scholarly journals Chromatin-Mediated Restriction of Nuclear Factor 1/CTF Binding in a Repressed and Hormone-Activated Promoter In Vivo

2004 ◽  
Vol 24 (7) ◽  
pp. 3036-3047 ◽  
Author(s):  
Sergey Belikov ◽  
Carolina Åstrand ◽  
Per-Henrik Holmqvist ◽  
Örjan Wrange
Keyword(s):  
Dna Binding ◽  
Chromatin Remodeling ◽  
Gene Networks ◽  
Fold Increase ◽  
Micrococcal Nuclease ◽  
Specific Gene ◽  
Nuclear Factor ◽  
Mmtv Promoter ◽  
Nuclear Factor 1

ABSTRACT Mouse mammary tumor virus (MMTV) promoter-driven transcription is induced by glucocorticoid hormone via binding of the glucocorticoid receptor (GR). The MMTV promoter also harbors a binding site for nuclear factor 1 (NF1). NF1 and GR were expressed in Xenopus oocytes; this revealed GR-NF1 cooperativity both in terms of DNA binding and chromatin remodeling but not transcription. A fraction of NF1 sites were occupied in a hormone-dependent fashion, but a significant and NF1 concentration-dependent fraction were constitutively bound. Activation of the MMTV promoter resulted in an ∼50-fold increase in the NF1 accessibility for its DNA site. The hormone-dependent component of NF1 binding was dissociated by addition of a GR antagonist; however, the antagonist RU486, which supports partial GR-DNA binding, also maintained partial NF1 binding. Hence GR-NF1 cooperativity is independent of agonist-driven chromatin remodeling. NF1 induced the formation of a micrococcal-nuclease-resistant protein-DNA complex containing the DNA segment from −185 to −55, the MMTV enhanceosome. Coexpression of NF1 and Oct1 resulted in a significant stimulation of hormone-induced MMTV transcription and also in increased basal transcription. We propose that hormone-independent NF1 binding may be involved in maintaining transcriptional competence and establishment of tissue-specific gene networks.

Overexpression of Myc suppresses CCAAT transcription factor/nuclear factor 1-dependent promoters in vivo

1993 ◽  
Vol 13 (5) ◽  
pp. 3093-3102
Author(s):  
B S Yang ◽  
J D Gilbert ◽  
S O Freytag
Keyword(s):  
Transcription Factor ◽  
Dna Binding ◽  
Regulatory Elements ◽  
Specific Gene ◽  
Quiescent State ◽  
Nuclear Factor ◽  
Nuclear Factor 1 ◽  
Transcription Factor Nuclear Factor ◽  
In Cells

Overexpression of Myc in cells can suppress the transcription of specific genes. Because several of these genes have common transcriptional regulatory elements, we investigated the possibility that this effect of Myc is mediated through a specific transcription factor. In vitro DNA-binding assays detect only one form of CCAAT transcription factor/nuclear factor 1 (CTF/NF-1) in quiescent 3T3-L1 cells. By contrast, quiescent 3T3-L1 cells that stably overexpress either c-Myc or N-Myc contain at least three forms of CTF/NF-1. Biochemical characterization of the various CTF/NF-1 forms showed that they have the same native molecular weight but differ in charge density. The more negatively charged CTF/NF-1 forms present in Myc-overexpressing cells are converted into that found in normal cells by treatment with acid phosphatase, suggesting that they represent a more phosphorylated form of the CTF/NF-1 protein. The various CTF/NF-1 forms have a similar DNA-binding affinity. Transfection experiments demonstrated that transcription from CTF/NF-1-dependent promoters is specifically suppressed in cells that stably overexpress c-Myc. This effect requires CTF/NF-1 binding. CTF/NF-1-dependent promoter activity is also suppressed in 3T3-L1 cells during active growth (relative to the quiescent state). Interestingly, actively growing 3T3-L1 cells contain forms of CTF/NF-1 similar to those in quiescent cells that stably overexpress c-Myc. Thus, the CTF/NF-1 forms present in cells that express high amounts of c-Myc correlate with a lower transcription rate of CTF/NF-1-dependent promoters in vivo. Our results provide a basis for the suppression of specific gene transcription by c-Myc.

scholarly journals Overexpression of Myc suppresses CCAAT transcription factor/nuclear factor 1-dependent promoters in vivo.

