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1.
PLoS Genet ; 8(3): e1002584, 2012.
Artigo em Inglês | MEDLINE | ID: mdl-22479191

RESUMO

Chronic kidney disease (CKD) is an important public health problem with a genetic component. We performed genome-wide association studies in up to 130,600 European ancestry participants overall, and stratified for key CKD risk factors. We uncovered 6 new loci in association with estimated glomerular filtration rate (eGFR), the primary clinical measure of CKD, in or near MPPED2, DDX1, SLC47A1, CDK12, CASP9, and INO80. Morpholino knockdown of mpped2 and casp9 in zebrafish embryos revealed podocyte and tubular abnormalities with altered dextran clearance, suggesting a role for these genes in renal function. By providing new insights into genes that regulate renal function, these results could further our understanding of the pathogenesis of CKD.


Assuntos
Estudo de Associação Genômica Ampla , Taxa de Filtração Glomerular/genética , Falência Renal Crônica/genética , Rim/fisiopatologia , Peixe-Zebra/genética , ATPases Associadas a Diversas Atividades Celulares , Negro ou Afro-Americano/genética , Idoso , Animais , Caspase 9/genética , Quinases Ciclina-Dependentes/genética , RNA Helicases DEAD-box/genética , DNA Helicases/genética , Proteínas de Ligação a DNA , Feminino , Seguimentos , Técnicas de Silenciamento de Genes , Humanos , Falência Renal Crônica/patologia , Masculino , Pessoa de Meia-Idade , Diester Fosfórico Hidrolases/genética , População Branca/genética
2.
BMC Genomics ; 15: 532, 2014 Jun 27.
Artigo em Inglês | MEDLINE | ID: mdl-24973796

RESUMO

BACKGROUND: Gene expression genetic studies in human tissues and cells identify cis- and trans-acting expression quantitative trait loci (eQTLs). These eQTLs provide insights into regulatory mechanisms underlying disease risk. However, few studies systematically characterized eQTL results across cell and tissues types. We synthesized eQTL results from >50 datasets, including new primary data from human brain, peripheral plaque and kidney samples, in order to discover features of human eQTLs. RESULTS: We find a substantial number of robust cis-eQTLs and far fewer trans-eQTLs consistent across tissues. Analysis of 45 full human GWAS scans indicates eQTLs are enriched overall, and above nSNPs, among positive statistical signals in genetic mapping studies, and account for a significant fraction of the strongest human trait effects. Expression QTLs are enriched for gene centricity, higher population allele frequencies, in housekeeping genes, and for coincidence with regulatory features, though there is little evidence of 5' or 3' positional bias. Several regulatory categories are not enriched including microRNAs and their predicted binding sites and long, intergenic non-coding RNAs. Among the most tissue-ubiquitous cis-eQTLs, there is enrichment for genes involved in xenobiotic metabolism and mitochondrial function, suggesting these eQTLs may have adaptive origins. Several strong eQTLs (CDK5RAP2, NBPFs) coincide with regions of reported human lineage selection. The intersection of new kidney and plaque eQTLs with related GWAS suggest possible gene prioritization. For example, butyrophilins are now linked to arterial pathogenesis via multiple genetic and expression studies. Expression QTL and GWAS results are made available as a community resource through the NHLBI GRASP database [http://apps.nhlbi.nih.gov/grasp/]. CONCLUSIONS: Expression QTLs inform the interpretation of human trait variability, and may account for a greater fraction of phenotypic variability than protein-coding variants. The synthesis of available tissue eQTL data highlights many strong cis-eQTLs that may have important biologic roles and could serve as positive controls in future studies. Our results indicate some strong tissue-ubiquitous eQTLs may have adaptive origins in humans. Efforts to expand the genetic, splicing and tissue coverage of known eQTLs will provide further insights into human gene regulation.


