scholarly journals Characterization of a novel Foxa (hepatocyte nuclear factor-3) site in the glucagon promoter that is conserved between rodents and humans

2005 ◽  
Vol 389 (3) ◽  
pp. 831-841 ◽  
Author(s):  
Sanjeev K. Sharma ◽  
Ulrike Leinemann ◽  
Regine Ratke ◽  
Elke Oetjen ◽  
Roland Blume ◽  
...  
Keyword(s):  
Pancreatic Islet ◽  
Transcriptional Activation ◽  
Hepatic Glucose Production ◽  
Glucose Production ◽  
Comparative Sequence Analysis ◽  
Nuclear Factor ◽  
Hepatocyte Nuclear Factor ◽  
Enhancer Activity ◽  
Enhancer Element ◽  
Nuclear Extracts

The pancreatic islet hormone glucagon stimulates hepatic glucose production and thus maintains blood glucose levels in the fasting state. Transcription factors of the Foxa [Fox (forkhead box) subclass A; also known as HNF-3 (hepatocyte nuclear factor-3)] family are required for cell-specific activation of the glucagon gene in pancreatic islet α-cells. However, their action on the glucagon gene is poorly understood. In the present study, comparative sequence analysis and molecular characterization using protein–DNA binding and transient transfection assays revealed that the well-characterized Foxa-binding site in the G2 enhancer element of the rat glucagon gene is not conserved in humans and that the human G2 sequence lacks basal enhancer activity. A novel Foxa site was identified that is conserved in rats, mice and humans. It mediates activation of the glucagon gene by Foxa proteins and confers cell-specific promoter activity in glucagon-producing pancreatic islet α-cell lines. In contrast with previously identified Foxa-binding sites in the glucagon promoter, which bind nuclear Foxa2, the novel Foxa site was found to bind preferentially Foxa1 in nuclear extracts of a glucagon-producing pancreatic islet α-cell line, offering a mechanism that explains the decrease in glucagon gene expression in Foxa1-deficient mice. This site is located just upstream of the TATA box (between −30 and −50), suggesting a role for Foxa proteins in addition to direct transcriptional activation, such as a role in opening the chromatin at the start site of transcription of the glucagon gene.

scholarly journals Modulation of liver-specific transcription by interactions between hepatocyte nuclear factor 3 and nuclear factor 1 binding DNA in close apposition.

1993 ◽  
Vol 13 (4) ◽  
pp. 2401-2410 ◽  
Author(s):  
D A Jackson ◽  
K E Rowader ◽  
K Stevens ◽  
C Jiang ◽  
P Milos ◽  
...  
Keyword(s):  
Transcriptional Activation ◽  
Mutational Analysis ◽  
Regulatory Element ◽  
Nuclear Factor ◽  
Hepatocyte Nuclear Factor ◽  
In Vitro Differentiation ◽  
Enhancer Activity ◽  
Activation Of Transcription ◽  
Nuclear Factor 1

The liver-specific enhancer of the serum albumin gene contains an essential segment, designated eH, which binds the hepatocyte nuclear factor 3 alpha (HNF3 alpha) and ubiquitous nuclear factor 1/CCAAT transcription factor (NF1/CTF) proteins in tight apposition. We previously showed that activation of transcription by the eH site was correlated with an increase in intracellular HNF3 alpha levels during the in vitro differentiation of the hepatic cell line H2.35. We now show that transfection of an HNF3 alpha cDNA expression vector into dedifferentiated H2.35 cells is sufficient to induce transcription from the eH site. Mutational analysis of the enhancer demonstrates that NF1/CTF cooperates with HNF3 alpha to induce enhancer activity. However, when the eH site is removed from the context of the enhancer, NF1/CTF can inhibit transcriptional activation by HNF3 alpha. We conclude that the ternary complex of HNF3 alpha, NF1/CTF, and the eH site forms a novel, composite regulatory element that is sensitive to the local DNA sequence environment and suggest that the transcriptional stimulatory activity of NF1/CTF depends on its higher-order interactions with other proteins during hepatocyte differentiation.

