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1.
Nat Commun ; 11(1): 4544, 2020 09 11.
Artigo em Inglês | MEDLINE | ID: mdl-32917861

RESUMO

Stratification of enhancers by signal strength in ChIP-seq assays has resulted in the establishment of super-enhancers as a widespread and useful tool for identifying cell type-specific, highly expressed genes and associated pathways. We examine a distinct method of stratification that focuses on peak breadth, termed hyperacetylated chromatin domains (HCDs), which classifies broad regions exhibiting histone modifications associated with gene activation. We find that this analysis serves to identify genes that are both more highly expressed and more closely aligned to cell identity than super-enhancer analysis does using multiple data sets. Moreover, genetic manipulations of selected gene loci suggest that some enhancers located within HCDs work at least in part via a distinct mechanism involving the modulation of histone modifications across domains and that this activity can be imported into a heterologous gene locus. In addition, such genetic dissection reveals that the super-enhancer concept can obscure important functions of constituent elements.


Assuntos
Cromatina/metabolismo , Elementos Facilitadores Genéticos/genética , Loci Gênicos/genética , Ativação Transcricional , Acetilação , Animais , Linhagem Celular Tumoral , Imunoprecipitação da Cromatina , Sequenciamento de Cromatina por Imunoprecipitação , Conjuntos de Dados como Assunto , Embrião de Mamíferos , Eritroblastos , Feminino , Feto , Código das Histonas/genética , Histonas/genética , Histonas/metabolismo , Humanos , Camundongos , Regiões Promotoras Genéticas/genética , RNA-Seq
2.
Nat Commun ; 11(1): 4060, 2020 08 13.
Artigo em Inglês | MEDLINE | ID: mdl-32792512

RESUMO

Chromatin modifiers affect spatiotemporal gene expression programs that underlie organismal development. The Polycomb repressive complex 2 (PRC2) is a crucial chromatin modifier in executing neurodevelopmental programs. Here, we find that PRC2 interacts with the nucleic acid-binding protein Ybx1. In the mouse embryo in vivo, Ybx1 is required for forebrain specification and restricting mid-hindbrain growth. In neural progenitor cells (NPCs), Ybx1 controls self-renewal and neuronal differentiation. Mechanistically, Ybx1 highly overlaps PRC2 binding genome-wide, controls PRC2 distribution, and inhibits H3K27me3 levels. These functions are consistent with Ybx1-mediated promotion of genes involved in forebrain specification, cell proliferation, or neuronal differentiation. In Ybx1-knockout NPCs, H3K27me3 reduction by PRC2 enzymatic inhibitor or genetic depletion partially rescues gene expression and NPC functions. Our findings suggest that Ybx1 fine-tunes PRC2 activities to regulate spatiotemporal gene expression in embryonic neural development and uncover a crucial epigenetic mechanism balancing forebrain-hindbrain lineages and self-renewal-differentiation choices in NPCs.


Assuntos
Encéfalo/embriologia , Encéfalo/metabolismo , Histona-Lisina N-Metiltransferase/metabolismo , Fatores de Transcrição/metabolismo , Animais , Western Blotting , Diferenciação Celular/genética , Diferenciação Celular/fisiologia , Proliferação de Células/genética , Proliferação de Células/fisiologia , Células Cultivadas , Imunoprecipitação da Cromatina , Drosophila , Epigênese Genética/genética , Citometria de Fluxo , Imunofluorescência , Histona-Lisina N-Metiltransferase/genética , Imunoprecipitação , Camundongos , Camundongos Knockout , Processamento de Proteína Pós-Traducional/genética , Processamento de Proteína Pós-Traducional/fisiologia , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Fatores de Transcrição/genética
3.
Nat Commun ; 11(1): 4133, 2020 08 17.
Artigo em Inglês | MEDLINE | ID: mdl-32807777

RESUMO

Chromatin organization is critical for cell growth, differentiation, and disease development, however, its functions in peripheral myelination and myelin repair remain elusive. In this report, we demonstrate that the CCCTC-binding factor (CTCF), a crucial chromatin organizer, is essential for Schwann cell myelination and myelin regeneration after nerve injury. Inhibition of CTCF or its deletion blocks Schwann cell differentiation at the pro-myelinating stage, whereas overexpression of CTCF promotes the myelination program. We find that CTCF establishes chromatin interaction loops between enhancer and promoter regulatory elements and promotes expression of a key pro-myelinogenic factor EGR2. In addition, CTCF interacts with SUZ12, a component of polycomb-repressive-complex 2 (PRC2), to repress the transcriptional program associated with negative regulation of Schwann cell maturation. Together, our findings reveal a dual role of CTCF-dependent chromatin organization in promoting myelinogenic programs and recruiting chromatin-repressive complexes to block Schwann cell differentiation inhibitors to control peripheral myelination and repair.


