Oxygen-regulated erythropoietin gene expression is dependent on a CpG methylation-free hypoxia-inducible factor-1 DNA-binding site

1998 ◽  
Vol 253 (3) ◽  
pp. 771-777 ◽  
Author(s):  
Roland H. Wenger ◽  
Ivica Kvietikova ◽  
Andreas Rolfs ◽  
Gieri Camenisch ◽  
Max Gassmann
Keyword(s):  
Gene Expression ◽  
Dna Binding ◽  
Binding Site ◽  
Hypoxia Inducible Factor ◽  
Cpg Methylation ◽  
Hypoxia Inducible Factor 1 ◽  
Dna Binding Site

scholarly journals Hesx1 homeodomain protein represses transcription as a monomer and antagonises transactivation of specific sites as a homodimer

2002 ◽  
Vol 28 (3) ◽  
pp. 193-205 ◽  
Author(s):  
J Quirk ◽  
P Brown
Keyword(s):  
Gene Expression ◽  
Dna Binding ◽  
Binding Site ◽  
Anterior Pituitary ◽  
Homeodomain Protein ◽  
Dependent Manner ◽  
Gene Promoters ◽  
Differentiation Markers ◽  
Proximal Half ◽  
Dna Binding Site

The homeobox repressor Hesx1, expressed throughout Rathke's pouch and required for normal pituitary development, has been implicated in anterior pituitary pathogenesis in man. Prolonged expression of Hesx1 delays the appearance of anterior pituitary terminal differentiation markers in mice, particularly the gonadotroph hormones. We tested if Hesx1 could modulate gonadotrophin gene expression directly, and found that Hesx1 repressed both common alpha subunit (alpha GSU) and luteinising hormone beta-subunit (LH beta) gene promoters. Repression mapped to the Pitx1 homeodomain protein transactivation site in the proximal alpha GSU promoter, but did not map to the equivalent site on LH beta. Hesx1 repression of the alpha GSU Pitx1 site was overridden by co-transfection of Pitx1. In contrast, Hesx1 antagonised Pitx1 transactivation of LH beta in a dose-dependent manner. This was due to monomeric binding of Hesx1 on alpha GSU and homodimerisation on LH beta. The homodimerisation site comprises the Pitx1 DNA binding site and a proximal binding site, and mutation of either inhibited homodimer formation. Conversion of the LH beta Pitx1 DNA binding site to an alpha GSU-type did not promote homodimer formation, arguing that Hesx1 has pronounced site selectivity. Furthermore, mutation of the proximal half of the homodimerisation site blocked Hesx1 antagonisation of Pitx1 transactivation. We conclude that Hesx1 monomers repress gene expression, and homodimers block specific transactivation sites.

scholarly journals Crystal Structure of the DNA-Binding Domain of Human Herpesvirus 6A Immediate Early Protein 2

2017 ◽  
Vol 91 (21) ◽  
Author(s):  
Mitsuhiro Nishimura ◽  
Junjie Wang ◽  
Aika Wakata ◽  
Kento Sakamoto ◽  
Yasuko Mori
Keyword(s):  
Gene Expression ◽  
Dna Binding ◽  
Binding Site ◽  
Human Herpesvirus ◽  
Binding Domain ◽  
Immediate Early ◽  
Dna Binding Domain ◽  
Factor Binding ◽  
Early Protein ◽  
Dna Binding Site

ABSTRACT Immediate early proteins of human herpesvirus 6A (HHV-6A) are expressed at the outset of lytic infection and thereby regulate viral gene expression. Immediate early protein 2 (IE2) of HHV-6A is a transactivator that drives a variety of promoters. The C-terminal region of HHV-6A IE2 is shared among IE2 homologs in betaherpesviruses and is involved in dimerization, DNA binding, and transcription factor binding. In this study, the structure of the IE2 C-terminal domain (IE2-CTD) was determined by X-ray crystallography at a resolution of 2.5 Å. IE2-CTD forms a homodimer stabilized by a β-barrel core with two interchanging long loops. Unexpectedly, the core structure resembles those of the gammaherpesvirus factors EBNA1 of Epstein-Barr virus and LANA of Kaposi sarcoma-associated herpesvirus, but the interchanging loops are longer in IE2-CTD and form helix-turn-helix (HTH)-like motifs at their tips. The HTH and surrounding α-helices form a structural feature specific to the IE2 group. The apparent DNA-binding site (based on structural similarity with EBNA1 and LANA) resides on the opposite side of the HTH-like motifs, surrounded by positive electrostatic potential. Mapping analysis of conserved residues on the three-dimensional structure delineated a potential factor-binding site adjacent to the expected DNA-binding site. The predicted bi- or tripartite functional sites indicate a role for IE2-CTD as an adapter connecting the promoter and transcriptional factors that drive gene expression. IMPORTANCE Human herpesvirus 6A (HHV-6A) and HHV-6B belong to betaherpesvirus subfamily. Both viruses establish lifelong latency after primary infection, and their reactivation poses a significant risk to immunocompromised patients. Immediate early protein 2 (IE2) of HHV-6A and HHV-6B is a transactivator that triggers viral replication and contains a DNA-binding domain shared with other betaherpesviruses such as human herpesvirus 7 and human cytomegalovirus. In this study, an atomic structure of the DNA-binding domain of HHV-6A IE2 was determined and analyzed, enabling a structure-based understanding of the functions of IE2, specifically DNA recognition and interaction with transcription factors. Unexpectedly, the dimeric core resembles the DNA-binding domain of transcription regulators from gammaherpesviruses, showing structural conservation as a DNA-binding domain but with its own unique structural features. These findings facilitate further characterization of this key viral transactivator.

