Master transcription factors form interconnected circuitry and orchestrate transcriptional networks in oesophageal adenocarcinoma

ObjectiveWhile oesophageal squamous cell carcinoma remains infrequent in Western populations, the incidence of oesophageal adenocarcinoma (EAC) has increased sixfold to eightfold over the past four decades. We aimed to characterise oesophageal cancer-specific and subtypes-specific gene regulation patterns and their upstream transcription factors (TFs). DesignTo identify regulatory elements, we profiled fresh-frozen oesophageal normal samples, tumours and cell lines with chromatin immunoprecipitation sequencing (ChIP-Seq). Mathematical modelling was performed to establish (super)-enhancers landscapes and interconnected transcriptional circuitry formed by master TFs. Coregulation and cooperation between master TFs were investigated by ChIP-Seq, circularised chromosome conformation capture sequencing and luciferase assay. Biological functions of candidate factors were evaluated both in vitro and in vivo.ResultsWe found widespread and pervasive alterations of the (super)-enhancer reservoir in both subtypes of oesophageal cancer, leading to transcriptional activation of a myriad of novel oncogenes and signalling pathways, some of which may be exploited pharmacologically (eg, leukemia inhibitory factor (LIF) pathway). Focusing on EAC, we bioinformatically reconstructed and functionally validated an interconnected circuitry formed by four master TFs—ELF3, KLF5, GATA6 and EHF—which promoted each other’s expression by interacting with each super-enhancer. Downstream, these master TFs occupied almost all EAC super-enhancers and cooperatively orchestrated EAC transcriptome. Each TF within the transcriptional circuitry was highly and specifically expressed in EAC and functionally promoted EAC cell proliferation and survival.ConclusionsBy establishing cancer-specific and subtype-specific features of the EAC epigenome, our findings promise to transform understanding of the transcriptional dysregulation and addiction of EAC, while providing molecular clues to develop novel therapeutic modalities against this malignancy.

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Participation of the yeast activator Abf1 in meiosis-specific expression of the HOP1 gene.

Molecular and Cellular Biology ◽

10.1128/mcb.16.6.2777 ◽

1996 ◽

Vol 16 (6) ◽

pp. 2777-2786 ◽

Cited By ~ 24

Author(s):

V Gailus-Durner ◽

J Xie ◽

C Chintamaneni ◽

A K Vershon

Keyword(s):

Transcriptional Activation ◽

Mutational Analysis ◽

Consensus Sequence ◽

Promoter Sequence ◽

Regulatory Elements ◽

Specific Gene ◽

Specific Expression ◽

Autonomously Replicating Sequence

The meiosis-specific gene HOP1, which encodes a component of the synaptonemal complex, is controlled through two regulatory elements, UASH and URS1H. Sites similar to URS1H have been identified in the promoter region of virtually every early meiosis-specific gene, as well as in many promoters of nonmeiotic genes, and it has been shown that the proteins that bind to this site function to regulate meiotic and nonmeiotic transcription. Sites similar to the UASH site have been found in a number of meiotic and nonmeiotic genes as well. Since it has been shown that UASH functions as an activator site in vegetative haploid cells, it seemed likely that the factors binding to this site regulate both meiotic and nonmeiotic transcription. We purified the factor binding to the UASH element of the HOP1 promoter. Sequence analysis identified the protein as Abf1 (autonomously replicating sequence-binding factor 1), a multifunctional protein involved in DNA replication, silencing, and transcriptional regulation. We show by mutational analysis of the UASH site, that positions outside of the proposed UASH consensus sequence (TNTGN[A/T]GT) are required for DNA binding in vitro and transcriptional activation in vivo. A new UASH consensus sequence derived from this mutational analysis closely matches a consensus Abf1 binding site. We also show that an Abf1 site from a nonmeiotic gene can replace the function of the UASH site in the HOP1 promoter. Taken together, these results show that Abf1 functions to regulate meiotic gene expression.