Assuntos
Locos de Características Quantitativas , Linhagem Celular , Análise por Conglomerados , Perfilação da Expressão Gênica , Frequência do Gene , Estudo de Associação Genômica Ampla , Humanos , Especificidade de Órgãos , Polimorfismo de Nucleotídeo Único , Transcriptoma
3.
Hum Mol Genet ; 21(24): 5329-43, 2012 Dec 15.
Artigo em Inglês | MEDLINE | ID: mdl-22962313

RESUMO

In conducting genome-wide association studies (GWAS), analytical approaches leveraging biological information may further understanding of the pathophysiology of clinical traits. To discover novel associations with estimated glomerular filtration rate (eGFR), a measure of kidney function, we developed a strategy for integrating prior biological knowledge into the existing GWAS data for eGFR from the CKDGen Consortium. Our strategy focuses on single nucleotide polymorphism (SNPs) in genes that are connected by functional evidence, determined by literature mining and gene ontology (GO) hierarchies, to genes near previously validated eGFR associations. It then requires association thresholds consistent with multiple testing, and finally evaluates novel candidates by independent replication. Among the samples of European ancestry, we identified a genome-wide significant SNP in FBXL20 (P = 5.6 × 10(-9)) in meta-analysis of all available data, and additional SNPs at the INHBC, LRP2, PLEKHA1, SLC3A2 and SLC7A6 genes meeting multiple-testing corrected significance for replication and overall P-values of 4.5 × 10(-4)-2.2 × 10(-7). Neither the novel PLEKHA1 nor FBXL20 associations, both further supported by association with eGFR among African Americans and with transcript abundance, would have been implicated by eGFR candidate gene approaches. LRP2, encoding the megalin receptor, was identified through connection with the previously known eGFR gene DAB2 and extends understanding of the megalin system in kidney function. These findings highlight integration of existing genome-wide association data with independent biological knowledge to uncover novel candidate eGFR associations, including candidates lacking known connections to kidney-specific pathways. The strategy may also be applicable to other clinical phenotypes, although more testing will be needed to assess its potential for discovery in general.


Assuntos
Estudo de Associação Genômica Ampla/métodos , Polimorfismo de Nucleotídeo Único/genética , Sistemas de Transporte de Aminoácidos Básicos/genética , Cadeia Pesada da Proteína-1 Reguladora de Fusão/genética , Predisposição Genética para Doença/genética , Taxa de Filtração Glomerular/genética , Taxa de Filtração Glomerular/fisiologia , Humanos , Subunidades beta de Inibinas/genética , Peptídeos e Proteínas de Sinalização Intracelular/genética , Proteína-2 Relacionada a Receptor de Lipoproteína de Baixa Densidade/genética , Proteínas de Membrana/genética
4.
JCO Oncol Pract ; 20(1): 145-153, 2024 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-37556776

RESUMO

PURPOSE: Identification and targeting of actionable oncogenic drivers (AODs) in advanced non-small-cell lung cancer (NSCLC) has dramatically improved outcomes. However, genomic testing uptake is variable and hampered by factors including slow turnaround time, frequently resulting in initial non-tyrosine kinase inhibitor (TKI) treatment. We investigate how this behavior affects outcomes. METHODS: This retrospective analysis of real-world, deidentified data from the Integra Connect Database included adults with stage IV NSCLC newly diagnosed from January 1, 2018, to December 31, 2020, with mutations of EGFR, ALK, ROS1, BRAF, MET, RET, ERBB2, or NTRK. Outcomes were reported as time to next treatment or death (TTNT) and overall survival (OS). RESULTS: Five hundred ten patients harboring AODs were identified and grouped as follows: group A (n = 379) were treated after the AOD was reported and served as the comparator. One hundred thirty-one patients treated before their AOD report were divided into group B (n = 47) who were initially started on chemotherapy and/or checkpoint inhibitor but switched to appropriate TKI within 35 days and group C (n = 84) who were also started empirically on non-TKI and did not switch within 35 days. Survival (OS) was significantly superior in group A compared with group C; TTNT was significantly superior in group A compared with groups B and C. CONCLUSION: For patients harboring AODs in advanced NSCLC, initial treatment before receipt of genomic test results yields significantly inferior outcomes and should be avoided. Molecular profiling panels with rapid turnaround times are essential to optimize patient outcomes and should be standard of care.