Assuntos
Fator de Ligação a CCCTC/metabolismo , Cromatina/metabolismo , Proteína 2 de Resposta de Crescimento Precoce/metabolismo , Bainha de Mielina/metabolismo , Complexo Repressor Polycomb 2/metabolismo , Animais , Fator de Ligação a CCCTC/genética , Células Cultivadas , Imunoprecipitação da Cromatina , Proteína 2 de Resposta de Crescimento Precoce/genética , Camundongos , Bainha de Mielina/genética , Complexo Repressor Polycomb 2/genética , Ratos , Células de Schwann/metabolismo
4.
Nat Commun ; 11(1): 4055, 2020 08 13.
Artigo em Inglês | MEDLINE | ID: mdl-32792504

RESUMO

Although metastasis is the most common cause of cancer deaths, metastasis-intrinsic dependencies remain largely uncharacterized. We previously reported that metastatic pancreatic cancers were dependent on the glucose-metabolizing enzyme phosphogluconate dehydrogenase (PGD). Surprisingly, PGD catalysis was constitutively elevated without activating mutations, suggesting a non-genetic basis for enhanced activity. Here we report a metabolic adaptation that stably activates PGD to reprogram metastatic chromatin. High PGD catalysis prevents transcriptional up-regulation of thioredoxin-interacting protein (TXNIP), a gene that negatively regulates glucose import. This allows glucose consumption rates to rise in support of PGD, while simultaneously facilitating epigenetic reprogramming through a glucose-fueled histone hyperacetylation pathway. Restoring TXNIP normalizes glucose consumption, lowers PGD catalysis, reverses hyperacetylation, represses malignant transcripts, and impairs metastatic tumorigenesis. We propose that PGD-driven suppression of TXNIP allows pancreatic cancers to avidly consume glucose. This renders PGD constitutively activated and enables metaboloepigenetic selection of additional traits that increase fitness along glucose-replete metastatic routes.


Assuntos
Cromatina/metabolismo , Glucose/metabolismo , Neoplasias Pancreáticas/metabolismo , Neoplasias Pancreáticas/patologia , Animais , Transporte Biológico/genética , Transporte Biológico/fisiologia , Proteínas de Transporte/genética , Proteínas de Transporte/metabolismo , Proteínas de Ciclo Celular/genética , Proteínas de Ciclo Celular/metabolismo , Reprogramação Celular/genética , Reprogramação Celular/fisiologia , Imunoprecipitação da Cromatina , Epigênese Genética/genética , Camundongos , Camundongos Nus , Neoplasias Pancreáticas/genética , Fosfogluconato Desidrogenase/genética , Fosfogluconato Desidrogenase/metabolismo , Tiorredoxinas/genética , Tiorredoxinas/metabolismo
5.
BMC Bioinformatics ; 21(1): 281, 2020 Jul 02.
Artigo em Inglês | MEDLINE | ID: mdl-32615918

RESUMO

BACKGROUND: During transcription, numerous transcription factors (TFs) bind to targets in a highly coordinated manner to control the gene expression. Alterations in groups of TF-binding profiles (i.e. "co-binding changes") can affect the co-regulating associations between TFs (i.e. "rewiring the co-regulator network"). This, in turn, can potentially drive downstream expression changes, phenotypic variation, and even disease. However, quantification of co-regulatory network rewiring has not been comprehensively studied. RESULTS: To address this, we propose DiNeR, a computational method to directly construct a differential TF co-regulation network from paired disease-to-normal ChIP-seq data. Specifically, DiNeR uses a graphical model to capture the gained and lost edges in the co-regulation network. Then, it adopts a stability-based, sparsity-tuning criterion -- by sub-sampling the complete binding profiles to remove spurious edges -- to report only significant co-regulation alterations. Finally, DiNeR highlights hubs in the resultant differential network as key TFs associated with disease. We assembled genome-wide binding profiles of 104 TFs in the K562 and GM12878 cell lines, which loosely model the transition between normal and cancerous states in chronic myeloid leukemia (CML). In total, we identified 351 significantly altered TF co-regulation pairs. In particular, we found that the co-binding of the tumor suppressor BRCA1 and RNA polymerase II, a well-known transcriptional pair in healthy cells, was disrupted in tumors. Thus, DiNeR successfully extracted hub regulators and discovered well-known risk genes. CONCLUSIONS: Our method DiNeR makes it possible to quantify changes in co-regulatory networks and identify alterations to TF co-binding patterns, highlighting key disease regulators. Our method DiNeR makes it possible to quantify changes in co-regulatory networks and identify alterations to TF co-binding patterns, highlighting key disease regulators.