Modulation of liver-specific transcription by interactions between hepatocyte nuclear factor 3 and nuclear factor 1 binding DNA in close apposition

1993 ◽  
Vol 13 (4) ◽  
pp. 2401-2410
Author(s):  
D A Jackson ◽  
K E Rowader ◽  
K Stevens ◽  
C Jiang ◽  
P Milos ◽  
...  
Keyword(s):  
Transcriptional Activation ◽  
Mutational Analysis ◽  
Regulatory Element ◽  
Nuclear Factor ◽  
Hepatocyte Nuclear Factor ◽  
In Vitro Differentiation ◽  
Enhancer Activity ◽  
Activation Of Transcription ◽  
Nuclear Factor 1

The liver-specific enhancer of the serum albumin gene contains an essential segment, designated eH, which binds the hepatocyte nuclear factor 3 alpha (HNF3 alpha) and ubiquitous nuclear factor 1/CCAAT transcription factor (NF1/CTF) proteins in tight apposition. We previously showed that activation of transcription by the eH site was correlated with an increase in intracellular HNF3 alpha levels during the in vitro differentiation of the hepatic cell line H2.35. We now show that transfection of an HNF3 alpha cDNA expression vector into dedifferentiated H2.35 cells is sufficient to induce transcription from the eH site. Mutational analysis of the enhancer demonstrates that NF1/CTF cooperates with HNF3 alpha to induce enhancer activity. However, when the eH site is removed from the context of the enhancer, NF1/CTF can inhibit transcriptional activation by HNF3 alpha. We conclude that the ternary complex of HNF3 alpha, NF1/CTF, and the eH site forms a novel, composite regulatory element that is sensitive to the local DNA sequence environment and suggest that the transcriptional stimulatory activity of NF1/CTF depends on its higher-order interactions with other proteins during hepatocyte differentiation.

scholarly journals The mechanism by which the human apolipoprotein B gene reducer operates involves blocking of transcriptional activation by hepatocyte nuclear factor 3.

1993 ◽  
Vol 13 (3) ◽  
pp. 1534-1546 ◽  
Author(s):  
B Paulweber ◽  
F Sandhofer ◽  
B Levy-Wilson
Keyword(s):  
Binding Site ◽  
Transcriptional Activation ◽  
Apolipoprotein B ◽  
Regulatory Protein ◽  
Nuclear Factor ◽  
Hepatocyte Nuclear Factor ◽  
Apo B ◽  
Human Apolipoprotein ◽  
Colon Carcinoma Cells ◽  
Mediator Protein

Previously, we showed that when a DNA fragment extending from -3067 to -2734 of the human apolipoprotein B (apo-B) gene is inserted immediately upstream of an apo-B promoter segment (-139 to +121), transcription from this promoter is reduced by about 10-fold in cultured colon carcinoma cells (CaCo-2) but not in cultured hepatoma cells (HepG2). We postulated that this reducer operates by a mechanism involving active repression of a transcriptional activator that binds to the segment from -111 to -88 of the apo-B promoter (B. Paulweber and B. Levy-Wilson, J. Biol. Chem. 266:24161-24168 1991). In the current study, the reducer element has been localized to a 24-bp sequence from -2801 to -2778 of the apo-B gene that contains a binding site for the negative regulatory protein ARP-1. Furthermore, we have demonstrated that the transcription factor hepatocyte nuclear factor 3 alpha (HNF-3 alpha) binds to the sequence 5'-TGTTTGCTTTTC-3' from -95 to -106 of the apo-B promoter, to stimulate transcription. Transcriptional activation by HNF-3 is repressed when the reducer sequence is inserted immediately upstream of the HNF-3 binding site, suggesting a mechanism by which the reducer-bound protein blocks the activation promoted by HNF-3. Data from cotransfection experiments in which ARP-1 is overexpressed in the absence of its binding site suggest that ARP-1 interacts either directly or via a mediator protein with proteins recognizing the HNF-3 site and that this interaction is sufficient to repress transcriptional activation by HNF-3. Because transcriptional activation by Sp1 is not affected by the reducer, it is unlikely that the reducer interacts directly with basic components of the transcriptional machinery.

scholarly journals Regulation of the T-cell receptor delta enhancer by functional cooperation between c-Myb and core-binding factors.

1994 ◽  
Vol 14 (1) ◽  
pp. 473-483 ◽  
Author(s):  
C Hernandez-Munain ◽  
M S Krangel
Keyword(s):  
T Cell ◽  
T Cell Receptor ◽  
Transcriptional Activation ◽  
Cell Receptor ◽  
Enhancer Activity ◽  
Control Of Gene Expression ◽  
Enhancer Element ◽  
The Core ◽  
Core Site ◽  
Factor C

A T-cell-specific transcriptional enhancer lies within the J delta 3-C delta intron of the human T-cell receptor (TCR) delta gene. The 30-bp minimal enhancer element denoted delta E3 carries a core sequence (TGTGGTTT) that binds a T-cell-specific factor, and that is necessary but not sufficient for transcriptional activation. Here we demonstrate that the transcription factor c-Myb regulates TCR delta enhancer activity through a binding site in delta E3 that is adjacent to the core site. Both v-Myb and c-Myb bind specifically to delta E3. The Myb site is necessary for enhancer activity, because a mutation that eliminates Myb binding abolishes transcriptional activation by the delta E3 element and by the 370-bp TCR delta enhancer. Transfection of cells with a c-Myb expression construct upregulates delta E3 enhancer activity, whereas treatment of cells with an antisense c-myb oligonucleotide inhibits delta E3 enhancer activity. Since intact Myb and core sites are both required for delta E3 function, our data argue that c-Myb and core binding factors must cooperate to mediate transcriptional activation through delta E3. Efficient cooperation depends on the relative positioning of the Myb and core sites, since only one of two overlapping Myb sites within delta E3 is functional and alterations of the distance between this site and the core site disrupt enhancer activity. Cooperative regulation by c-Myb and core-binding factors is likely to play an important role in the control of gene expression during T-cell development.

scholarly journals Negative autoregulation of the neu gene is mediated by a novel enhancer.