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HMGN1 Modulates Estrogen-Mediated Transcriptional Activation through Interactions with Specific DNA-Binding Transcription Factors

Molecular and Cellular Biology ◽

10.1128/mcb.01724-07 ◽

2007 ◽

Vol 27 (24) ◽

pp. 8859-8873 ◽

Cited By ~ 10

Author(s):

Nan Zhu ◽

Ulla Hansen

Keyword(s):

Transcription Factors ◽

Protein Interactions ◽

Transcriptional Activation ◽

Serum Response Factor ◽

Estrogen Receptor Α ◽

Order Structure ◽

Specific Gene ◽

Estrogen Induction

ABSTRACT HMGN1, an abundant nucleosomal binding protein, can affect both the chromatin higher order structure and the modification of nucleosomal histones, but it alters the expression of only a subset of genes. We investigated specific gene targeting by HMGN1 in the context of estrogen induction of gene expression. Knockdown and overexpression experiments indicated that HMGN1 limits the induction of several estrogen-regulated genes, including TFF1 and FOS, which are induced by estrogen through entirely distinct mechanisms. HMGN1 specifically interacts with estrogen receptor α (ERα), both in vitro and in vivo. At the TFF1 promoter, estrogen increases HMGN1 association through recruitment by the ERα. HMGN1 S20E/S24E, although deficient in binding nucleosomal DNA, still interacts with ERα and, strikingly, still represses estrogen-driven activation of the TFF1 gene. On the FOS promoter, which lacks the ERα binding sites, constitutively bound serum response factor (SRF) mediates estrogen stimulation. HMGN1 also interacts specifically with SRF, but HMGN1 S20E/S24E does not. Consistent with the protein interactions, only wild-type HMGN1 significantly inhibits the estrogen-driven activation of the FOS gene. Mechanistically, the inhibition of estrogen induction of several ERα-associated genes, including TFF1, by HMGN1 correlates with decreased levels of acetylation of Lys9 on histone H3. Together, these findings indicate that HMGN1 regulates the expression of particular genes via specific protein-protein interactions with transcription factors at target gene regulatory regions.

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Subnuclear compartmentalization of sequence-specific transcription factors and regulation of eukaryotic gene expression

Biochemistry and Cell Biology ◽

10.1139/o05-062 ◽

2005 ◽

Vol 83 (4) ◽

pp. 535-547 ◽

Cited By ~ 7

Author(s):

Gareth N Corry ◽

D Alan Underhill

Keyword(s):

Gene Expression ◽

Transcription Factor ◽

Transcription Factors ◽

Transcriptional Activation ◽

Current Knowledge ◽

Nuclear Architecture ◽

Subnuclear Localization ◽

Modular Structures

To date, the majority of the research regarding eukaryotic transcription factors has focused on characterizing their function primarily through in vitro methods. These studies have revealed that transcription factors are essentially modular structures, containing separate regions that participate in such activities as DNA binding, protein–protein interaction, and transcriptional activation or repression. To fully comprehend the behavior of a given transcription factor, however, these domains must be analyzed in the context of the entire protein, and in certain cases the context of a multiprotein complex. Furthermore, it must be appreciated that transcription factors function in the nucleus, where they must contend with a variety of factors, including the nuclear architecture, chromatin domains, chromosome territories, and cell-cycle-associated processes. Recent examinations of transcription factors in the nucleus have clarified the behavior of these proteins in vivo and have increased our understanding of how gene expression is regulated in eukaryotes. Here, we review the current knowledge regarding sequence-specific transcription factor compartmentalization within the nucleus and discuss its impact on the regulation of such processes as activation or repression of gene expression and interaction with coregulatory factors.Key words: transcription, subnuclear localization, chromatin, gene expression, nuclear architecture.

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Expression of laminin γ 1 cultured hepatocytes involves repeated CTC and GC elements in the LAMC1 promoter

Biochemical Journal ◽

10.1042/bj3130745 ◽

1996 ◽

Vol 313 (3) ◽

pp. 745-752 ◽

Cited By ~ 5

Author(s):

Françoise LEVAVASSEUR ◽

Jocelyne LIÉTARD ◽

Kohei OGAWA ◽

Nathalie THÉRET ◽

Peter D. BURBELO ◽

...