Assuntos
Redes Reguladoras de Genes , Modelos Genéticos , Software , Imunoprecipitação da Cromatina , Regulação da Expressão Gênica , Genoma , Humanos , Células K562 , Leucemia Mielogênica Crônica BCR-ABL Positiva/genética , Ligação Proteica , Fatores de Transcrição/metabolismo , Transcrição Genética
6.
Nat Commun ; 11(1): 3520, 2020 07 14.
Artigo em Inglês | MEDLINE | ID: mdl-32665551

RESUMO

PRDM (PRDI-BF1 and RIZ homology domain containing) family members are sequence-specific transcriptional regulators involved in cell identity and fate determination, often dysregulated in cancer. The PRDM15 gene is of particular interest, given its low expression in adult tissues and its overexpression in B-cell lymphomas. Despite its well characterized role in stem cell biology and during early development, the role of PRDM15 in cancer remains obscure. Herein, we demonstrate that while PRDM15 is largely dispensable for mouse adult somatic cell homeostasis in vivo, it plays a critical role in B-cell lymphomagenesis. Mechanistically, PRDM15 regulates a transcriptional program that sustains the activity of the PI3K/AKT/mTOR pathway and glycolysis in B-cell lymphomas. Abrogation of PRDM15 induces a metabolic crisis and selective death of lymphoma cells. Collectively, our data demonstrate that PRDM15 fuels the metabolic requirement of B-cell lymphomas and validate it as an attractive and previously unrecognized target in oncology.


Assuntos
Proteínas de Ligação a DNA/metabolismo , Fatores de Transcrição/metabolismo , Animais , Apoptose/genética , Apoptose/fisiologia , Western Blotting , Sobrevivência Celular/genética , Sobrevivência Celular/fisiologia , Imunoprecipitação da Cromatina , Biologia Computacional , Proteínas de Ligação a DNA/genética , Feminino , Citometria de Fluxo , Regulação da Expressão Gênica/genética , Regulação da Expressão Gênica/fisiologia , Humanos , Linfoma/genética , Linfoma/metabolismo , Camundongos , Camundongos SCID , Fosfatidilinositol 3-Quinases/genética , Fosfatidilinositol 3-Quinases/metabolismo , Distribuição Aleatória , Fatores de Transcrição/genética , Transcriptoma/genética
7.
Plant Mol Biol ; 104(1-2): 113-136, 2020 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-32627097

RESUMO

KEY MESSAGE: Present study revealed a complex relationship among histone H3 methylation (examined using H3K4/K27me3 marks), cytosine DNA methylation and differential gene expression during Lr28 mediated leaf rust resistance in wheat. During the present study, genome-wide histone modifications were examined in a pair of near isogenic lines (NILs) (with and without Lr28 in the background of cv. HD2329). The two histone marks used included H3K4me3 (an activation mark) and H3K27me3 (a repression mark). The results were compared with levels of expression (using RNA-seq) and DNA methylation (MeDIP) data obtained using the same pair of NILs. Some of the salient features of the present study include the following: (i) large scale differential binding sites (DBS) were available for only H3K4me3 in the susceptible cultivar, but for both H3K4me3 and H3K27me3 in its resistant NIL; (ii) DBSs for H3K27me3 mark were more abundant (> 80%) in intergenic regions, whereas DBSs for H3K4me3 were distributed in all genomic regions including exons, introns, intergenic, TTS (transcription termination sites) and promoters; (iii) fourteen (14) genes associated with DBSs showed co-localization for both the marks; (iv) only a small fraction (7% for H3K4me3 and 12% for H3K27me3) of genes associated with DBSs matched with the levels of gene expression inferred from RNA-seq data; (v) validation studies using qRT-PCR were conducted on 26 selected representative genes; results for only 11 genes could be validated. The proteins encoded by important genes involved in promoting infection included domains generally carried by R gene proteins such as Mlo like protein, protein kinases and purple acid phosphatase. Similarly, proteins encoded by genes involved in resistance included those carrying domains for lectin kinase, R gene, aspartyl protease, etc. Overall, the results suggest a very complex network of downstream genes that are expressed during compatible and incompatible interactions; some of the genes identified during the present study may be used in future validation studies involving RNAi/overexpression approaches.


Assuntos
Basidiomycota/metabolismo , Resistência à Doença/genética , Genes de Plantas/genética , Genoma de Planta/genética , Histonas/genética , Doenças das Plantas/genética , Triticum/genética , Triticum/metabolismo , Imunoprecipitação da Cromatina , Metilação de DNA , Regulação da Expressão Gênica de Plantas , Ligação Genética , Histonas/metabolismo , Anotação de Sequência Molecular , Doenças das Plantas/microbiologia , Folhas de Planta/genética , Folhas de Planta/microbiologia , Regiões Promotoras Genéticas , Reprodutibilidade dos Testes , Alinhamento de Sequência , Análise de Sequência , Análise de Sequência de RNA , Transcrição Genética , Triticum/microbiologia
8.
Nat Commun ; 11(1): 2922, 2020 06 10.
Artigo em Inglês | MEDLINE | ID: mdl-32523103

RESUMO

The conversion of white adipocytes to thermogenic beige adipocytes represents a potential mechanism to treat obesity and related metabolic disorders. However, the mechanisms involved in converting white to beige adipose tissue remain incompletely understood. Here we show profound beiging in a genetic mouse model lacking the transcriptional repressor Krüppel-like factor 3 (KLF3). Bone marrow transplants from these animals confer the beige phenotype on wild type recipients. Analysis of the cellular and molecular changes reveal an accumulation of eosinophils in adipose tissue. We examine the transcriptomic profile of adipose-resident eosinophils and posit that KLF3 regulates adipose tissue function via transcriptional control of secreted molecules linked to beiging. Furthermore, we provide evidence that eosinophils may directly act on adipocytes to drive beiging and highlight the critical role of these little-understood immune cells in thermogenesis.