Assuntos
Carcinoma Pulmonar de Células não Pequenas , Neoplasias Pulmonares , Adulto , Humanos , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Carcinoma Pulmonar de Células não Pequenas/genética , Neoplasias Pulmonares/tratamento farmacológico , Neoplasias Pulmonares/genética , Estudos Retrospectivos , Proteínas Tirosina Quinases/genética , Proteínas Proto-Oncogênicas/genética , Mutação
5.
J Cell Biochem ; 113(11): 3313-29, 2012 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-22644811

RESUMO

Although it is well known that chromosomes are non-randomly organized during interphase, it is not completely clear whether higher-order chromatin structure is transmitted from mother to daughter cells. Therefore, we addressed the question of how chromatin is rearranged during interphase and whether heterochromatin pattern is transmitted after mitosis. We additionally tested the similarity of chromatin arrangement in sister interphase nuclei. We noticed a very active cell rotation during interphase, especially when histone hyperacetylation was induced or transcription was inhibited. This natural phenomenon can influence the analysis of nuclear arrangement. Using photoconversion of Dendra2-tagged core histone H4 we showed that the distribution of chromatin in daughter interphase nuclei differed from that in mother cells. Similarly, the nuclear distribution of heterochromatin protein 1ß (HP1ß) was not completely identical in mother and daughter cells. However, identity between mother and daughter cells was in many cases evidenced by nucleolar composition. Moreover, morphology of nucleoli, HP1ß protein, Cajal bodies, chromosome territories, and gene transcripts were identical in sister cell nuclei. We conclude that the arrangement of interphase chromatin is not transmitted through mitosis, but the nuclear pattern is identical in naturally synchronized sister cells. It is also necessary to take into account the possibility that cell rotation and the degree of chromatin condensation during functionally specific cell cycle phases might influence our view of nuclear architecture.


Assuntos
Nucléolo Celular/ultraestrutura , Corpos Enovelados/ultraestrutura , Heterocromatina/genética , Interfase/genética , Mitose/genética , Animais , Linhagem Celular , Nucléolo Celular/efeitos dos fármacos , Nucléolo Celular/genética , Homólogo 5 da Proteína Cromobox , Proteínas Cromossômicas não Histona/genética , Proteínas Cromossômicas não Histona/metabolismo , Corpos Enovelados/efeitos dos fármacos , Corpos Enovelados/genética , Dactinomicina/farmacologia , Corantes Fluorescentes , Heterocromatina/efeitos dos fármacos , Heterocromatina/ultraestrutura , Inibidores de Histona Desacetilases/farmacologia , Histonas/genética , Histonas/metabolismo , Humanos , Ácidos Hidroxâmicos/farmacologia , Interfase/efeitos dos fármacos , Camundongos , Microscopia de Fluorescência , Mitose/efeitos dos fármacos , Processos Fotoquímicos , Inibidores da Síntese de Proteínas/farmacologia , RNA Mensageiro/biossíntese
6.
Proc Natl Acad Sci U S A ; 106(10): 3812-7, 2009 Mar 10.
Artigo em Inglês | MEDLINE | ID: mdl-19234129

RESUMO

Genome function in higher eukaryotes involves major changes in the spatial organization of the chromatin fiber. Nevertheless, our understanding of chromatin folding is remarkably limited. Polymer models have been used to describe chromatin folding. However, none of the proposed models gives a satisfactory explanation of experimental data. In particularly, they ignore that each chromosome occupies a confined space, i.e., the chromosome territory. Here, we present a polymer model that is able to describe key properties of chromatin over length scales ranging from 0.5 to 75 Mb. This random loop (RL) model assumes a self-avoiding random walk folding of the polymer backbone and defines a probability P for 2 monomers to interact, creating loops of a broad size range. Model predictions are compared with systematic measurements of chromatin folding of the q-arms of chromosomes 1 and 11. The RL model can explain our observed data and suggests that on the tens-of-megabases length scale P is small, i.e., 10-30 loops per 100 Mb. This is sufficient to enforce folding inside the confined space of a chromosome territory. On the 0.5- to 3-Mb length scale chromatin compaction differs in different subchromosomal domains. This aspect of chromatin structure is incorporated in the RL model by introducing heterogeneity along the fiber contour length due to different local looping probabilities. The RL model creates a quantitative and predictive framework for the identification of nuclear components that are responsible for chromatin-chromatin interactions and determine the 3-dimensional organization of the chromatin fiber.