Keyword(s):

Transcriptional Activation ◽

Hepatoma Cell ◽

Primary Cultures ◽

Regulatory Elements ◽

Hepatoma Cells ◽

Basement Membranes ◽

Major Protein ◽

Cultured Hepatocytes

Laminin γ1 chain is present in all basement membranes and is expressed at high levels in various diseases, such as hepatic fibrosis. We have identified cis- and trans-acting elements involved in the regulation of this gene in normal rat liver, as well as in hepatocyte primary cultures and hepatoma cell lines. Northern-blot analyses showed that laminin γ1 mRNA was barely detectable in freshly isolated hepatocytes and expressed at high levels in hepatocyte primary cultures, as early as 4 h after liver dissociation. Actinomycin D and cycloheximide treatment in vivo and in vitro indicated that laminin γ1 overexpression in cultured hepatocytes was under the control of transcriptional mechanisms. Transfection of deletion mutants of the 5´ flanking region of murine LAMC1 gene in hepatoma cells that constitutively express laminin γ1 indicated that regulatory elements were located between -594 bp and -94 bp. This segment included GC- and CTC-containing motifs. Gel-shift analyses showed that two complexes were resolved with different affinity for the CTC sequence depending on the location of the GC box. The pattern of complex formation with nuclear factors from freshly isolated and cultured hepatocytes was different from that obtained with total liver and similar to that with hepatoma cells. Southwestern analysis indicated that several polypeptides bound the CTC-rich sequence. Affinity chromatography demonstrated that a Mr 60000 polypeptide was a major protein binding to the CTC motif. This polypeptide is probably involved in the transcriptional activation of various proto-oncogenes and extracellular matrix genes that are expressed at high levels in both hepatoma cells and early hepatocyte cultures.

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Targeting of p38 Mitogen-Activated Protein Kinases to MEF2 Transcription Factors

Molecular and Cellular Biology ◽

10.1128/mcb.19.6.4028 ◽

1999 ◽

Vol 19 (6) ◽

pp. 4028-4038 ◽

Cited By ~ 172

Author(s):

Shen-Hsi Yang ◽

Alex Galanis ◽

Andrew D. Sharrocks

Keyword(s):

Transcription Factors ◽

Transcriptional Activation ◽

Cellular Response ◽

Map Kinases ◽

Biological Response ◽

Extracellular Signals ◽

Extracellular Signal ◽

Mitogen Activated Protein

ABSTRACT Mitogen-activated protein (MAP) kinase-mediated signalling to the nucleus is an important event in the conversion of extracellular signals into a cellular response. However, the existence of multiple MAP kinases which phosphorylate similar phosphoacceptor motifs poses a problem in maintaining substrate specificity and hence the correct biological response. Both the extracellular signal-regulated kinase (ERK) and c-Jun NH2-terminal kinase (JNK) subfamilies of MAP kinases use a second specificity determinant and require docking to their transcription factor substrates to achieve maximal substrate activation. In this study, we demonstrate that among the different MAP kinases, the MADS-box transcription factors MEF2A and MEF2C are preferentially phosphorylated and activated by the p38 subfamily members p38α and p38β2. The efficiency of phosphorylation in vitro and transcriptional activation in vivo of MEF2A and MEF2C by these p38 subtypes requires the presence of a kinase docking domain (D-domain). Furthermore, the D-domain from MEF2A is sufficient to confer p38 responsiveness on different transcription factors, and reciprocal effects are observed upon the introduction of alternative D-domains into MEF2A. These results therefore contribute to our understanding of signalling to MEF2 transcription factors and demonstrate that the requirement for substrate binding by MAP kinases is an important facet of three different subclasses of MAP kinases (ERK, JNK, and p38).

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Activating Signal Cointegrator 2 Belongs to a Novel Steady-State Complex That Contains a Subset of Trithorax Group Proteins

Molecular and Cellular Biology ◽

10.1128/mcb.23.1.140-149.2003 ◽

2003 ◽

Vol 23 (1) ◽

pp. 140-149 ◽

Cited By ~ 158

Author(s):

Young-Hwa Goo ◽

Young Chang Sohn ◽

Dae-Hwan Kim ◽

Seung-Whan Kim ◽

Min-Jung Kang ◽

...