Assuntos
Tecido Adiposo/metabolismo , Eosinófilos/metabolismo , Fatores de Transcrição Kruppel-Like/metabolismo , Transdução de Sinais/fisiologia , Adiposidade/genética , Adiposidade/fisiologia , Animais , Células COS , Chlorocebus aethiops , Imunoprecipitação da Cromatina , Citometria de Fluxo , Fatores de Transcrição Kruppel-Like/genética , Masculino , Camundongos , Obesidade/metabolismo , Transdução de Sinais/genética , Software
9.
PLoS One ; 15(6): e0235343, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32584896

RESUMO

Triple Negative Breast Cancer (TNBC) is a heterogeneous disease lacking known molecular drivers and effective targeted therapies. Cytotoxic chemotherapy remains the mainstay of treatment for TNBCs, which have significantly poorer survival rates compared to other breast cancer subtypes. In addition to changes within the coding genome, aberrant enhancer activity is a well-established contributor to tumorigenesis. Here we use H3K27Ac chromatin immunoprecipitation followed by sequencing (ChIP-Seq) to map the active cis-regulatory landscape in TNBC. We identify distinct disease subtypes associated with specific enhancer activity, and over 2,500 unique superenhancers acquired by tumor cells but absent from normal breast tissue. To identify potential actionable disease drivers, we probed the dependency on genes that associate with tumor-specific enhancers by CRISPR screening. In this way we identify a number of tumor-specific dependencies, including a previously uncharacterized dependency on the TGFß pseudo-receptor BAMBI.


Assuntos
Elementos Facilitadores Genéticos/genética , Oncogenes/genética , Neoplasias de Mama Triplo Negativas/patologia , Linhagem Celular Tumoral , Imunoprecipitação da Cromatina , Feminino , Edição de Genes , Regulação Neoplásica da Expressão Gênica , Histonas/química , Histonas/genética , Histonas/metabolismo , Humanos , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , RNA Guia/metabolismo , Neoplasias de Mama Triplo Negativas/genética
10.
Nucleic Acids Res ; 48(11): 5967-5985, 2020 06 19.
Artigo em Inglês | MEDLINE | ID: mdl-32406921

RESUMO

During infection of a host, Pseudomonas aeruginosa orchestrates global gene expression to adapt to the host environment and counter the immune attacks. P. aeruginosa harbours hundreds of regulatory genes that play essential roles in controlling gene expression. However, their contributions to the bacterial pathogenesis remain largely unknown. In this study, we analysed the transcriptomic profile of P. aeruginosa cells isolated from lungs of infected mice and examined the roles of upregulated regulatory genes in bacterial virulence. Mutation of a novel regulatory gene pvrA (PA2957) attenuated the bacterial virulence in an acute pneumonia model. Chromatin immunoprecipitation (ChIP)-Seq and genetic analyses revealed that PvrA directly regulates genes involved in phosphatidylcholine utilization and fatty acid catabolism. Mutation of the pvrA resulted in defective bacterial growth when phosphatidylcholine or palmitic acid was used as the sole carbon source. We further demonstrated that palmitoyl coenzyme A is a ligand for the PvrA, enhancing the binding affinity of PvrA to its target promoters. An arginine residue at position 136 was found to be essential for PvrA to bind palmitoyl coenzyme A. Overall, our results revealed a novel regulatory pathway that controls genes involved in phosphatidylcholine and fatty acid utilization and contributes to the bacterial virulence.


Assuntos
Proteínas de Bactérias/metabolismo , Ácidos Graxos/química , Ácidos Graxos/metabolismo , Genes Bacterianos/genética , Pseudomonas aeruginosa/metabolismo , Pseudomonas aeruginosa/patogenicidade , Animais , Arginina/metabolismo , Sequência de Bases , Imunoprecipitação da Cromatina , Modelos Animais de Doenças , Perfilação da Expressão Gênica , Regulação Bacteriana da Expressão Gênica , Ligantes , Camundongos , Modelos Moleculares , Mutação , Ácido Palmítico/metabolismo , Palmitoil Coenzima A/metabolismo , Fosfatidilcolinas/metabolismo , Pneumonia Bacteriana/microbiologia , Regiões Promotoras Genéticas , Pseudomonas aeruginosa/genética , Transcriptoma , Virulência/genética
11.
Nat Commun ; 11(1): 2156, 2020 05 01.
Artigo em Inglês | MEDLINE | ID: mdl-32358485

