Transcriptional regulation of the Xenopus laevis B.beta. fibrinogen subunit gene by glucocorticoids and hepatocyte nuclear factor 1: Analysis by transfection into primary liver cells

Roberts, Lewis R.; Nichols, LaNita A.; Holland, Lené J.

doi:10.1021/bi00094a020

Cited by 12 publications

(20 citation statements)

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“…A second consequence of HNF4 expression in H5 cells is the reexpression of a subset of genes whose expression is characteristic of hepatocytes and of well-differentiated cells of the H4II lineage. The genes coding for ␣1-AT, ␤Fib, and TTR all possess well-characterized HNF4 or HNF1 binding sites in their regulatory regions (18,33,50). Since both the ␣1-AT and ␤Fib genes are weakly expressed in H5 cells subjected to longterm DEX treatment, both genes posses the potential to be expressed: the presence of HNF4tag and HNF1 permits real-ization of this potential.…”

Section: Phenotypic Consequences Of Expression Of Hnf4mentioning

confidence: 99%

Hepatocyte Nuclear Factor 4 Expression Overcomes Repression of the Hepatic Phenotype in Dedifferentiated Hepatoma Cells

Späth

Weiss

1997

Molecular and Cellular Biology

109

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The capacity of the liver-enriched transcription factor hepatocyte nuclear factor 4 (HNF4) to direct redifferentiation of dedifferentiated rat hepatoma cells was investigated by stable transfection of epitope-tagged HNF4 cDNA into H5 variant cells. HNF4-producing cells expressed the previously silent HNF1 gene and showed activation of some hepatic functions, including alpha1-antitrypsin, beta-fibrinogen, and transthyretin, but not of the endogenous HNF4 gene. Expression of the other hepatocyte-enriched transcription factors was not modified. Treatment of the HNF4tag-expressing cells with dexamethasone induced expression of the transgene by 10-fold, resulting in enhanced expression of target genes of both glucocorticoid hormones and HNF4. The set of activated hepatic genes was extended by treatment of cells with the demethylating agent 5-azacytidine followed by selection in dexamethasone-containing glucose-free medium. Some of the colonies that developed reexpressed the entire set of hepatic functions tested. Fusion of HNF4tag-producing H5 cells with well-differentiated Fao cells showed that only those hybrids which maintained expression of HNF4tag were protected from complete extinction, including that of the Fao HNF4 gene. Thus, H5 cells must produce an extinguisher of the HNF4 gene. In addition, this result implies that HNF4 itself, or its target HNF1, is a positive regulator of HNF4. In conclusion, HNF4tag expression overcomes repression of the hepatic phenotype of the H5 cell without abolishing its potential to extinguish an active genome. Taken together, these results predict that expression of HNF4 should be sufficient to establish heritable expression of many parameters of the hepatic differentiated state.

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Section: Phenotypic Consequences Of Expression Of Hnf4mentioning

confidence: 99%

Hepatocyte Nuclear Factor 4 Expression Overcomes Repression of the Hepatic Phenotype in Dedifferentiated Hepatoma Cells

Späth

Weiss

1997

Molecular and Cellular Biology

109

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“…This holds true also for fibrinogen genes. In Xenopus laevis, a single GRE at bases -148 to -162 of the ␤ fibrinogen gene is present (28), while glucocorticoid regulation of the ␥ fibrinogen gene requires at least three nearby GREs and an accessory factor binding site, all within the first 187 bp of the promoter (34). In the human ␥ fibrinogen gene, we have shown that a GRE at position -1116 to -1102 confers dexamethasone inducibility.…”

Section: Discussionmentioning

confidence: 82%

Identification of a Glucocorticoid Response Element in the Human γ Chain Fibrinogen Promoter