scholarly journals t(8;21)(q22;q22) fusion proteins preferentially bind to duplicated AML1/RUNX1 DNA-binding sequences to differentially regulate gene expression

Blood ◽  
2008 ◽  
Vol 112 (4) ◽  
pp. 1392-1401 ◽  
Author(s):  
Akiko J. Okumura ◽  
Luke F. Peterson ◽  
Fumihiko Okumura ◽  
Anita Boyapati ◽  
Dong-Er Zhang
Keyword(s):  
Gene Expression ◽  
Dna Binding ◽  
Binding Site ◽  
Dna Sequences ◽  
Fusion Proteins ◽  
Target Genes ◽  
Regulatory Elements ◽  
Chromosome Abnormalities ◽  
Cancer Development ◽  
Dna Binding Site

AbstractChromosome abnormalities are frequently associated with cancer development. The 8;21(q22;q22) chromosomal translocation is one of the most common chromosome abnormalities identified in leukemia. It generates fusion proteins between AML1 and ETO. Since AML1 is a well-defined DNA-binding protein, AML1-ETO fusion proteins have been recognized as DNA-binding proteins interacting with the same consensus DNA-binding site as AML1. The alteration of AML1 target gene expression due to the presence of AML1-ETO is related to the development of leukemia. Here, using a 25-bp random double-stranded oligonucleotide library and a polymerase chain reaction (PCR)-based DNA-binding site screen, we show that compared with native AML1, AML1-ETO fusion proteins preferentially bind to DNA sequences with duplicated AML1 consensus sites. This finding is further confirmed by both in vitro and in vivo DNA-protein interaction assays. These results suggest that AML1-ETO fusion proteins have a selective preference for certain AML1 target genes that contain multimerized AML1 consensus sites in their regulatory elements. Such selected regulation provides an important molecular mechanism for the dysregulation of gene expression during cancer development.

Interleukin-1β and Tumor Necrosis Factor- Stimulate DNA Binding of Hypoxia-Inducible Factor-1

Blood ◽  
1999 ◽  
Vol 94 (5) ◽  
pp. 1561-1567 ◽  
Author(s):  
Thomas Hellwig-Bürgel ◽  
Karen Rutkowski ◽  
Eric Metzen ◽  
Joachim Fandrey ◽  
Wolfgang Jelkmann
Keyword(s):  
Gene Expression ◽  
Tumor Necrosis Factor ◽  
Dna Binding ◽  
Tumor Necrosis ◽  
Hypoxia Inducible Factor ◽  
Interleukin 1Β ◽  
Mrna Levels ◽  
Human Hepatoma Cells ◽  
Hypoxia Inducible Factor 1 ◽  
Necrosis Factor

The rate of transcription of several genes encoding proteins involved in O2 and energy homeostasis is controlled by hypoxia-inducible factor-1 (HIF-1), a heterodimeric DNA binding complex composed of  and β subunits. HIF-1 is considered the primarytrans-acting factor for the erythropoietin (EPO) and vascular endothelial growth factor (VEGF) genes. Since EPO gene expression is inhibited by the proinflammatory cytokines interleukin-1β (IL-1β) and tumor necrosis factor- (TNF-), while no such effect has been reported with respect to the VEGF gene, we investigated the effects of IL-1β and TNF- on the activation of the HIF-1 DNA-binding complex and the amount of HIF-1 protein in human hepatoma cells in culture. Under normoxic conditions, both cytokines caused a moderate activation of HIF-1 DNA binding. In hypoxia, cytokines strongly increased HIF-1 activity compared with the effect of hypoxia alone. Only IL-1β increased HIF-1 protein levels. In transient transfection experiments, HIF-1–driven reporter gene expression was augmented by cytokines only under hypoxic conditions. In contrast to their effect on EPO synthesis, neither IL-1β nor TNF- decreased VEGF production. The mRNA levels of HIF-1 and VEGF were unaffected. Thus, cytokine-induced inhibition of EPO production is not mediated by impairment of HIF-1 function. We propose that HIF-1 may be involved in modulating gene expression during inflammation.