RESUMO

Colorectal cancer (CRC) is the most common gastrointestinal malignancy in the U.S.A. and approximately 50% of patients develop metastatic disease (mCRC). Despite our understanding of long non-coding RNAs (lncRNAs) in primary colon cancer, their role in mCRC and treatment resistance remains poorly characterized. Therefore, through transcriptome sequencing of normal, primary, and distant mCRC tissues we find 148 differentially expressed RNAs Associated with Metastasis (RAMS). We prioritize RAMS11 due to its association with poor disease-free survival and promotion of aggressive phenotypes in vitro and in vivo. A FDA-approved drug high-throughput viability assay shows that elevated RAMS11 expression increases resistance to topoisomerase inhibitors. Subsequent experiments demonstrate RAMS11-dependent recruitment of Chromobox protein 4 (CBX4) transcriptionally activates Topoisomerase II alpha (TOP2α). Overall, recent clinical trials using topoisomerase inhibitors coupled with our findings of RAMS11-dependent regulation of TOP2α supports the potential use of RAMS11 as a biomarker and therapeutic target for mCRC.


Assuntos
Neoplasias Colorretais/genética , Neoplasias Colorretais/patologia , RNA Longo não Codificante/genética , RNA Longo não Codificante/metabolismo , Animais , Western Blotting , Células CACO-2 , Linhagem Celular Tumoral , Imunoprecipitação da Cromatina , Biologia Computacional , DNA Topoisomerases Tipo II/metabolismo , Progressão da Doença , Éxons/genética , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Regulação Neoplásica da Expressão Gênica/genética , Células HCT116 , Células HT29 , Humanos , Ligases/metabolismo , Camundongos , Proteínas do Grupo Polycomb/metabolismo , RNA-Seq , Reação em Cadeia da Polimerase em Tempo Real , Inibidores da Topoisomerase/farmacologia
12.
PLoS Negl Trop Dis ; 14(5): e0008262, 2020 05.
Artigo em Inglês | MEDLINE | ID: mdl-32469928

RESUMO

Adhesion of T. cruzi trypomastigotes to components of the extracellular matrix (ECM) is an important step in mammalian host cell invasion. We have recently described a significant increase in the tyrosine nitration levels of histones H2A and H4 when trypomastigotes are incubated with components of the ECM. In this work, we used chromatin immunoprecipitation (ChIP) with an anti-nitrotyrosine antibody followed by mass spectrometry to identify nitrated DNA binding proteins in T. cruzi and to detect alterations in nitration levels induced upon parasite incubation with the ECM. Histone H1, H2B, H2A and H3 were detected among the 9 most abundant nitrated DNA binding proteins using this proteomic approach. One nitrated tyrosine residue (Y29) was identified in Histone H2B in the MS/MS spectrum. In addition, we observed a significant increase in the nitration levels of histones H1, H2B, H2A and H4 upon parasite incubation with ECM. Finally, we used ChIP-Seq to map global changes in the DNA binding profile of nitrated proteins. We observed a significant change in the binding pattern of nitrated proteins to DNA after parasite incubation with ECM. This work provides the first global profile of nitrated DNA binding proteins in T. cruzi and additional evidence for modification in the nitration profile of histones upon parasite incubation with ECM. Our data also indicate that the parasite interaction with the ECM induces alterations in chromatin structure, possibly affecting nuclear functions.


Assuntos
Matriz Extracelular/parasitologia , Histonas/análise , Processamento de Proteína Pós-Traducional , Proteínas de Protozoários/análise , Trypanosoma cruzi/química , Trypanosoma cruzi/crescimento & desenvolvimento , Imunoprecipitação da Cromatina , Matriz Extracelular/metabolismo , Histonas/metabolismo , Espectrometria de Massas , Nitrosação , Proteômica , Proteínas de Protozoários/metabolismo , Tirosina/análogos & derivados , Tirosina/imunologia
13.
Gene ; 753: 144805, 2020 Aug 30.
Artigo em Inglês | MEDLINE | ID: mdl-32445923

RESUMO

Genomic variants in both ADTRP and TFPI genes are associated with risk of coronary artery disease (CAD). ADTRP regulates TFPI expression and endothelial cell functions involved in the initiation of atherosclerotic CAD. ADTRP also specifies primitive myelopoiesis and definitive hematopoiesis by upregulating TFPI expression. However, the underlying molecular mechanism is unknown. Here we show that transcription factor POU1F1 is the key by which ADTRP regulates TFPI expression. Luciferase reporter assays, chromatin-immunoprecipitation (ChIP) and electrophoretic mobility shift assay (EMSA) in combination with analysis of large and small deletions of the TFPI promoter/regulatory region were used to identify the molecular mechanism by which ADTRP regulates TFPI expression. Genetic association was assessed using case-control association analysis and phenome-wide association analysis (PhenGWA). ADTRP regulates TFPI expression at the transcription level in a dose-dependent manner. The ADTRP-response element was localized to a 50 bp region between -806 bp and -756 bp upstream of TFPI transcription start site, which contains a binding site for POU1F1. Deletion of POU1F1-binding site or knockdown of POU1F1 expression abolished ADTRP-mediated transcription of TFPI. ChIP and EMSA demonstrated that POU1F1 binds to the ADTRP response element. Genetic analysis identified significant association between POU1F1 variants and risk of CAD. PhenGWA identified other phenotypic traits associated with the ADTRP-POU1F1-TFPI axis such as lymphocyte count (ADTRP), waist circumference (TFPI), and standing height (POU1F1). These data identify POU1F1 as a transcription factor that regulates TFPI transcription in response to ADTRP, and link POU1F1 variants to risk of CAD for the first time.