1993 ◽  
Vol 13 (5) ◽  
pp. 3093-3102 ◽  
Author(s):  
B S Yang ◽  
J D Gilbert ◽  
S O Freytag
Keyword(s):  
Transcription Factor ◽  
Dna Binding ◽  
Regulatory Elements ◽  
Specific Gene ◽  
Quiescent State ◽  
Nuclear Factor ◽  
Nuclear Factor 1 ◽  
Transcription Factor Nuclear Factor ◽  
In Cells

Overexpression of Myc in cells can suppress the transcription of specific genes. Because several of these genes have common transcriptional regulatory elements, we investigated the possibility that this effect of Myc is mediated through a specific transcription factor. In vitro DNA-binding assays detect only one form of CCAAT transcription factor/nuclear factor 1 (CTF/NF-1) in quiescent 3T3-L1 cells. By contrast, quiescent 3T3-L1 cells that stably overexpress either c-Myc or N-Myc contain at least three forms of CTF/NF-1. Biochemical characterization of the various CTF/NF-1 forms showed that they have the same native molecular weight but differ in charge density. The more negatively charged CTF/NF-1 forms present in Myc-overexpressing cells are converted into that found in normal cells by treatment with acid phosphatase, suggesting that they represent a more phosphorylated form of the CTF/NF-1 protein. The various CTF/NF-1 forms have a similar DNA-binding affinity. Transfection experiments demonstrated that transcription from CTF/NF-1-dependent promoters is specifically suppressed in cells that stably overexpress c-Myc. This effect requires CTF/NF-1 binding. CTF/NF-1-dependent promoter activity is also suppressed in 3T3-L1 cells during active growth (relative to the quiescent state). Interestingly, actively growing 3T3-L1 cells contain forms of CTF/NF-1 similar to those in quiescent cells that stably overexpress c-Myc. Thus, the CTF/NF-1 forms present in cells that express high amounts of c-Myc correlate with a lower transcription rate of CTF/NF-1-dependent promoters in vivo. Our results provide a basis for the suppression of specific gene transcription by c-Myc.

scholarly journals Nuclear Factor 1 Is Required for Both Hormone-Dependent Chromatin Remodeling and Transcriptional Activation of the Mouse Mammary Tumor Virus Promoter

2003 ◽  
Vol 23 (3) ◽  
pp. 887-898 ◽  
Author(s):  
Pratibha B. Hebbar ◽  
Trevor K. Archer
Keyword(s):  
Binding Site ◽  
Chromatin Remodeling ◽  
Transcriptional Activation ◽  
Mouse Mammary Tumor Virus ◽  
Nuclear Factor ◽  
Mammary Tumor Virus ◽  
Mouse Mammary Tumor ◽  
Tumor Virus ◽  
Mmtv Promoter ◽  
Nuclear Factor 1

ABSTRACT The mouse mammary tumor virus (MMTV) promoter has been used as a model to study how the glucocorticoid receptor (GR) remodels chromatin to allow other transcription factors to bind and activate transcription. To dissect the precise role of nuclear factor 1 (NF1) in chromatin remodeling and transcriptional activation, we used linker-scanning mutants of transcription factor binding sites on the MMTV promoter. We compared the NF1 mutant MMTV promoter in the context of transiently transfected templates (transient transfection) and templates organized as chromatin (stable transfection) to understand the effect of chromatin on factor binding and transcription. We show that on a transiently transfected template, mutation in the NF1 binding site reduces both basal and hormone-dependent transcription. This suggests that NF1 is required for transcription in the absence of organized chromatin. We also found that binding of NF1 on a transiently transfected template is independent of mutation in hormone response elements or the octamer transcription factor (OTF) binding site. In contrast, the binding of OTF proteins to a transiently transfected template was found to be dependent on the binding of NF1, which may imply that NF1 has a stabilizing effect on OTF binding. On a chromatin template, mutation in the NF1 binding site does not affect the positioning of nucleosomes on the promoter. We also show that in the absence of NF1 binding, GR-mediated chromatin remodeling of nucleosome B is reduced and hormone-dependent activation of transcription is abolished. Further, we demonstrate that NF1 is required for both the association of BRG1 chromatin remodeling complex and the GR on the promoter in vivo. These results suggest the novel possibility that NF1 may participate in chromatin remodeling activities in addition to directly enhancing transcription and that in the absence of its binding site the GR is unable to effectively bind the promoter and recruit the remodeling complex.