scholarly journals Selective DNA Binding and Association with the CREB Binding Protein Coactivator Contribute to Differential Activation of Alpha/Beta Interferon Genes by Interferon Regulatory Factors 3 and 7

2000 ◽  
Vol 20 (17) ◽  
pp. 6342-6353 ◽  
Author(s):  
Rongtuan Lin ◽  
Pierre Génin ◽  
Yaël Mamane ◽  
John Hiscott
Keyword(s):  
Gene Expression ◽  
Dna Binding ◽  
Binding Site ◽  
Binding Protein ◽  
Creb Binding Protein ◽  
Regulatory Factors ◽  
Transactivation Activity ◽  
Dna Binding Site ◽  
Alpha Beta ◽  
Selective Dna Binding

ABSTRACT Recent studies implicate the interferon (IFN) regulatory factors (IRF) IRF-3 and IRF-7 as key activators of the alpha/beta IFN (IFN-α/β) genes as well as the RANTES chemokine gene. Using coexpression analysis, the human IFNB, IFNA1, and RANTES promoters were stimulated by IRF-3 coexpression, whereas the IFNA4, IFNA7, and IFNA14 promoters were preferentially induced by IRF-7 only. Chimeric proteins containing combinations of different IRF-7 and IRF-3 domains were also tested, and the results provided evidence of distinct DNA binding properties of IRF-3 and IRF-7, as well as a preferential association of IRF-3 with the CREB binding protein (CBP) coactivator. Interestingly, some of these fusion proteins led to supraphysiological levels of IFN promoter activation. DNA binding site selection studies demonstrated that IRF-3 and IRF-7 bound to the 5′-GAAANNGAAANN-3′ consensus motif found in many virus-inducible genes; however, a single nucleotide substitution in either of the GAAA half-site motifs eliminated IRF-3 binding and transactivation activity but did not affect IRF-7 interaction or transactivation activity. These studies demonstrate that IRF-3 possesses a restricted DNA binding site specificity and interacts with CBP, whereas IRF-7 has a broader DNA binding specificity that contributes to its capacity to stimulate delayed-type IFN gene expression. These results provide an explanation for the differential regulation of IFN-α/β gene expression by IRF-3 and IRF-7 and suggest that these factors have complementary rather than redundant roles in the activation of the IFN-α/β genes.

Interleukin-1β and Tumor Necrosis Factor- Stimulate DNA Binding of Hypoxia-Inducible Factor-1

Blood ◽  
1999 ◽  
Vol 94 (5) ◽  
pp. 1561-1567 ◽  
Author(s):  
Thomas Hellwig-Bürgel ◽  
Karen Rutkowski ◽  
Eric Metzen ◽  
Joachim Fandrey ◽  
Wolfgang Jelkmann
Keyword(s):  
Gene Expression ◽  
Tumor Necrosis Factor ◽  
Dna Binding ◽  
Tumor Necrosis ◽  
Hypoxia Inducible Factor ◽  
Interleukin 1Β ◽  
Mrna Levels ◽  
Human Hepatoma Cells ◽  
Hypoxia Inducible Factor 1 ◽  
Necrosis Factor

Abstract The rate of transcription of several genes encoding proteins involved in O2 and energy homeostasis is controlled by hypoxia-inducible factor-1 (HIF-1), a heterodimeric DNA binding complex composed of  and β subunits. HIF-1 is considered the primarytrans-acting factor for the erythropoietin (EPO) and vascular endothelial growth factor (VEGF) genes. Since EPO gene expression is inhibited by the proinflammatory cytokines interleukin-1β (IL-1β) and tumor necrosis factor- (TNF-), while no such effect has been reported with respect to the VEGF gene, we investigated the effects of IL-1β and TNF- on the activation of the HIF-1 DNA-binding complex and the amount of HIF-1 protein in human hepatoma cells in culture. Under normoxic conditions, both cytokines caused a moderate activation of HIF-1 DNA binding. In hypoxia, cytokines strongly increased HIF-1 activity compared with the effect of hypoxia alone. Only IL-1β increased HIF-1 protein levels. In transient transfection experiments, HIF-1–driven reporter gene expression was augmented by cytokines only under hypoxic conditions. In contrast to their effect on EPO synthesis, neither IL-1β nor TNF- decreased VEGF production. The mRNA levels of HIF-1 and VEGF were unaffected. Thus, cytokine-induced inhibition of EPO production is not mediated by impairment of HIF-1 function. We propose that HIF-1 may be involved in modulating gene expression during inflammation.

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