scholarly journals Enhancer-bound LDB1 regulates a corticotrope promoter-pausing repression program

2015 ◽  
Vol 112 (5) ◽  
pp. 1380-1385 ◽  
Author(s):  
Feng Zhang ◽  
Bogdan Tanasa ◽  
Daria Merkurjev ◽  
Chijen Lin ◽  
Xiaoyuan Song ◽  
...  
Keyword(s):  
Target Gene ◽  
Target Genes ◽  
Binding Protein ◽  
Homeodomain Protein ◽  
Gene Promoters ◽  
Cell Type ◽  
Lim Domain ◽  
Transcriptional Initiation ◽  
Nucleosome Remodeling ◽  
Enhancer Binding Protein

Substantial evidence supports the hypothesis that enhancers are critical regulators of cell-type determination, orchestrating both positive and negative transcriptional programs; however, the basic mechanisms by which enhancers orchestrate interactions with cognate promoters during activation and repression events remain incompletely understood. Here we report the required actions of LIM domain-binding protein 1 (LDB1)/cofactor of LIM homeodomain protein 2/nuclear LIM interactor, interacting with the enhancer-binding protein achaete-scute complex homolog 1, to mediate looping to target gene promoters and target gene regulation in corticotrope cells. LDB1-mediated enhancer:promoter looping appears to be required for both activation and repression of these target genes. Although LDB1-dependent activated genes are regulated at the level of transcriptional initiation, the LDB1-dependent repressed transcription units appear to be regulated primarily at the level of promoter pausing, with LDB1 regulating recruitment of metastasis-associated 1 family, member 2, a component of the nucleosome remodeling deacetylase complex, on these negative enhancers, required for the repressive enhancer function. These results indicate that LDB1-dependent looping events can deliver repressive cargo to cognate promoters to mediate promoter pausing events in a pituitary cell type.

scholarly journals Target Gene Specificity of E2F and Pocket Protein Family Members in Living Cells

2000 ◽  
Vol 20 (16) ◽  
pp. 5797-5807 ◽  
Author(s):  
Julie Wells ◽  
Kathryn E. Boyd ◽  
Christopher J. Fry ◽  
Stephanie M. Bartley ◽  
Peggy J. Farnham
Keyword(s):  
Cell Cycle ◽  
Target Gene ◽  
Target Genes ◽  
Protein Complexes ◽  
Current Model ◽  
Family Members ◽  
Living Cells ◽  
Gene Promoters ◽  
Pocket Protein ◽  
Protein Dna Interactions

ABSTRACT E2F-mediated transcription is thought to involve binding of an E2F-pocket protein complex to promoters in the G0 phase of the cell cycle and release of the pocket protein in late G1, followed by release of E2F in S phase. We have tested this model by monitoring protein-DNA interactions in living cells using a formaldehyde cross-linking and immunoprecipitation assay. We find that E2F target genes are bound by distinct E2F-pocket protein complexes which change as cells progress through the cell cycle. We also find that certain E2F target gene promoters are bound by pocket proteins when such promoters are transcriptionally active. Our data indicate that the current model applies only to certain E2F target genes and suggest that Rb family members may regulate transcription in both G0 and S phases. Finally, we find that a given promoter can be bound by one of several different E2F-pocket protein complexes at a given time in the cell cycle, suggesting that cell cycle-regulated transcription is a stochastic, not a predetermined, process.

scholarly journals Regulation of Poly(ADP-ribose) Polymerase-1-dependent Gene Expression through Promoter-directed Recruitment of a Nuclear NAD+ Synthase

2012 ◽  
Vol 287 (15) ◽  
pp. 12405-12416 ◽  
Author(s):  
Tong Zhang ◽  
Jhoanna G. Berrocal ◽  
Jie Yao ◽  
Michelle E. DuMond ◽  
Raga Krishnakumar ◽  
...  
Keyword(s):  
Enzymatic Activity ◽  
Target Gene ◽  
Target Genes ◽  
Enzymatic Activities ◽  
Gene Promoters ◽  
Protein Coding ◽  
Photon Excitation ◽  
Gene Sets ◽  
Cellular Compartments ◽  
Parp 1

