Spatially specific expression of Hoxb4 is dependent on the ubiquitous transcription factor NFY

Development ◽  
2002 ◽  
Vol 129 (16) ◽  
pp. 3887-3899 ◽  
Author(s):  
Jonathan Gilthorpe ◽  
Marie Vandromme ◽  
Tim Brend ◽  
Alejandro Gutman ◽  
Dennis Summerbell ◽  
...  
Keyword(s):  
Gene Expression ◽  
Binding Sites ◽  
Specific Interaction ◽  
Transcriptional Regulators ◽  
Heterologous Promoter ◽  
Specific Expression ◽  
Enhancer Activity ◽  
Hox Gene ◽  
Two Factors ◽  
Enhancer Function

Understanding how boundaries and domains of Hox gene expression are determined is critical to elucidating the means by which the embryo is patterned along the anteroposterior axis. We have performed a detailed analysis of the mouse Hoxb4 intron enhancer to identify upstream transcriptional regulators. In the context of an heterologous promoter, this enhancer can establish the appropriate anterior boundary of mesodermal expression but is unable to maintain it, showing that a specific interaction with its own promoter is important for maintenance. Enhancer function depends on a motif that contains overlapping binding sites for the transcription factors NFY and YY1. Specific mutations that either abolish or reduce NFY binding show that it is crucial for enhancer activity. The NFY/YY1 motif is reiterated in the Hoxb4 promoter and is known to be required for its activity. As these two factors are able to mediate opposing transcriptional effects by reorganizing the local chromatin environment, the relative levels of NFY and YY1 binding could represent a mechanism for balancing activation and repression of Hoxb4 through the same site.

Regionalization of Sonic hedgehog transcription along the anteroposterior axis of the mouse central nervous system is regulated by Hnf3-dependent and -independent mechanisms

Development ◽  
1999 ◽  
Vol 126 (2) ◽  
pp. 281-292 ◽  
Author(s):  
D.J. Epstein ◽  
A.P. McMahon ◽  
A.L. Joyner
Keyword(s):  
Gene Expression ◽  
Sonic Hedgehog ◽  
Binding Sites ◽  
Transcriptional Regulators ◽  
Regulatory Sequences ◽  
Reporter Expression ◽  
Enhancer Activity ◽  
Ventral Midline ◽  
Genomic Clones ◽  
Anteroposterior Axis

The axial midline mesoderm and the ventral midline of the neural tube, the floor plate, share the property of being a source of the secreted protein, Sonic hedgehog (Shh), which has the capacity to induce a variety of ventral cell types along the length of the mouse CNS. To gain insight into the mechanisms by which Shh transcription is initiated in these tissues, we set out to identify the cis-acting sequences regulating Shh gene expression. As an approach, we have tested genomic clones encompassing 35 kb of the Shh locus for their ability to direct a lacZ reporter gene to the temporally and spatially restricted confines of the Shh expression domains in transgenic mice. Three enhancers were identified that directed lacZ expression to distinct regions along the anteroposterior axis including the ventral midline of the spinal cord, hindbrain, rostral midbrain and caudal diencephalon, suggesting that multiple transcriptional regulators are required to initiate Shh gene expression within the CNS. In addition, regulatory sequences were also identified that directed reporter expression to the notochord, albeit, under limited circumstances. Sequence analysis of the genomic clones responsible for enhancer activity from a variety of organisms, including mouse, chicken and human, have identified highly conserved binding sites for the hepatocyte nuclear factor 3 (Hnf3) family of transcriptional regulators in some, but not all, of the enhancers. Moreover, the generation of mutations in the Hnf3-binding sites showed their requirement in certain, but not all, aspects of Shh reporter expression. Taken together, our results support the existence of Hnf3-dependent and -independent mechanisms in the direct activation of Shh transcription within the CNS and axial mesoderm.

scholarly journals Molecular analysis of the zeste-white interaction reveals a promoter-proximal element essential for distant enhancer-promoter communication.