Assuntos
Doença da Artéria Coronariana/metabolismo , Lipoproteínas/biossíntese , Proteínas de Membrana/metabolismo , Fator de Transcrição Pit-1/metabolismo , Aterosclerose/genética , Estudos de Casos e Controles , Linhagem Celular , Imunoprecipitação da Cromatina/métodos , Doença da Artéria Coronariana/genética , Bases de Dados Genéticas , Células Endoteliais/metabolismo , Genes Homeobox , Células HeLa , Humanos , Lipoproteínas/genética , Lipoproteínas/metabolismo , Proteínas de Membrana/genética , Proteínas de Membrana/fisiologia , Regiões Promotoras Genéticas , Elementos de Resposta , Sítio de Iniciação de Transcrição , Transcrição Genética
14.
Proc Natl Acad Sci U S A ; 117(21): 11471-11482, 2020 05 26.
Artigo em Inglês | MEDLINE | ID: mdl-32385160

RESUMO

Lineage plasticity is a prominent feature of pancreatic ductal adenocarcinoma (PDA) cells, which can occur via deregulation of lineage-specifying transcription factors. Here, we show that the zinc finger protein ZBED2 is aberrantly expressed in PDA and alters tumor cell identity in this disease. Unexpectedly, our epigenomic experiments reveal that ZBED2 is a sequence-specific transcriptional repressor of IFN-stimulated genes, which occurs through antagonism of IFN regulatory factor 1 (IRF1)-mediated transcriptional activation at cooccupied promoter elements. Consequently, ZBED2 attenuates the transcriptional output and growth arrest phenotypes downstream of IFN signaling in multiple PDA cell line models. We also found that ZBED2 is preferentially expressed in the squamous molecular subtype of human PDA, in association with inferior patient survival outcomes. Consistent with this observation, we show that ZBED2 can repress the pancreatic progenitor transcriptional program, enhance motility, and promote invasion in PDA cells. Collectively, our findings suggest that high ZBED2 expression is acquired during PDA progression to suppress the IFN response pathway and to promote lineage plasticity in this disease.


Assuntos
Carcinoma Ductal Pancreático/patologia , Proteínas de Ligação a DNA/metabolismo , Fator Regulador 1 de Interferon/metabolismo , Neoplasias Pancreáticas/patologia , Fatores de Transcrição/metabolismo , Animais , Carcinoma Ductal Pancreático/genética , Carcinoma Ductal Pancreático/metabolismo , Carcinoma Ductal Pancreático/mortalidade , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Imunoprecipitação da Cromatina , Proteínas de Ligação a DNA/genética , Regulação Neoplásica da Expressão Gênica , Humanos , Fator Regulador 1 de Interferon/genética , Interferon gama/farmacologia , Camundongos , Neoplasias Pancreáticas/genética , Neoplasias Pancreáticas/metabolismo , Neoplasias Pancreáticas/mortalidade , Regiões Promotoras Genéticas , Análise de Sobrevida , Fatores de Transcrição/genética
15.
Life Sci ; 253: 117693, 2020 Jul 15.
Artigo em Inglês | MEDLINE | ID: mdl-32325133

RESUMO

AIMS: Thyroid cancer is a common endocrine malignancy and sex hormone plays an important role in it. We have previously shown that activation of estrogen receptor (ER) α promotes thyroid cancer cell proliferation and invasion. Here, we attempted to investigate the role of ETS variant 5 (ETV5) on estrogen drived thyroid malignancy. MAIN METHODS: Ten patients with follicular thyroid cancer were enrolled in this study. Cell proliferation and migration ability were analyzed by CCK-8 assay and cell migration assay, respectively. Chromatin immunoprecipitation-PCR and luciferase assay were conducted to analyze the relationship of ETV5 and PIK3CA. KEY FINDINGS: ETV5 is highly expressed in thyroid tissues from patients with follicular thyroid cancer as well as in FTC133 cells. 17b-estradiol or overexpression of ERα induced an increase in ETV5 protein level in FTC133 cells. Knockdown of ETV5 inhibited FTC133 cell proliferation, migration, and epithelial-mesenchymal transition, while 17b-estradiol could not correct this effect. Additionally, the level of PIK3CA was markedly decreased in ETV5 knockdown cells and had a positive correlation with ETV5 in thyroid cancer patients. Chromatin immunoprecipitation-PCR analysis and luciferase assay confirmed that ETV5 directly targeted PIK3CA and that ETV5 was bound to the promoter region of PIK3CA. In addition, PIK3CA overexpression abrogated ETV5-induced cell growth, migration and epithelial-mesenchymal transition. SIGNIFICANCE: ETV5 enhanced cell proliferation, migration, and epithelial-mesenchymal transition through the PIK3CA signaling pathway, indicating that ETV5 may be a therapeutic target in thyroid cancer.