A direct requirement of nuclear factor-κB for suppression of apoptosis in ventricular myocytes

2000 ◽  
Vol 279 (3) ◽  
pp. H939-H945 ◽  
Author(s):  
Shareef Mustapha ◽  
Alla Kirshner ◽  
Danielle De Moissac ◽  
Lorrie A. Kirshenbaum
Keyword(s):  
Dna Binding ◽  
Gene Transcription ◽  
Ventricular Myocytes ◽  
Vital Staining ◽  
Fold Increase ◽  
Nuclear Factor Κb ◽  
Nuclear Factor ◽  
Cytoplasmic Factor ◽  
Tnf Α ◽  
Factor Α

Nuclear factor-κB (NF-κB) is a ubiquitously expressed cellular factor regulated by the cytoplasmic factor inhibitor protein κBα (IκBα). Activation of NF-κB by cytokines, including tumor necrosis factor-α (TNF-α), requires the phosphorylation and degradation of IκBα. An anti-apoptotic role for NF-κB has recently been suggested. In the present study, we ascertained whether death-promoting signals and apoptosis mediated by TNF-α are suppressed by NF-κB in postnatal ventricular myocytes. Stimulation of myocytes with TNF-α resulted in a 12.1-fold increase ( P < 0.01) in NF-κB-dependent gene transcription and DNA binding compared with controls. This was accompanied by a corresponding increase in the NF-κB target protein A20 as determined by Western blot analysis. Vital staining revealed that TNF-α was not cytotoxic to myocytes and did not provoke apoptosis. Adenovirus-mediated delivery of a nonphosphorylatable form of IκBα to inactivate NF-κB prevented TNF-α-stimulated NF-κB-dependent gene transcription and nuclear NF-κB DNA binding. Importantly, myocytes stimulated with TNF-α and defective for NF-κB activation resulted in a 2.2-fold increase ( P < 0.001) in apoptosis. To our knowledge, the data provide the first indication that a functional NF-κB signaling pathway is crucial for suppressing death-promoting signals mediated by TNF-α in ventricular myocytes.

scholarly journals Androgen and glucocorticoid receptor direct distinct transcriptional programs by receptor-specific and shared DNA binding sites

2021 ◽  
Vol 49 (7) ◽  
pp. 3856-3875
Author(s):  
Marina Kulik ◽  
Melissa Bothe ◽  
Gözde Kibar ◽  
Alisa Fuchs ◽  
Stefanie Schöne ◽  
...  
Keyword(s):  
Gene Regulation ◽  
Transcription Factor ◽  
Dna Binding ◽  
Receptor Binding ◽  
Binding Sites ◽  
Specific Gene ◽  
Functional Diversification ◽  
Enhancer Activity ◽  
Sequence Composition

Abstract The glucocorticoid (GR) and androgen (AR) receptors execute unique functions in vivo, yet have nearly identical DNA binding specificities. To identify mechanisms that facilitate functional diversification among these transcription factor paralogs, we studied them in an equivalent cellular context. Analysis of chromatin and sequence suggest that divergent binding, and corresponding gene regulation, are driven by different abilities of AR and GR to interact with relatively inaccessible chromatin. Divergent genomic binding patterns can also be the result of subtle differences in DNA binding preference between AR and GR. Furthermore, the sequence composition of large regions (&gt;10 kb) surrounding selectively occupied binding sites differs significantly, indicating a role for the sequence environment in guiding AR and GR to distinct binding sites. The comparison of binding sites that are shared shows that the specificity paradox can also be resolved by differences in the events that occur downstream of receptor binding. Specifically, shared binding sites display receptor-specific enhancer activity, cofactor recruitment and changes in histone modifications. Genomic deletion of shared binding sites demonstrates their contribution to directing receptor-specific gene regulation. Together, these data suggest that differences in genomic occupancy as well as divergence in the events that occur downstream of receptor binding direct functional diversification among transcription factor paralogs.

