scholarly journals Upstream stimulatory factor (USF) and neurogenic differentiation/beta-cell E box transactivator 2 (NeuroD/BETA2) contribute to islet-specific glucose-6-phosphatase catalytic-subunit-related protein (IGRP) gene expression

2003 ◽  
Vol 371 (3) ◽  
pp. 675-686 ◽  
Author(s):  
Cyrus C. MARTIN ◽  
Christina A. SVITEK ◽  
James K. OESER ◽  
Eva HENDERSON ◽  
Roland STEIN ◽  
...  
Keyword(s):  
Gene Expression ◽  
Binding Sites ◽  
Fusion Gene ◽  
Catalytic Subunit ◽  
Related Protein ◽  
Upstream Stimulatory Factor ◽  
Factor Binding ◽  
E Box ◽  
Stimulatory Factor

Islet-specific glucose-6-phosphatase (G6Pase) catalytic-subunit-related protein (IGRP) is a homologue of the catalytic subunit of G6Pase, the enzyme that catalyses the final step of the gluconeogenic pathway. The analysis of IGRP-chloramphenicol acetyltransferase (CAT) fusion-gene expression through transient transfection of islet-derived βTC-3 cells revealed that multiple promoter regions, located between −306 and −97, are required for maximal IGRP-CAT fusion-gene expression. These regions correlated with trans-acting factor-binding sites in the IGRP promoter that were identified in βTC-3 cells in situ using the ligation-mediated PCR (LMPCR) footprinting technique. However, the LMPCR data also revealed additional trans-acting factor-binding sites located between −97 and +1 that overlap two E-box motifs, even though this region by itself conferred minimal fusion-gene expression. The data presented here show that these E-box motifs are important for IGRP promoter activity, but that their action is only manifest in the presence of distal promoter elements. Thus mutation of either E-box motif in the context of the −306 to +3 IGRP promoter region reduces fusion-gene expression. These two E-box motifs have distinct sequences and preferentially bind NeuroD/BETA2 neurogenic differentiation/β-cell E box transactivator 2 and upstream stimulatory factor (USF) in vitro, consistent with the binding of both factors to the IGRP promoter in situ, as determined using the chromatin-immunoprecipitation (ChIP) assay. Based on experiments using mutated IGRP promoter constructs, we propose a model to explain how the ubiquitously expressed USF could contribute to islet-specific IGRP gene expression.

scholarly journals Foxa2 and MafA regulate islet-specific glucose-6-phosphatase catalytic subunit-related protein gene expression

2008 ◽  
Vol 41 (5) ◽  
pp. 315-328 ◽  
Author(s):  
Cyrus C Martin ◽  
Brian P Flemming ◽  
Yingda Wang ◽  
James K Oeser ◽  
Richard M O'Brien
Keyword(s):  
Gene Expression ◽  
Type 1 Diabetes ◽  
Transcription Factors ◽  
Binding Sites ◽  
Fusion Gene ◽  
Fasting Blood Glucose ◽  
Catalytic Subunit ◽  
Related Protein ◽  
Min6 Cells

Islet-specific glucose-6-phosphatase catalytic subunit-related protein (IGRP/G6PC2) is a major autoantigen in both mouse and human type 1 diabetes. IGRP is selectively expressed in islet β cells and polymorphisms in the IGRP gene have recently been associated with variations in fasting blood glucose levels and cardiovascular-associated mortality in humans. Chromatin immunoprecipitation (ChIP) assays have shown that the IGRP promoter binds the islet-enriched transcription factors Pax-6 and BETA2. We show here, again using ChIP assays, that the IGRP promoter also binds the islet-enriched transcription factors MafA and Foxa2. Single binding sites for these factors were identified in the proximal IGRP promoter, mutation of which resulted in decreased IGRP fusion gene expression in βTC-3, Hamster insulinoma tumor (HIT), and Min6 cells. ChiP assays have shown that the islet-enriched transcription factor Pdx-1 also binds the IGRP promoter, but mutational analysis of four Pdx-1 binding sites in the proximal IGRP promoter revealed surprisingly little effect of Pdx-1 binding on IGRP fusion gene expression in βTC-3 cells. In contrast, in both HIT and Min6 cells mutation of these four Pdx-1 binding sites resulted in a ∼50% reduction in fusion gene expression. These data suggest that the same group of islet-enriched transcription factors, namely Pdx-1, Pax-6, MafA, BETA2, and Foxa2, directly or indirectly regulate expression of the two major autoantigens in type 1 diabetes.