Assuntos
Adenocarcinoma Folicular/patologia , Classe I de Fosfatidilinositol 3-Quinases/metabolismo , Proteínas de Ligação a DNA/genética , Neoplasias da Glândula Tireoide/patologia , Fatores de Transcrição/genética , Adenocarcinoma Folicular/genética , Adulto , Linhagem Celular Tumoral , Movimento Celular/genética , Proliferação de Células/genética , Imunoprecipitação da Cromatina , Progressão da Doença , Transição Epitelial-Mesenquimal/genética , Receptor alfa de Estrogênio/genética , Feminino , Técnicas de Silenciamento de Genes , Humanos , Pessoa de Meia-Idade , Transdução de Sinais , Neoplasias da Glândula Tireoide/genética
16.
Mol Cell ; 78(3): 459-476.e13, 2020 05 07.
Artigo em Inglês | MEDLINE | ID: mdl-32240602

RESUMO

The cyclin-dependent kinase 1 (Cdk1) drives cell division. To uncover additional functions of Cdk1, we generated knockin mice expressing an analog-sensitive version of Cdk1 in place of wild-type Cdk1. In our study, we focused on embryonic stem cells (ESCs), because this cell type displays particularly high Cdk1 activity. We found that in ESCs, a large fraction of Cdk1 substrates is localized on chromatin. Cdk1 phosphorylates many proteins involved in epigenetic regulation, including writers and erasers of all major histone marks. Consistent with these findings, inhibition of Cdk1 altered histone-modification status of ESCs. High levels of Cdk1 in ESCs phosphorylate and partially inactivate Dot1l, the H3K79 methyltransferase responsible for placing activating marks on gene bodies. Decrease of Cdk1 activity during ESC differentiation de-represses Dot1l, thereby allowing coordinated expression of differentiation genes. These analyses indicate that Cdk1 functions to maintain the epigenetic identity of ESCs.


Assuntos
Proteína Quinase CDC2/metabolismo , Células-Tronco Embrionárias/fisiologia , Epigênese Genética , Trifosfato de Adenosina/análogos & derivados , Trifosfato de Adenosina/metabolismo , Animais , Proteína Quinase CDC2/genética , Diferenciação Celular , Células Cultivadas , Imunoprecipitação da Cromatina/métodos , Feminino , Histona-Lisina N-Metiltransferase/genética , Histona-Lisina N-Metiltransferase/metabolismo , Humanos , Células MCF-7 , Masculino , Camundongos , Camundongos Knockout , Fosforilação , Proteínas de Saccharomyces cerevisiae/metabolismo
17.
PLoS Comput Biol ; 16(4): e1007800, 2020 04.
Artigo em Inglês | MEDLINE | ID: mdl-32251445

RESUMO

Transcriptomic profiling is an immensely powerful hypothesis generating tool. However, accurately predicting the transcription factors (TFs) and cofactors that drive transcriptomic differences between samples is challenging. A number of algorithms draw on ChIP-seq tracks to define TFs and cofactors behind gene changes. These approaches assign TFs and cofactors to genes via a binary designation of 'target', or 'non-target' followed by Fisher Exact Tests to assess enrichment of TFs and cofactors. ENCODE archives 2314 ChIP-seq tracks of 684 TFs and cofactors assayed across a 117 human cell lines under a multitude of growth and maintenance conditions. The algorithm presented herein, Mining Algorithm for GenetIc Controllers (MAGIC), uses ENCODE ChIP-seq data to look for statistical enrichment of TFs and cofactors in gene bodies and flanking regions in gene lists without an a priori binary classification of genes as targets or non-targets. When compared to other TF mining resources, MAGIC displayed favourable performance in predicting TFs and cofactors that drive gene changes in 4 settings: 1) A cell line expressing or lacking single TF, 2) Breast tumors divided along PAM50 designations 3) Whole brain samples from WT mice or mice lacking a single TF in a particular neuronal subtype 4) Single cell RNAseq analysis of neurons divided by Immediate Early Gene expression levels. In summary, MAGIC is a standalone application that produces meaningful predictions of TFs and cofactors in transcriptomic experiments.