Asselta¹,

Duga²,

Modugno³

et al. 1998

Thromb Haemost

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SummaryThe effect of the synthetic glucocorticoid hormone dexamethasone on human γ chain fibrinogen gene expression was examined. The whole promoter region of 3.8 kb of this gene and progressive 5’-deletions were inserted into a promoterless expression vector, upstream of the luciferase gene and transiently transfected into the human hepatoma HepG2 cells, in the presence or in the absence of dexamethasone stimulation. Deletion analysis allowed to identify a region located between –1359 and –954 bp upstream from the transcription start site, involved in hormone inducibility. On the basis of a computer-assisted analysis, a putative GRE was found in this region at bases –1116 to –1102. Specific point mutations eliminating this putative GRE led to complete loss of glucocorticoid inducibility, thus indicating its functional role. Binding of the rat glucocorticoid receptor to this site was demonstrated by mobility-shift assays.

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“…The value for Bβ-136 represents a non-induced baseline from a construct previously shown to be unresponsive to glucocorticoids (19). The percentages from six to ten experiments were averaged and the standard error of the mean (SEM) was calculated.…”

Section: Luciferase and β-Gal Reporter Assaysmentioning

confidence: 99%

Flanking Sequence Composition Differentially Affects the Binding and Functional Characteristics of Glucocorticoid Receptor Homo- and Heterodimers

2006

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The core binding sites for a multitude of transcription factors have been identified and characterized, but these sequences cannot fully account for the nuances of cell-specific and gene-specific control of gene transcription. Many factors may contribute to the precise responsiveness of a gene to a particular transcriptional regulatory protein, including the nucleotides in close proximity to the core binding site for that protein. Here, we examine two flanking sequences bordering a site in the γ-fibrinogen gene regulatory region that binds a heterodimer of the Xenopus glucocorticoid receptor accessory factor (XGRAF) and the glucocorticoid receptor (GR). Mutation of the upstream flank results in a decrease in XGRAF binding, but little change in hormone induction. However, alteration of the downstream flank adjacent to the GR binding site causes a decrease in both GR monomer binding and hormone induction. Conversion of the XGRAF:GR binding site to a full glucocorticoid response element (GRE) alters the role of the flanking sequences. A full GRE in this position requires the wild type upstream flank to bind GR homodimer and induce transcription to maximal levels. In contrast, mutation of the downstream flank is not detrimental to either binding or function of the GR dimer. Thus, flanking sequence composition and dimer partner combine to influence GR function, underscoring the complexities involved in the identification of authentic transcription factor response elements.The dynamic interaction of transcription factors with DNA regulates gene expression, conferring temporal and cell-specific control in response to a wide range of intracellular and extracellular cues. However, these protein:DNA interactions are not always easily predicted, as the exact sequence of a binding site can alter its function in unexpected ways. For instance, particular sequences may influence the recruitment of co-factors by altering the conformation of the bound transcription factor via specific interactions with individual nucleotides (1-3). If a site binds transcription factors as a dimer, both spacing and orientation of the two half sites affect how the site behaves (4,5). Also, not all functional binding sites are a good match to the consensus sequence compiled from known binding sites for a particular factor (6).In addition, the interactions between transcription factors and DNA do not necessarily rely solely on the sequence of the actual binding site, but may be influenced by other DNA elements. Sequences flanking the binding site can affect response element utilization by altering the protein conformation of a factor bound to the DNA (7). Nearby sequences may bind

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Transcriptional regulation of the Xenopus laevis B.beta. fibrinogen subunit gene by glucocorticoids and hepatocyte nuclear factor 1: Analysis by transfection into primary liver cells

Cited by 12 publications

References 50 publications

Hepatocyte Nuclear Factor 4 Expression Overcomes Repression of the Hepatic Phenotype in Dedifferentiated Hepatoma Cells

Hepatocyte Nuclear Factor 4 Expression Overcomes Repression of the Hepatic Phenotype in Dedifferentiated Hepatoma Cells

Identification of a Glucocorticoid Response Element in the Human γ Chain Fibrinogen Promoter

Flanking Sequence Composition Differentially Affects the Binding and Functional Characteristics of Glucocorticoid Receptor Homo- and Heterodimers

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