Genetics ◽  
1992 ◽  
Vol 131 (1) ◽  
pp. 79-90 ◽  
Author(s):  
S Qian ◽  
B Varjavand ◽  
V Pirrotta
Keyword(s):  
Transcription Start Site ◽  
Binding Sites ◽  
Start Site ◽  
Transcription Start ◽  
Heterologous Promoter ◽  
Specific Expression ◽  
Enhancer Activity ◽  
Directed Mutation ◽  
Sequence Deletion ◽  
Promoter Interaction

Abstract We have analyzed the eye and testis enhancers located 1 kb upstream of the transcription start site of the white gene. Both enhancers confer the corresponding tissue-specific expression on a heterologous promoter as well as on the white promoter. The eye determinant consists of multiple elements, each able to stimulate eye-specific expression. It also contains five binding sites for the zeste protein while the immediately adjacent testis element contains none. Site-directed mutation of these zeste binding sites abolishes the zeste-white interaction but does not significantly affect the eye enhancer activity, indicating that they are not important for the eye enhancer activity per se. Other zeste binding sites just upstream of the promoter are not necessary for the zeste-white interaction. We conclude that the overlap of the eye enhancer with the zeste binding sites is responsible for the zeste-white interaction and explains why this interaction affects eye but not testis expression. Sequence deletion or substitution experiments suggested that the white promoter is internal to the transcription start site; the zeste protein is not required for distant enhancer action but a 95-bp promoter-proximal sequence is essential for distant enhancer-promoter interaction. This element may serve as an anchor to stabilize formation of a loop that brings the enhancer to the vicinity of the promoter.

scholarly journals An unexpected contribution of lincRNA splicing to enhancer function

2018 ◽  
Author(s):  
Jennifer Y. Tan ◽  
Adriano Biasini ◽  
Robert S. Young ◽  
Ana C. Marques
Keyword(s):  
Gene Expression ◽  
Target Gene ◽  
Gene Expression Regulation ◽  
Transcriptional Regulators ◽  
Regulatory Function ◽  
Nucleotide Polymorphisms ◽  
Chromosomal Dna ◽  
Neighboring Gene ◽  
Enhancer Activity ◽  
Enhancer Function

ABSTRACTTranscription is common at active mammalian enhancers sometimes giving rise to stable and unidirectionally transcribed enhancer-associated long intergenic noncoding RNAs (elincRNAs). ElincRNA expression is associated with changes in neighboring gene product abundance and local chromosomal topology, suggesting that transcription at these loci contributes to gene expression regulation in cis. Despite the lack of evidence supporting sequence-dependent functions for most elincRNAs, splicing of these transcripts is unexpectedly common. Whether elincRNA splicing is a mere consequence of their cognate enhancer activity or if it directly impacts enhancer-associated cis-regulation remains unanswered.Here we show that elincRNAs are efficiently and rapidly spliced and that their processing rate is strongly associated with their cognate enhancer activity. This association is supported by: their enrichment in enhancer-specific chromatin signatures; elevated binding of co-transcriptional regulators, including CBP and p300; increased local intra-chromosomal DNA contacts; and strengthened cis-regulation on target gene expression. Using nucleotide polymorphisms at elincRNA splice sites, we found that elincRNA splicing enhances their transcription and directly impacts cis-regulatory function of their cognate enhancers. Importantly, up to 90% of human elincRNAs have nucleotide variants that are associated with both their splicing and the expression levels of their proximal genes.Our results highlight an unexpected contribution of elincRNA splicing to enhancer function.

The snail repressor establishes a muscle/notochord boundary in the Ciona embryo

Development ◽  
1998 ◽  
Vol 125 (13) ◽  
pp. 2511-2520 ◽  
Author(s):  
S. Fujiwara ◽  
J.C. Corbo ◽  
M. Levine
Keyword(s):  
Gene Expression ◽  
Dna Binding ◽  
Binding Sites ◽  
Promoter Region ◽  
Muscle Cells ◽  
Specific Pattern ◽  
Binding Assays ◽  
Heterologous Promoter ◽  
Present Evidence ◽  
Tail Muscle