NMNAT-1 and PARP-1, two key enzymes in the NAD+ metabolic pathway, localize to the nucleus where integration of their enzymatic activities has the potential to control a variety of nuclear processes. Using a variety of biochemical, molecular, cell-based, and genomic assays, we show that NMNAT-1 and PARP-1 physically and functionally interact at target gene promoters in MCF-7 cells. Specifically, we show that PARP-1 recruits NMNAT-1 to promoters where it produces NAD+ to support PARP-1 catalytic activity, but also enhances the enzymatic activity of PARP-1 independently of NAD+ production. Furthermore, using two-photon excitation microscopy, we show that NMNAT-1 catalyzes the production of NAD+ in a nuclear pool that may be distinct from other cellular compartments. In expression microarray experiments, depletion of NMNAT-1 or PARP-1 alters the expression of about 200 protein-coding genes each, with about 10% overlap between the two gene sets. NMNAT-1 enzymatic activity is required for PARP-1-dependent poly(ADP-ribosyl)ation at the promoters of commonly regulated target genes, as well as the expression of those target genes. Collectively, our studies link the enzymatic activities of NMNAT-1 and PARP-1 to the regulation of a set of common target genes through functional interactions at target gene promoters.

scholarly journals The Herpesvirus Saimiri Replication and Transcription Activator Acts Synergistically with CCAAT Enhancer Binding Protein Alpha To Activate the DNA Polymerase Promoter

2005 ◽  
Vol 79 (21) ◽  
pp. 13548-13560 ◽  
Author(s):  
Louise Wakenshaw ◽  
Matthew S. Walters ◽  
Adrian Whitehouse
Keyword(s):  
Dna Polymerase ◽  
Binding Protein ◽  
Lytic Replication ◽  
Early Gene ◽  
Herpesvirus Saimiri ◽  
Sequence Motif ◽  
Gene Promoters ◽  
Transcription Activator ◽  
Enhancer Binding Protein ◽  
Ccaat Enhancer Binding Protein

ABSTRACT The open reading frame (ORF) 50 gene product, also known as the replication and transcription activator (Rta), is an immediate-early gene which is well conserved among all gamma-2 herpesviruses and plays a pivotal role in regulating the latent-lytic switch. Herpesvirus saimiri (HVS) ORF 50a functions as a sequence-specific transactivator capable of activating delayed-early (DE) gene expression via binding directly to an ORF 50 response element (RE) within the respective promoter. Analysis of the ORF 50 REs have identified two distinct types within HVS gene promoters. The first comprises a consensus sequence motif, CCN9GG, the second an AT-rich sequence. Here we demonstrate that ORF 50a is capable of transactivating the DE ORF 9 promoter which encodes the DNA polymerase. Deletion analysis of the ORF 9 promoter mapped the ORF 50 RE to a 95-bp region situated 126 bp upstream of the initiation codon. Gel retardation analysis further mapped the RE to a 28-bp fragment, which was able to confer ORF 50 responsiveness on an enhancerless simian virus 40 minimal promoter. Furthermore, sequence analysis identified multiple CCAAT enhancer binding protein alpha (C/EBPα) binding sites within the ORF 9 promoter and specifically two within the close vicinity of the AT-rich ORF 50 RE. Analysis demonstrated that the HVS ORF 50a and C/EBPα proteins associate with the ORF 9 promoter in vivo, interact directly, and synergistically activate the ORF 9 promoter by binding to adjacent binding motifs. Overall, these data suggest a cooperative interaction between HVS ORF 50a and C/EBPα proteins to activate the DNA polymerase promoter during early stages of the lytic replication cycle.

scholarly journals Coactivation of Estrogen Receptor β by Gonadotropin-Induced Cofactor GIOT-4

2008 ◽  
Vol 29 (1) ◽  
pp. 83-92 ◽  
Author(s):  
Madoka Kouzu-Fujita ◽  
Yoshihiro Mezaki ◽  
Shun Sawatsubashi ◽  
Takahiro Matsumoto ◽  
Ikuko Yamaoka ◽  
...  
Keyword(s):  
Target Gene ◽  
Target Genes ◽  
Gene Promoters ◽  
Ovulatory Cycle ◽  
Independent Manner ◽  
Ovarian Granulosa Cell ◽  
Transcriptional Regulatory Mechanism ◽  
Transcriptional Regulatory ◽  
Specific Stage ◽  
Regulatory Convergence

ABSTRACT Estrogen exerts its diverse effects through two subtypes of estrogen receptors (ER), ERα and ERβ. Each subtype has its own distinct function and expression pattern in its target tissues. Little, however, is known about the transcriptional regulatory mechanism of ERβ in the major ERβ-expressing tissues. Using biochemical methods, we identified and described a novel ERβ coactivator. This protein, designated GIOT-4, was biochemically purified from 293F cells. It coactivated ERβ in ovarian granulosa cells. GIOT-4 expression was induced by stimulation with follicle-stimulating hormone (FSH). GIOT-4 recruited an SWI/SNF-type complex in a ligand-independent manner to ERβ as an ER subtype-specific physical bridging factor and induced subsequent histone modifications in the ERβ target gene promoters in a human ovarian granulosa cell line (KGN). Indeed, two ERβ-specific target genes were upregulated by FSH at a specific stage of a normal ovulatory cycle in intact mice. These findings imply the presence of a novel regulatory convergence between the gonadotropin signaling cascade and ERβ-mediated transcription in the ovary.