Keyword(s):

Transcription Factors ◽

Steady State ◽

Nuclear Receptors ◽

Transcriptional Activation ◽

Retinoic Acid Receptor ◽

Dependent Manner ◽

Trithorax Group ◽

Trithorax Group Proteins

ABSTRACT Many transcription coactivators interact with nuclear receptors in a ligand- and C-terminal transactivation function (AF2)-dependent manner. These include activating signal cointegrator 2 (ASC-2), a recently isolated transcriptional coactivator molecule, which is amplified in human cancers and stimulates transactivation by nuclear receptors and numerous other transcription factors. In this report, we show that ASC-2 belongs to a steady-state complex of approximately 2 MDa (ASC-2 complex [ASCOM]) in HeLa nuclei. ASCOM contains retinoblastoma-binding protein RBQ-3, α/β-tubulins, and trithorax group proteins ALR-1, ALR-2, HALR, and ASH2. In particular, ALR-1/2 and HALR contain a highly conserved 130- to 140-amino-acid motif termed the SET domain, which was recently implicated in histone H3 lysine-specific methylation activities. Indeed, recombinant ALR-1, HALR, and immunopurified ASCOM exhibit very weak but specific H3-lysine 4 methylation activities in vitro, and transactivation by retinoic acid receptor appears to involve ligand-dependent recruitment of ASCOM and subsequent transient H3-lysine 4 methylation of the promoter region in vivo. Thus, ASCOM may represent a distinct coactivator complex of nuclear receptors. Further characterization of ASCOM will lead to a better understanding of how nuclear receptors and other transcription factors mediate transcriptional activation.

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Cyclic AMP-dependent protein kinase inhibits the activity of myogenic helix-loop-helix proteins.

Molecular and Cellular Biology ◽

10.1128/mcb.12.10.4478 ◽

1992 ◽

Vol 12 (10) ◽

pp. 4478-4485 ◽

Cited By ~ 68

Author(s):

L Li ◽

R Heller-Harrison ◽

M Czech ◽

E N Olson

Keyword(s):

Protein Kinase ◽

Cyclic Amp ◽

Transcriptional Activation ◽

Consensus Sequence ◽

Signal Transduction Pathway ◽

Specific Gene ◽

Dependent Protein Kinase ◽

Dependent Protein

Differentiation of skeletal muscle cells is inhibited by the cyclic AMP (cAMP) signal transduction pathway. Here we report that the catalytic subunit of cAMP-dependent protein kinase (PKA) can substitute for cAMP and suppress muscle-specific transcription by silencing the activity of the MyoD family of regulatory factors, which includes MyoD, myogenin, myf5, and MRF4. Repression by the PKA catalytic (C) subunit is directed at the consensus sequence CANNTG, the target for DNA binding and transcriptional activation by these myogenic regulators. Phosphopeptide mapping of myogenin in vitro and in vivo revealed two PKA phosphorylation sites, both within the basic region. However, repression of myogenin function by PKA does not require direct phosphorylation of these sites but instead involves an indirect mechanism with one or more intermediate steps. Regulation of the transcriptional activity of the MyoD family by modulation of the cAMP signaling pathway may account for the inhibitory effects of certain peptide growth factors on muscle-specific gene expression and may also determine the responsiveness of different cell types to myogenic conversion by these myogenic regulators.

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Cyclic AMP-dependent protein kinase inhibits the activity of myogenic helix-loop-helix proteins

Molecular and Cellular Biology ◽

10.1128/mcb.12.10.4478-4485.1992 ◽

1992 ◽

Vol 12 (10) ◽

pp. 4478-4485

Author(s):

L Li ◽

R Heller-Harrison ◽

M Czech ◽

E N Olson

Keyword(s):

Protein Kinase ◽

Cyclic Amp ◽

Transcriptional Activation ◽

Consensus Sequence ◽

Signal Transduction Pathway ◽

Specific Gene ◽

Dependent Protein Kinase ◽

Dependent Protein

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Meloxicam, a Selective COX-2 Inhibitor, Mediates Hypoxia-Inducible Factor- (HIF-) 1α Signaling in Hepatocellular Carcinoma

Oxidative Medicine and Cellular Longevity ◽

10.1155/2020/7079308 ◽

2020 ◽

Vol 2020 ◽

pp. 1-15

Author(s):

Yinghong Zhou ◽

Xiaofeng Dong ◽

Peng Xiu ◽

Xin Wang ◽

Jianrong Yang ◽

...