Previous studies have identified a minimal 434 bp enhancer from the promoter region of the Ciona Brachyury gene (Ci-Bra), which is sufficient to direct a notochord-specific pattern of gene expression. Here we present evidence that a Ciona homolog of snail (Ci-sna) encodes a repressor of the Ci-Bra enhancer in the tail muscles. DNA-binding assays identified four Ci-Sna-binding sites in the Ci-Bra enhancer, and mutations in these sites cause otherwise normal Ci-Bra/lacZ transgenes to be misexpressed in ectopic tissues, particularly the tail muscles. Selective misexpression of Ci-sna using a heterologous promoter results in the repression of Ci-Bra/lacZ transgenes in the notochord. Moreover, the conversion of the Ci-Sna repressor into an activator results in the ectopic induction of Ci-Bra/lacZ transgenes in the muscles, and also causes an intermixing of notochord and muscle cells during tail morphogenesis. These results suggest that Ci-Sna functions as a boundary repressor, which subdivides the mesoderm into separate notochord and tail muscle lineages.

scholarly journals Regulation of the human T-cell receptor alpha gene enhancer: multiple ubiquitous and T-cell-specific nuclear proteins interact with four hypomethylated enhancer elements.

1990 ◽  
Vol 10 (9) ◽  
pp. 4720-4727 ◽  
Author(s):  
I C Ho ◽  
J M Leiden
Keyword(s):  
T Cell ◽  
Protein Binding ◽  
Binding Sites ◽  
Nuclear Protein ◽  
Cell Receptor ◽  
Enhancer Activity ◽  
Protein Binding Sites ◽  
Human T Cell ◽  
Alpha Gene ◽  
Enhancer Function

Transcription of human T-cell receptor (TCR) alpha genes is regulated by a T-cell-specific transcriptional enhancer that is located 4.5 kilobases 3' of the C alpha gene segment. Previous studies have demonstrated that this enhancer contains at least five nuclear protein-binding sites called T alpha 1 to T alpha 5. In the studies described in this report, we have determined the molecular requirements for human TCR alpha enhancer function. In vitro mutagenesis and deletion analyses demonstrated that full enhancer activity is retained in a 116-base-pair fragment containing the T alpha 1 and T alpha 2 nuclear protein-binding sites and that both of these sites are required for full enhancer function. Functional enhancer activity requires that the T alpha 1 and T alpha 2 binding sites be separated by more than 15 and fewer than 85 base pairs. However, the sequence of this spacer region and the relative phase of the two binding sites on the DNA helix do not affect enhancer function. Deletion and mutation analyses demonstrated that the T alpha 3 and T alpha 4 nuclear protein-binding sites are not necessary or sufficient for TCR alpha enhancer activity. However, a fragment containing these two sites was able to compensate for T alpha 1 and T alpha 2 mutations that otherwise abolished enhancer activity. Electrophoretic mobility shift analyses of the TCR alpha enhancer binding proteins revealed that the T alpha 1, T alpha 3, and T alpha 4 binding proteins are expressed in a variety of T-cell and non-T-cell tumor cell lines. In contrast, one of the two T alpha 2 binding activities was detected only in T-cell nuclear extracts. The activity of the TCR alpha enhancer does not appear to be regulated solely at the level of DNA methylation on that the enhancer sequences were found to be identically hypomethylated in B and T cells as compared with fibroblasts. Taken together, these results suggest that TCR alpha enhancer activity is regulated by the interaction of multiple T-cell-specific and ubiquitous nuclear proteins with partially redundant cis-acting enhancer elements that are hypomethylated in cells of the lymphoid lineage.

scholarly journals Direct and widespread role for the nuclear receptor EcR in mediating the response to ecdysone in Drosophila

2019 ◽  
Vol 116 (20) ◽  
pp. 9893-9902 ◽  
Author(s):  
Christopher M. Uyehara ◽  
Daniel J. McKay
Keyword(s):  
Gene Expression ◽  
Transcription Factor ◽  
Nuclear Receptor ◽  
Binding Sites ◽  
Transcriptional Responses ◽  
Enhancer Activity ◽  
Developmental Transitions ◽  
Experimental Systems ◽  
The Impact