scholarly journals A PIT-1 Homeodomain Mutant Blocks the Intranuclear Recruitment Of the CCAAT/Enhancer Binding Protein α Required for Prolactin Gene Transcription

2003 ◽  
Vol 17 (2) ◽  
pp. 209-222 ◽  
Author(s):  
John F. Enwright ◽  
Margaret A. Kawecki-Crook ◽  
Ty C. Voss ◽  
Fred Schaufele ◽  
Richard N. Day
Keyword(s):  
Gene Transcription ◽  
Binding Protein ◽  
Hormone Deficiency ◽  
Centromeric Heterochromatin ◽  
Pituitary Hormone ◽  
Homeodomain Protein ◽  
Pituitary Cells ◽  
Enhancer Binding Protein ◽  
Ccaat Enhancer Binding Protein ◽  
Prl Gene

Abstract The pituitary-specific homeodomain protein Pit-1 cooperates with other transcription factors, including CCAAT/enhancer binding protein α (C/EBPα), in the regulation of pituitary lactotrope gene transcription. Here, we correlate cooperative activation of prolactin (PRL) gene transcription by Pit-1 and C/EBPα with changes in the subnuclear localization of these factors in living pituitary cells. Transiently expressed C/EBPα induced PRL gene transcription in pituitary GHFT1–5 cells, whereas the coexpression of Pit-1 and C/EBPα in HeLa cells demonstrated their cooperativity at the PRL promoter. Individually expressed Pit-1 or C/EBPα, fused to color variants of fluorescent proteins, occupied different subnuclear compartments in living pituitary cells. When coexpressed, Pit-1 recruited C/EBPα from regions of transcriptionally quiescent centromeric heterochromatin to the nuclear regions occupied by Pit-1. The homeodomain region of Pit-1 was necessary for the recruitment of C/EBPα. A point mutation in the Pit-1 homeodomain associated with the syndrome of combined pituitary hormone deficiency in humans also failed to recruit C/EBPα. This Pit-1 mutant functioned as a dominant inhibitor of PRL gene transcription and, instead of recruiting C/EBPα, was itself recruited by C/EBPα to centromeric heterochromatin. Together our results suggest that the intranuclear positioning of these factors determines whether they activate or silence PRL promoter activity.

scholarly journals EAGLE: an algorithm that utilizes a small number of genomic features to predict tissue/cell type-specific enhancer-gene interactions

2019 ◽  
Author(s):  
Tianshun Gao ◽  
Jiang Qian
Keyword(s):  
Target Gene ◽  
Target Genes ◽  
Cross Validation ◽  
Cell Types ◽  
Gene Interactions ◽  
Tissue Cell ◽  
Cell Type ◽  
Genomic Features ◽  
Link Type ◽  
Cell Type Specific

AbstractLong-range regulation by distal enhancers is crucial for many biological processes. The existing methods for enhancer-target gene prediction often require many genomic features. This makes them difficult to be applied to many cell types, in which the relevant datasets are not always available. Here, we design a tool EAGLE, an enhancer and gene learning ensemble method for identification of Enhancer-Gene (EG) interactions. Unlike existing tools, EAGLE used only six features derived from the genomic features of enhancers and gene expression datasets. Cross-validation revealed that EAGLE outperformed other existing methods. Enrichment analyses on special transcriptional factors, epigenetic modifications, and eQTLs demonstrated that EAGLE could distinguish the interacting pairs from non- interacting ones. Finally, EAGLE was applied to mouse and human genomes and identified 7,680,203 and 7,437,255 EG interactions involving 31,375 and 43,724 genes, 138,547 and 177,062 enhancers across 89 and 110 tissue/cell types in mouse and human, respectively. The obtained interactions are accessible through an interactive database enhanceratlas.org. The EAGLE method is available at https://github.com/EvansGao/EAGLE and the predicted datasets are available in http://www.enhanceratlas.org/.Author summaryEnhancers are DNA sequences that interact with promoters and activate target genes. Since enhancers often located far from the target genes and the nearest genes are not always the targets of the enhancers, the prediction of enhancer-target gene relationships is a big challenge. Although a few computational tools are designed for the prediction of enhancer-target genes, it’s difficult to apply them in most tissue/cell types due to a lack of enough genomic datasets. Here we proposed a new method, EAGLE, which utilizes a small number of genomic features to predict tissue/cell type-specific enhancer-gene interactions. Comparing with other existing tools, EAGLE displayed a better performance in the 10-fold cross-validation and cross-sample test. Moreover, the predictions by EAGLE were validated by other independent evidence such as the enrichment of relevant transcriptional factors, epigenetic modifications, and eQTLs.Finally, we integrated the enhancer-target relationships obtained from human and mouse genomes into an interactive database EnhancerAtlas, http://www.enhanceratlas.org/.

