MEDLINE | ID: mdl-12742653

RESUMO

Maternal infection during pregnancy is associated with increased risk for neurodevelopmental disorders. Lipopolysaccharide (LPS) or saline was administered to rats to model maternal infection, and levels of brain-derived neurotrophic factor (BDNF) and nerve growth factor (NGF) in maternal plasma, placenta, amniotic fluid, fetal liver/spleen, fetal brain, and cerebral cortex after birth were determined by ELISA or semiquantitative Western blot analysis. BDNF expression was significantly increased in the fetal brain (p=0.039); NGF expression was significantly increased in neonatal cortex (p=0.0009). Neurotrophic factor expression was also altered in other tissues of the maternal-fetal unit. Abnormal expression of neurotrophic factors represents a potential mechanism through which maternal infection increases risk for neurodevelopmental disorders.


Assuntos
Fator Neurotrófico Derivado do Encéfalo/biossíntese , Encéfalo/metabolismo , Infecções por Escherichia coli/metabolismo , Troca Materno-Fetal/imunologia , Fator de Crescimento Neural/biossíntese , Complicações Infecciosas na Gravidez/metabolismo , Animais , Animais Recém-Nascidos , Encéfalo/embriologia , Encéfalo/imunologia , Fator Neurotrófico Derivado do Encéfalo/antagonistas & inibidores , Fator Neurotrófico Derivado do Encéfalo/sangue , Infecções por Escherichia coli/sangue , Infecções por Escherichia coli/imunologia , Feminino , Feto , Injeções Intraperitoneais , Lipopolissacarídeos/toxicidade , Fígado/embriologia , Fígado/imunologia , Fígado/metabolismo , Masculino , Placenta/imunologia , Placenta/metabolismo , Gravidez , Complicações Infecciosas na Gravidez/sangue , Complicações Infecciosas na Gravidez/imunologia , Ratos , Ratos Sprague-Dawley , Baço/embriologia , Baço/imunologia , Baço/metabolismo , Fator de Necrose Tumoral alfa/biossíntese , Regulação para Cima/imunologia
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