Assuntos
Cromatina/química , Fibroblastos/citologia , Interfase , Conformação de Ácido Nucleico , Células Cultivadas , Feminino , Humanos , Modelos Moleculares
7.
Mol Cell Biol ; 27(12): 4475-87, 2007 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-17420274

RESUMO

The three-dimensional (3D) organization of the chromosomal fiber in the human interphase nucleus is an important but poorly understood aspect of gene regulation. Here we quantitatively analyze and compare the 3D structures of two types of genomic domains as defined by the human transcriptome map. While ridges are gene dense and show high expression levels, antiridges, on the other hand, are gene poor and carry genes that are expressed at low levels. We show that ridges are in general less condensed, more irregularly shaped, and located more closely to the nuclear center than antiridges. Six human cell lines that display different gene expression patterns and karyotypes share these structural parameters of chromatin. This shows that the chromatin structures of these two types of genomic domains are largely independent of tissue-specific variations in gene expression and differentiation state. Moreover, we show that there is remarkably little intermingling of chromatin from different parts of the same chromosome in a chromosome territory, neither from adjacent nor from distant parts. This suggests that the chromosomal fiber has a compact structure that sterically suppresses intermingling. Together, our results reveal novel general aspects of 3D chromosome architecture that are related to genome structure and function.


Assuntos
Cromossomos Humanos , Genoma Humano , Interfase , Mapeamento Físico do Cromossomo , Transcrição Gênica , Linhagem Celular , Linhagem Celular Tumoral , Cromatina/genética , Células HeLa , Humanos , Processamento de Imagem Assistida por Computador , Hibridização in Situ Fluorescente , Modelos Genéticos , Análise de Sequência com Séries de Oligonucleotídeos
8.
Eur J Cancer ; 75: 63-72, 2017 04.
Artigo em Inglês | MEDLINE | ID: mdl-28214660

RESUMO

Neuroblastoma is predominantly characterised by chromosomal rearrangements. Next to V-Myc Avian Myelocytomatosis Viral Oncogene Neuroblastoma Derived Homolog (MYCN) amplification, chromosome 7 and 17q gains are frequently observed. We identified a neuroblastoma patient with a regional 7q36 gain, encompassing the enhancer of zeste homologue 2 (EZH2) gene. EZH2 is the histone methyltransferase of lysine 27 of histone H3 (H3K27me3) that forms the catalytic subunit of the polycomb repressive complex 2. H3K27me3 is commonly associated with the silencing of genes involved in cellular processes such as cell cycle regulation, cellular differentiation and cancer. High EZH2 expression correlated with poor prognosis and overall survival independent of MYCN amplification status. Unexpectedly, treatment of 3 EZH2-high expressing neuroblastoma cell lines (IMR32, CHP134 and NMB), with EZH2-specific inhibitors (GSK126 and EPZ6438) resulted in only a slight G1 arrest, despite maximum histone methyltransferase activity inhibition. Furthermore, colony formation in cell lines treated with the inhibitors was reduced only at concentrations much higher than necessary for complete inhibition of EZH2 histone methyltransferase activity. Knockdown of the complete protein with three independent shRNAs resulted in a strong apoptotic response and decreased cyclin D1 levels. This apoptotic response could be rescued by overexpressing EZH2ΔSET, a truncated form of wild-type EZH2 lacking the SET transactivation domain necessary for histone methyltransferase activity. Our findings suggest that high EZH2 expression, at least in neuroblastoma, has a survival function independent of its methyltransferase activity. This important finding highlights the need for studies on EZH2 beyond its methyltransferase function and the requirement for compounds that will target EZH2 as a complete protein.


Assuntos
Proteína Potenciadora do Homólogo 2 de Zeste/fisiologia , Histona-Lisina N-Metiltransferase/antagonistas & inibidores , Neuroblastoma/enzimologia , Benzamidas/farmacologia , Compostos de Bifenilo , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Sobrevivência Celular/fisiologia , Aberrações Cromossômicas , Cromossomos Humanos Par 7 , Regulação para Baixo/efeitos dos fármacos , Proteína Potenciadora do Homólogo 2 de Zeste/metabolismo , Inibidores Enzimáticos/farmacologia , Técnicas de Silenciamento de Genes , Rearranjo Gênico , Histona Metiltransferases , Histona-Lisina N-Metiltransferase/fisiologia , Humanos , Indóis/farmacologia , Morfolinas , Neuroblastoma/fisiopatologia , Prognóstico , Piridonas/farmacologia
9.
PLoS One ; 9(11): e112430, 2014.
Artigo em Inglês | MEDLINE | ID: mdl-25390934