scholarly journals Effective CRISPR interference of an endogenous gene via a single transgene in mice

2019 ◽  
Vol 9 (1) ◽  
Author(s):  
Ryan S. MacLeod ◽  
Keisha M. Cawley ◽  
Igor Gubrij ◽  
Intawat Nookaew ◽  
Melda Onal ◽  
...  
Keyword(s):  
Transgene Expression ◽  
Target Gene ◽  
Gene Deletion ◽  
Target Genes ◽  
Intermediate Phenotype ◽  
Mrna Levels ◽  
Loss Of Function ◽  
Cell Type ◽  
High Bone Mass ◽  
Crispr Interference

AbstractDrawbacks of conditional gene deletion in mice include the need for extensive breeding and, often, a lack of cell type-specificity. CRISPR interference (CRISPRi) is an alternative approach for loss-of-function studies that inhibits expression by guiding a transcriptional repressor to the transcription start-site of target genes. However, there has been limited exploration of CRISPRi in mice. We tested the effectiveness of a single CRISPRi transgene broadly expressing a single guide RNA and a catalytically dead Cas9 fused to the KRAB repressor domain to suppress a well-characterized target gene, Tnfsf11. The phenotype of CRISPRi transgenic mice was compared to mice with germline deletion of Tnfsf11, which are osteopetrotic and do not form lymph nodes. High transgene expression mimicked gene deletion, with failure of lymph node development and classic signs of osteopetrosis such as high bone mass and failure of tooth eruption. Mice with low transgene expression were normal and mice with medium expression displayed an intermediate phenotype. Transgene expression in tissues from these mice correlated inversely with Tnfsf11 mRNA levels. These results demonstrate that a single CRISPRi transgene can effectively suppress a target gene in mice and suggest that this approach may be useful for cell type-specific loss-of-function studies.

scholarly journals Identification of a TAL1 Target Gene Reveals a Positive Role for the LIM Domain-Binding Protein Ldb1 in Erythroid Gene Expression and Differentiation

2003 ◽  
Vol 23 (21) ◽  
pp. 7585-7599 ◽  
Author(s):  
Zhixiong Xu ◽  
Suming Huang ◽  
Long-Sheng Chang ◽  
Alan D. Agulnick ◽  
Stephen J. Brandt
Keyword(s):  
Gene Expression ◽  
Transcription Factor ◽  
Dna Binding ◽  
Target Genes ◽  
Binding Protein ◽  
Proximal Promoter ◽  
Lim Domain ◽  
Positive Role ◽  
E Box ◽  
Murine Erythroleukemia

ABSTRACT The TAL1 (or SCL) gene, originally identified from its involvement by a recurrent chromosomal translocation, encodes a basic helix-loop-helix transcription factor essential for erythropoiesis. Although presumed to regulate transcription, its target genes are largely unknown. We show here that a nuclear complex containing TAL1, its DNA-binding partner E47, zinc finger transcription factor GATA-1, LIM domain protein LMO2, and LIM domain-binding protein Ldb1 transactivates the protein 4.2 (P4.2) gene through two E box GATA elements in its proximal promoter. Binding of this complex to DNA was dependent on the integrity of both E box and GATA sites and was demonstrated to occur on the P4.2 promoter in cells. Maximal transcription in transiently transfected cells required both E box GATA elements and expression of all five components of the complex. This complex was shown, in addition, to be capable of linking in solution double-stranded oligonucleotides corresponding to the two P4.2 E box GATA elements. This DNA-linking activity required Ldb1 and increased with dimethyl sulfoxide-induced differentiation of murine erythroleukemia (MEL) cells. In contrast, enforced expression in MEL cells of dimerization-defective mutant Ldb1, as well as wild-type Ldb1, significantly decreased E box GATA DNA-binding activities, P4.2 promoter activity, and accumulation of P4.2 and β-globin mRNAs. These studies define a physiologic target for a TAL1- and GATA-1-containing ternary complex and reveal a positive role for Ldb1 in erythroid gene expression and differentiation.

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