Assuntos
Biologia Computacional/métodos , Perfilação da Expressão Gênica/métodos , Software , Fatores de Transcrição , Algoritmos , Animais , Linhagem Celular , Imunoprecipitação da Cromatina , Bases de Dados Genéticas , Humanos , Células MCF-7 , Camundongos , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo
18.
Proc Natl Acad Sci U S A ; 117(13): 7140-7149, 2020 03 31.
Artigo em Inglês | MEDLINE | ID: mdl-32188783

RESUMO

The recognition of cis-regulatory RNA motifs in human transcripts by RNA binding proteins (RBPs) is essential for gene regulation. The molecular features that determine RBP specificity are often poorly understood. Here, we combined NMR structural biology with high-throughput iCLIP approaches to identify a regulatory mechanism for U2AF2 RNA recognition. We found that the intrinsically disordered linker region connecting the two RNA recognition motif (RRM) domains of U2AF2 mediates autoinhibitory intramolecular interactions to reduce nonproductive binding to weak Py-tract RNAs. This proofreading favors binding of U2AF2 at stronger Py-tracts, as required to define 3' splice sites at early stages of spliceosome assembly. Mutations that impair the linker autoinhibition enhance the affinity for weak Py-tracts result in promiscuous binding of U2AF2 along mRNAs and impact on splicing fidelity. Our findings highlight an important role of intrinsically disordered linkers to modulate RNA interactions of multidomain RBPs.


Assuntos
RNA/metabolismo , Fator de Processamento U2AF/metabolismo , Animais , Bovinos , Imunoprecipitação da Cromatina/métodos , Humanos , Espectroscopia de Ressonância Magnética , Camundongos , Motivo de Reconhecimento de RNA , Ribonucleosídeo Difosfato Redutase/metabolismo
19.
Sheng Wu Gong Cheng Xue Bao ; 36(2): 341-352, 2020 Feb 25.
Artigo em Chinês | MEDLINE | ID: mdl-32148006

RESUMO

Sonication is one of the essential strategies of chromatin fragmentation in Chromatin immunoprecipitation (ChIP) assay. The impact of proteins with different molecular weights generated under different duration of sonication on the results of Chromatin immunoprecipitation sequencing (ChIP-seq) experiments involving the Polycomb Repressive Complex 2 (PRC2) related protein EZH2, which is a histone methyltransferase, and its product H3K27me3 was investigated. The results indicate that in the promoter region or nonpromoter region, there were hardly any differences among the H3K27me3 peaks annotated genes from different duration of sonication, which suggesting that the duration of sonication had little effect on histone ChIP-seq results. In contrast, in the promoter region newly gained EZH2 peaks annonated genes at 20 min sonication time were significantly clustered in the gene ontology (GO) pathways related to actin filament bundle compared with 10 min. In the nonpromoter region, compared with 10 min, the GO pathways of newly gained EZH2 peaks annonated genes at 20 min sonication time is much more than that of lost EZH2 peaks annonated genes. And in the nonpromoter region, compared with 20 min, the GO pathways of lost EZH2 peaks annonated genes at 30 min sonication time is much more than that of newly gained EZH2 peaks annonated genes. Most of these pathways are associated with RNA polymerase II (RNAPII), organ development and cell morphogenesis. These suggest that the genomic information of EZH2 will be lost if the duration of sonication is not enough or too long. Different duration of sonication mainly affect the EZH2 peaks in PRC2 unoccupied region and the bivalent promoter in the promoter region, as well as the PRC2 occupied region, PRC2 unoccupied region and the activated enhancer in the nonpromoter region. Therefore, the sonication for the chromatin related proteins with high molecular weights need to be optimized to make chromatin fragments size vary from 100 bp to 500 bp, which will yield relatively comprehensive genomic information of the target protein. For histones, which are of small molecular weights, duration of sonication has little effect on them.


Assuntos
Sonicação , Imunoprecipitação da Cromatina , Proteína Potenciadora do Homólogo 2 de Zeste , Histonas , Complexo Repressor Polycomb 2
20.
Life Sci ; 249: 117479, 2020 May 15.
Artigo em Inglês | MEDLINE | ID: mdl-32119959

RESUMO

A concomitant change of nucleus shape and chromosome conformation often happens in all-trans retinoic acid (ATRA)-induced differentiation of acute myeloid leukemia cells. However, the relation between the 3D chromosome architecture and the genome-wide epigenetic pattern for transcriptional regulation is poorly understood. In this study, high-throughput chromosome conformation capture (Hi-C) and chromosome immunoprecipitation (ChIP-seq) were employed to investigate the landscape of chromosome distal interaction and H3K4/27me3 in HL-60 cells treated with ATRA. We observed a general loss of topological associated domains (TADs) at PTPN11 during the differentiation of HL-60 cells. Furthermore, the significantly reduced enrichment of CCCTC binding factor (CTCF) near the boundary where PTPN11 located, as well as the decreased H3K4me3 and increased H3K27me3 enrichment at PTPN11 upon ATRA treatment was observed. Taken together, our study indicated a regulatory mechanism behind the silenced PTPN11 in HL-60 cells differentiation.


Assuntos
Diferenciação Celular/efeitos dos fármacos , Cromossomos Humanos , Leucemia Mieloide Aguda/patologia , Tretinoína/farmacologia , Imunoprecipitação da Cromatina , Células HL-60 , Histonas/metabolismo , Humanos , Leucemia Mieloide Aguda/genética
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