scholarly journals The DNA-binding protein HTa from Thermoplasma acidophilum is an archaeal histone analog

eLife ◽  
2019 ◽  
Vol 8 ◽  
Author(s):  
Antoine Hocher ◽  
Maria Rojec ◽  
Jacob B Swadling ◽  
Alexander Esin ◽  
Tobias Warnecke
Keyword(s):  
Dna Binding ◽  
Binding Protein ◽  
Dna Binding Protein ◽  
Coli Strain ◽  
Micrococcal Nuclease ◽  
Thermoplasma Acidophilum ◽  
Chromatin Architecture ◽  
Archaeal Histone

Histones are a principal constituent of chromatin in eukaryotes and fundamental to our understanding of eukaryotic gene regulation. In archaea, histones are widespread but not universal: several lineages have lost histone genes. What prompted or facilitated these losses and how archaea without histones organize their chromatin remains largely unknown. Here, we elucidate primary chromatin architecture in an archaeon without histones, Thermoplasma acidophilum, which harbors a HU family protein (HTa) that protects part of the genome from micrococcal nuclease digestion. Charting HTa-based chromatin architecture in vitro, in vivo and in an HTa-expressing E. coli strain, we present evidence that HTa is an archaeal histone analog. HTa preferentially binds to GC-rich sequences, exhibits invariant positioning throughout the growth cycle, and shows archaeal histone-like oligomerization behavior. Our results suggest that HTa, a DNA-binding protein of bacterial origin, has converged onto an architectural role filled by histones in other archaea.

scholarly journals Two-Step Synergism between the Progesterone Receptor and the DNA-Binding Domain of Nuclear Factor 1 on MMTV Minichromosomes

1999 ◽  
Vol 4 (1) ◽  
pp. 45-54 ◽  
Author(s):  
Luciano Di Croce ◽  
Ronald Koop ◽  
Patrizia Venditti ◽  
Hannes M Westphal ◽  
Karl P Nightingale ◽  
...  
Keyword(s):  
Dna Binding ◽  
Progesterone Receptor ◽  
Binding Domain ◽  
Nuclear Factor ◽  
Dna Binding Domain ◽  
Nuclear Factor 1

scholarly journals Effect of mechanical stretch on HIF-1α and MMP-2 expression in capillaries isolated from overloaded skeletal muscles: laser capture microdissection study

2005 ◽  
Vol 289 (3) ◽  
pp. H1315-H1320 ◽  
Author(s):  
Malgorzata Milkiewicz ◽  
Tara L. Haas
Keyword(s):  
Endothelial Cells ◽  
Laser Capture Microdissection ◽  
Skeletal Muscles ◽  
Hypoxia Inducible Factor ◽  
Fold Increase ◽  
Mechanical Stretch ◽  
Specific Gene ◽  
Capillary Endothelial Cells ◽  
Laser Capture

Under physiological nonhypoxic conditions, angiogenesis can be driven by mechanical forces. However, because of the limitations of the specific gene expression analysis of microvessels from in vivo experiments, the mechanisms regulating the coordinated expression of angiogenic factors implicated in the process remain intangible. In this study, the technique of laser capture microdissection (LCM) was adapted for the study of angiogenesis in skeletal muscles. With a combination of LCM and real-time quantitative PCR it was demonstrated that capillary endothelial cells produce matrix metalloproteinase (MMP)-2 and that mechanical stretch of capillaries within muscle tissue markedly increases MMP-2 mRNA (2.5-fold increase vs. control; P < 0.05). In addition, we showed that transcription factor hypoxia-inducible factor (HIF)-1α expression was 13.5-fold higher in capillaries subjected to stretch compared with controls ( P < 0.05). These findings demonstrate the feasibility of this approach to study angiogenic gene regulation and provide novel evidence of HIF-1α induction in stretched capillary endothelial cells.

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