RESUMO

Supercentenarians (110 years or older) are the world's oldest people. Seventy four are alive worldwide, with twenty two in the United States. We performed whole-genome sequencing on 17 supercentenarians to explore the genetic basis underlying extreme human longevity. We found no significant evidence of enrichment for a single rare protein-altering variant or for a gene harboring different rare protein altering variants in supercentenarian compared to control genomes. We followed up on the gene most enriched for rare protein-altering variants in our cohort of supercentenarians, TSHZ3, by sequencing it in a second cohort of 99 long-lived individuals but did not find a significant enrichment. The genome of one supercentenarian had a pathogenic mutation in DSC2, known to predispose to arrhythmogenic right ventricular cardiomyopathy, which is recommended to be reported to this individual as an incidental finding according to a recent position statement by the American College of Medical Genetics and Genomics. Even with this pathogenic mutation, the proband lived to over 110 years. The entire list of rare protein-altering variants and DNA sequence of all 17 supercentenarian genomes is available as a resource to assist the discovery of the genetic basis of extreme longevity in future studies.


Assuntos
Envelhecimento/genética , Genoma Humano , Longevidade/genética , Idoso de 80 Anos ou mais , Desmocolinas/genética , Feminino , Estudo de Associação Genômica Ampla , Proteínas de Homeodomínio/genética , Humanos , Masculino , Mutação , Análise de Sequência de DNA
10.
Genome Res ; 17(9): 1286-95, 2007 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-17693573

RESUMO

Transcription factor complexes bind to regulatory sequences of genes, providing a system of individual expression regulation. Targets of distinct transcription factors usually map throughout the genome, without clustering. Nevertheless, highly and weakly expressed genes do cluster in separate chromosomal domains with an average size of 80-90 genes. We therefore asked whether, besides transcription factors, an additional level of gene expression regulation exists that acts on chromosomal domains. Here we show that identical green fluorescent protein (GFP) reporter constructs integrated at 90 different chromosomal positions obtain expression levels that correspond to the activity of the domains of integration. These domains are up to 80 genes long and can exert an eightfold effect on the expression levels of integrated genes. 3D-FISH shows that active domains of integration have a more open chromatin structure than integration domains with weak activity. These results reveal a novel domain-wide regulatory mechanism that, together with transcription factors, exerts a dual control over gene transcription.


Assuntos
Regulação da Expressão Gênica , Genoma Humano , Linhagem Celular , Cromossomos Humanos , Genes Reporter , Proteínas de Fluorescência Verde/metabolismo , Humanos , Imageamento Tridimensional , Hibridização in Situ Fluorescente , Rim/citologia , Lentivirus/genética , Análise de Sequência com Séries de Oligonucleotídeos , Fosfoglicerato Quinase/genética , Mapeamento Físico do Cromossomo , Regiões Promotoras Genéticas , Transcrição Gênica , Transfecção
11.
J Bacteriol ; 186(22): 7754-62, 2004 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-15516590

RESUMO

Despite the fact that phosphoenolpyruvate carboxylase (PEPC) activity has been measured and in some cases even purified from some Archaea, the gene responsible for this activity has not been elucidated. Using sensitive sequence comparison methods, we detected a highly conserved, uncharacterized archaeal gene family that is distantly related to the catalytic core of the canonical PEPC. To verify the predicted function of this archaeal gene family, we cloned a representative from the hyperthermophilic acidophile Sulfolobus solfataricus and functionally produced the corresponding enzyme as a fusion with the Escherichia coli maltose-binding protein. The purified fusion protein indeed displayed highly thermostable PEPC activity. The structural and biochemical properties of the characterized archaeal-type PEPC (atPEPC) from S. solfataricus are in good agreement with previously reported biochemical analyses of other archaeal PEPC enzymes. The newly identified atPEPC, with its distinct properties, constitutes yet another example of the versatility of the enzymes of the central carbon metabolic pathways in the archaeal domain.


Assuntos
Proteínas Arqueais/metabolismo , Fosfoenolpiruvato Carboxilase/química , Fosfoenolpiruvato Carboxilase/metabolismo , Sulfolobus solfataricus/enzimologia , Sequência de Aminoácidos , Archaea/enzimologia , Proteínas Arqueais/química , Proteínas Arqueais/genética , Biologia Computacional/métodos , Escherichia coli/enzimologia , Escherichia coli/genética , Genoma Arqueal , Dados de Sequência Molecular , Fosfoenolpiruvato Carboxilase/genética , Filogenia , Análise de Sequência de DNA , Sulfolobus solfataricus/genética , Sulfolobus solfataricus/crescimento & desenvolvimento
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