Keyword(s):

Hepatocellular Carcinoma ◽

Membrane Potential ◽

Mitochondrial Membrane Potential ◽

Transcriptional Activation ◽

Mitochondrial Membrane ◽

Hypoxia Inducible Factor ◽

Luciferase Assay ◽

Cox 2

Hepatocellular carcinoma (HCC) is regarded as a leading cause of cancer-related deaths, and its progression is associated with hypoxia and the induction of hypoxia-inducible factor (HIF). Meloxicam, a selective cyclooxygenase-2 (COX-2) inhibitor, induces cell death in various malignancies. However, the underlying mechanism remains to be elucidated in HCC, especially under hypoxic conditions. The alteration of COX-2 and HIF-1α oncogenicity was evaluated in HCC specimens by tissue microarray. Cell viability, angiogenesis assays, and xenografted nude mice were used to evaluate the effects of meloxicam, along with flow cytometry to detect the cell cycle, apoptosis, and mitochondrial membrane potential (ΔΨm) of HCC. qRT-PCR, Western blotting, immunofluorescence, immunohistochemistry, luciferase assay, and RNAi were carried out to determine the HIF-1α signaling affected by meloxicam. In this study, we showed that meloxicam exerts antiproliferative and antiangiogenesis efficacy in vitro and in vivo and causes disruption of mitochondrial membrane potential (ΔΨm), thus leading to caspase-dependent apoptosis under hypoxic environments. Exposure to meloxicam significantly reduced HIF-1α transcriptional activation and expression through sequestering it in the cytoplasm and accelerating degradation via increasing the von Hippel-Lindau tumor suppressor protein (pVHL) in HCC. These data demonstrated that inhibition of HIF-1α by meloxicam could suppress angiogenesis and enhance apoptosis of HCC cells. This discovery highlights that COX-2 specific inhibitors may be a promising therapy in the treatment of HCC.

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Pmp22 super-enhancer deletion causes tomacula formation and conduction block in peripheral nerves

Human Molecular Genetics ◽

10.1093/hmg/ddaa082 ◽

2020 ◽

Vol 29 (10) ◽

pp. 1689-1699

Author(s):

Harrison Pantera ◽

Bo Hu ◽

Daniel Moiseev ◽

Chris Dunham ◽

Jibraan Rashid ◽

...

Keyword(s):

Peripheral Neuropathy ◽

Schwann Cell ◽

Copy Number ◽

Regulatory Elements ◽

Copy Number Variations ◽

Hereditary Neuropathy ◽

Peripheral Myelin Protein 22 ◽

Super Enhancer

Abstract Copy number variation of the peripheral nerve myelin gene Peripheral Myelin Protein 22 (PMP22) causes multiple forms of inherited peripheral neuropathy. The duplication of a 1.4 Mb segment surrounding this gene in chromosome 17p12 (c17p12) causes the most common form of Charcot-Marie-Tooth disease type 1A, whereas the reciprocal deletion of this gene causes a separate neuropathy termed hereditary neuropathy with liability to pressure palsies (HNPP). PMP22 is robustly induced in Schwann cells in early postnatal development, and several transcription factors and their cognate regulatory elements have been implicated in coordinating the gene’s proper expression. We previously found that a distal super-enhancer domain was important for Pmp22 expression in vitro, with particular impact on a Schwann cell-specific alternative promoter. Here, we investigate the consequences of deleting this super-enhancer in vivo. We find that loss of the super-enhancer in mice reduces Pmp22 expression throughout development and into adulthood, with greater impact on the Schwann cell-specific promoter. Additionally, these mice display tomacula formed by excessive myelin folding, a pathological hallmark of HNPP, as have been previously observed in heterozygous Pmp22 mice as well as sural biopsies from patients with HNPP. Our findings demonstrate a mechanism by which smaller copy number variations, not including the Pmp22 gene, are sufficient to reduce gene expression and phenocopy a peripheral neuropathy caused by the HNPP-associated deletion encompassing PMP22.

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