scholarly journals Upstream stimulatory factor 1 activates GATA5 expression through an E-box motif

2012 ◽  
Vol 446 (1) ◽  
pp. 89-98 ◽  
Author(s):  
Bohao Chen ◽  
Rona Hsu ◽  
Zhenping Li ◽  
Paul C. Kogut ◽  
Qingxia Du ◽  
...  
Keyword(s):  
Gene Expression ◽  
Promoter Hypermethylation ◽  
Site Directed Mutagenesis ◽  
Luciferase Reporter ◽  
Upstream Stimulatory Factor ◽  
Mobility Shift ◽  
Reverse Transcription Pcr ◽  
E Box ◽  
Upstream Stimulatory Factor 1 ◽  
Stimulatory Factor

Silencing of GATA5 gene expression as a result of promoter hypermethylation has been observed in lung, gastrointestinal and ovarian cancers. However, the regulation of GATA5 gene expression has been poorly understood. In the present study, we have demonstrated that an E (enhancer)-box in the GATA5 promoter (bp −118 to −113 in mice; bp −164 to −159 in humans) positively regulates GATA5 transcription by binding USF1 (upstream stimulatory factor 1). Using site-directed mutagenesis, EMSA (electrophoretic mobility-shift analysis) and affinity chromatography, we found that USF1 specifically binds to the E-box sequence (5′-CACGTG-3′), but not to a mutated E-box. CpG methylation of this E-box significantly diminished its binding of transcription factors. Mutation of the E-box within a GATA5 promoter fragment significantly decreased promoter activity in a luciferase reporter assay. Chromatin immunoprecipitation identified that USF1 physiologically interacts with the GATA5 promoter E-box in mouse intestinal mucosa, which has the highest GATA5 gene expression in mouse. Co-transfection with a USF1 expression plasmid significantly increased GATA5 promoter-driven luciferase transcription. Furthermore, real-time and RT (reverse transcription)–PCR analyses confirmed that overexpression of USF1 activates endogenous GATA5 gene expression in human bronchial epithelial cells. The present study provides the first evidence that USF1 activates GATA5 gene expression through the E-box motif and suggests a potential mechanism (disruption of the E-box) by which GATA5 promoter methylation reduces GATA5 expression in cancer.

scholarly journals The Basic Helix-Loop-Helix, Leucine Zipper Transcription Factor, USF (Upstream Stimulatory Factor), Is a Key Regulator of SF-1 (Steroidogenic Factor-1) Gene Expression in Pituitary Gonadotrope and Steroidogenic Cells

1998 ◽  
Vol 12 (5) ◽  
pp. 714-726 ◽  
Author(s):  
Adrienne N. Harris ◽  
Pamela L. Mellon
Keyword(s):  
Gene Expression ◽  
Molecular Mechanisms ◽  
Transcriptional Regulator ◽  
Orphan Nuclear Receptor ◽  
Upstream Stimulatory Factor ◽  
Specific Expression ◽  
Steroidogenic Factor 1 ◽  
Helix Loop Helix ◽  
E Box ◽  
Stimulatory Factor

Abstract Tissue-specific expression of the mammalian FTZ-F1 gene is essential for adrenal and gonadal development and sexual differentiation. The FTZ-F1 gene encodes an orphan nuclear receptor, termed SF-1 (steroidogenic factor-1) or Ad4BP, which is a primary transcriptional regulator of several hormone and steroidogenic enzyme genes that are critical for normal physiological function of the hypothalamic-pituitary-gonadal axis in reproduction. The objective of the current study is to understand the molecular mechanisms underlying transcriptional regulation of SF-1 gene expression in the pituitary. We have studied a series of deletion and point mutations in the SF-1 promoter region for transcriptional activity in αT3–1 and LβT2 (pituitary gonadotrope), CV-1, JEG-3, and Y1 (adrenocortical) cell lines. Our results indicate that maximal expression of the SF-1 promoter in all cell types requires an E box element at −82/−77. This E box sequence (CACGTG) is identical to the binding element for USF (upstream stimulatory factor), a member of the helix-loop-helix family of transcription factors. Studies of the SF-1 gene E box element using gel mobility shift and antibody supershift assays indicate that USF may be a key transcriptional regulator of SF-1 gene expression.