1992 ◽  
Vol 12 (6) ◽  
pp. 2739-2748 ◽  
Author(s):  
X Y Zhao ◽  
M C Hung
Keyword(s):  
Protein Complexes ◽  
Feedback Regulation ◽  
Specific Protein ◽  
Initiation Codon ◽  
Enhancer Activity ◽  
Enhancer Element ◽  
Nuclear Extracts ◽  
Gel Shift ◽  
Neu Oncogene ◽  
Transformed Cell Lines

In an attempt to study potential feedback regulation of the neu oncogene, we have found that the neu oncogene product specifically represses its own promoter activity. Deletion analysis indicated a 140-bp region (nucleotides -312 to -173 relative to the ATG initiation codon) in the rat neu promoter responsible for neu autorepression. Gel shift assays and methylation interference analysis further demonstrated that a GGTGGGGGGG sequence (nucleotides -243 to -234 relative to the ATG initiation codon) in this 140-bp region interacts with specific protein complexes. The GGTGGGGGGG sequence (GTG element), which functions as an enhancer, is sufficient to cause neu-mediated repression in a heterologous promoter. Furthermore, it produces different gel shift patterns with nuclear extracts from neu-transformed cell lines and their parental lines, suggesting that a transcriptional factor(s) interacting with this enhancer element has been perturbed by the introduction of neu. Taken together, the data presented in this report show that (i) the neu oncogene product autorepresses its own promoter, (ii) the neu promoter contains a novel enhancer, and (iii) neu autorepression is mediated through this enhancer, likely by inhibition of the enhancer activity.

Co-operative regulation of the transcription of human dihydrodiol dehydrogenase (DD)4/aldo–keto reductase (AKR)1C4 gene by hepatocyte nuclear factor (HNF)-4α/γ and HNF-1α

2001 ◽  
Vol 355 (2) ◽  
pp. 537-544 ◽  
Author(s):  
Takeshi OZEKI ◽  
Yoshiki TAKAHASHI ◽  
Toshiyuki KUME ◽  
Kazuo NAKAYAMA ◽  
Tsuyoshi YOKOI ◽  
...  
Keyword(s):  
Hepg2 Cells ◽  
Binding Motif ◽  
Nuclear Factor ◽  
Hepatocyte Nuclear Factor ◽  
Nuclear Extracts ◽  
Redox Active ◽  
Supershift Assay ◽  
Polycyclic Aromatic ◽  
Gel Shift Assay ◽  
Dihydrodiol Dehydrogenase

Human dihydrodiol dehydrogenase (DD) 4/aldo-keto reductase (AKR) 1C4 is a major isoform of hepatic DD that oxidizes trans-dihydrodiols of polycyclic aromatic hydrocarbons to reactive and redox-active o-quinones and that reduces several ketone-containing drugs. To investigate the mechanism of transcriptional regulation of the human DD4 gene, the 5′-flanking region of the gene was fused to the luciferase gene. The results of luciferase assays using HepG2 cells and of 1,10-phenanthroline-copper footprinting indicated that two positive regulatory regions were located in regions from -701 to -684 and from -682 to -666. The former region contained a putative hepatocyte nuclear factor (HNF)-4 binding motif, and the latter region contained an HNF-1 consensus binding sequence. DNA fragments of the HNF-4 or HNF-1 motif gave a shifted band in a gel-shift assay with nuclear extracts from HepG2 cells. The formation of the DNA-protein complex was inhibited by the HNF-4 or HNF-1 motif of the α1-antitrypsin gene. A supershift assay using antibodies to human HNF-4α, HNF-4γ and HNF-1α showed that HNF-4α and HNF-4γ bound to the HNF-4 motif, and that HNF-1α interacted with the HNF-1 motif. Introduction of mutations into the HNF-4 or HNF-1 motif lowered the luciferase activity to 10 or 8% respectively of that seen with the intact human DD4 gene. These results indicate that HNF-4α, HNF-4γ and HNF-1α regulate co-operatively the transcription of the human DD4 gene in HepG2 cells.

Sign in / Sign up

Export Citation Format

Share Document