scholarly journals Upstream stimulatory factor regulates Pdx-1 gene expression in differentiated pancreatic β-cells

1999 ◽  
Vol 341 (2) ◽  
pp. 315-322 ◽  
Author(s):  
Jin QIAN ◽  
Elizabeth N. KAYTOR ◽  
Howard C. TOWLE ◽  
L. Karl OLSON
Keyword(s):  
Gene Expression ◽  
Gene Promoter ◽  
Dominant Negative ◽  
Mrna Levels ◽  
Upstream Stimulatory Factor ◽  
Β Cells ◽  
Pancreatic Β Cells ◽  
Protein Levels ◽  
E Box ◽  
Stimulatory Factor

The homeobox gene Pdx-1 plays a key role in the development of the pancreas. In the adult, however, expression of the Pdx-1 gene is restricted to pancreatic β-cells and endocrine cells of duodenal epithelium. Recently, the transcription factor, upstream stimulatory factor (USF), has been shown to bind invitro to a mutationally sensitive E-box motif within the 5′-flanking region of the Pdx-1 gene [Sharma, Leonard, Lee, Chapman, Leiter and Montminy (1996) J. Biol. Chem. 271, 2294-2299]. In the present study, we show that USF not only binds to the Pdx-1 gene promoter but also functionally regulates the expression of the Pdx-1 gene in differentiated pancreatic β-cells. Adenovirus-mediated overexpression of a dominant negative form of USF2 decreased binding of endogenous USF to the E-box element by ~ 90%. This reduction in endogenous USF binding led to a greater than 50% decrease in Pdx-1 gene promoter activity, which, in turn, resulted in marked reductions in Pdx-1 mRNA and protein levels. Importantly, the lower Pdx-1 protein levels led to a greater than 50% reduction in Pdx-1 binding activity to the A3 element on the insulin gene promoter, and a significant reduction in insulin mRNA levels. Overall, our results show that USF functionally regulates Pdx-1 gene expression in differentiated pancreatic β-cells and provide the first functional data for a role of USF in the regulation of a normal cellular gene.

scholarly journals Bioinformatics Analysis of SNPs in IL-6 Gene Promoter of Jinghai Yellow Chickens

Genes ◽  
2018 ◽  
Vol 9 (9) ◽  
pp. 446 ◽  
Author(s):  
Shijie Xin ◽  
Xiaohui Wang ◽  
Guojun Dai ◽  
Jingjing Zhang ◽  
Tingting An ◽  
...  
Keyword(s):  
Gene Expression ◽  
Transcription Factor ◽  
Binding Sites ◽  
Promoter Region ◽  
Bioinformatics Analysis ◽  
Cpg Islands ◽  
Regulatory Region ◽  
Transcription Factor Binding Sites ◽  
Transcription Factor Binding ◽  
Factor Binding

The proinflammatory cytokine, interleukin-6 (IL-6), plays a critical role in many chronic inflammatory diseases, particularly inflammatory bowel disease. To investigate the regulation of IL-6 gene expression at the molecular level, genomic DNA sequencing of Jinghai yellow chickens (Gallus gallus) was performed to detect single-nucleotide polymorphisms (SNPs) in the region −2200 base pairs (bp) upstream to 500 bp downstream of IL-6. Transcription factor binding sites and CpG islands in the IL-6 promoter region were predicted using bioinformatics software. Twenty-eight SNP sites were identified in IL-6. Four of these 28 SNPs, three [−357 (G > A), −447 (C > G), and −663 (A > G)] in the 5′ regulatory region and one in the 3′ non-coding region [3177 (C > T)] are not labelled in GenBank. Bioinformatics analysis revealed 11 SNPs within the promoter region that altered putative transcription factor binding sites. Furthermore, the C-939G mutation in the promoter region may change the number of CpG islands, and SNPs in the 5′ regulatory region may influence IL-6 gene expression by altering transcription factor binding or CpG methylation status. Genetic diversity analysis revealed that the newly discovered A-663G site significantly deviated from Hardy-Weinberg equilibrium. These results provide a basis for further exploration of the promoter function of the IL-6 gene and the relationships of these SNPs to intestinal inflammation resistance in chickens.

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