The ecdysone pathway was among the first experimental systems employed to study the impact of steroid hormones on the genome. In Drosophila and other insects, ecdysone coordinates developmental transitions, including wholesale transformation of the larva into the adult during metamorphosis. Like other hormones, ecdysone controls gene expression through a nuclear receptor, which functions as a ligand-dependent transcription factor. Although it is clear that ecdysone elicits distinct transcriptional responses within its different target tissues, the role of its receptor, EcR, in regulating target gene expression is incompletely understood. In particular, EcR initiates a cascade of transcription factor expression in response to ecdysone, making it unclear which ecdysone-responsive genes are direct EcR targets. Here, we use the larval-to-prepupal transition of developing wings to examine the role of EcR in gene regulation. Genome-wide DNA binding profiles reveal that EcR exhibits widespread binding across the genome, including at many canonical ecdysone response genes. However, the majority of its binding sites reside at genes with wing-specific functions. We also find that EcR binding is temporally dynamic, with thousands of binding sites changing over time. RNA-seq reveals that EcR acts as both a temporal gate to block precocious entry to the next developmental stage as well as a temporal trigger to promote the subsequent program. Finally, transgenic reporter analysis indicates that EcR regulates not only temporal changes in target enhancer activity but also spatial patterns. Together, these studies define EcR as a multipurpose, direct regulator of gene expression, greatly expanding its role in coordinating developmental transitions.

scholarly journals A non-coding genetic variant maximally associated with serum urate levels is functionally linked to HNF4A-dependent PDZK1 expression

2018 ◽  
Author(s):  
Sarada Ketharnathan ◽  
Megan Leask ◽  
James Boocock ◽  
Amanda J. Phipps-Green ◽  
Jisha Antony ◽  
...  
Keyword(s):  
Gene Expression ◽  
Hepg2 Cells ◽  
Molecular Mechanisms ◽  
Genetic Variant ◽  
Fluorescent Protein ◽  
Serum Urate ◽  
Specific Gene ◽  
Specific Expression ◽  
Enhancer Activity ◽  
Tissue Specific

ABSTRACTSeveral dozen genetic variants associate with serum urate levels, but the precise molecular mechanisms by which they affect serum urate are unknown. Here we tested for functional linkage of the maximally-associated genetic variant rs1967017 at the PDZK1 locus to elevated PDZK1 expression.We performed expression quantitative trait locus (eQTL) and likelihood analyses followed by gene expression assays. Zebrafish were used to determine the ability of rs1967017 to direct tissue-specific gene expression. Luciferase assays in HEK293 and HepG2 cells measured the effect of rs1967017 on transcription amplitude.PAINTOR analysis revealed rs1967017 as most likely to be causal and rs1967017 was an eQTL for PDZK1 in the intestine. The region harboring rs1967017 was capable of directly driving green fluorescent protein expression in the kidney, liver and intestine of zebrafish embryos, consistent with a conserved ability to confer tissue-specific expression. The urate-increasing T-allele of rs1967017 strengthens a binding site for the transcription factor HNF4A. siRNA depletion of HNF4A reduced endogenous PDZK1 expression in HepG2 cells. Luciferase assays showed that the T-allele of rs1967017 gains enhancer activity relative to the urate-decreasing C-allele, with T-allele enhancer activity abrogated by HNF4A depletion. HNF4A physically binds the rs1967017 region, suggesting direct transcriptional regulation of PDZK1 by HNF4A.With other reports our data predict that the urate-raising T-allele of rs1967017 enhances HNF4A binding to the PDZK1 promoter, thereby increasing PDZK1 expression. As PDZK1 is a scaffold protein for many ion channel transporters, increased expression can be predicted to increase activity of urate transporters and alter excretion of urate.

scholarly journals Gene expression evolution in pattern-triggered immunity within Arabidopsis thaliana and across Brassicaceae species

2020 ◽  
Author(s):  
Thomas M. Winkelmüller ◽  
Frederickson Entila ◽  
Shajahan Anver ◽  
Anna Piasecka ◽  
Baoxing Song ◽  
...  
Keyword(s):  
Gene Expression ◽  
Arabidopsis Thaliana ◽  
Binding Sites ◽  
Microbial Control ◽  
Purifying Selection ◽  
Wrky Transcription Factor ◽  
Specific Gene ◽  
Specific Expression ◽  
Brassicaceae Species ◽  
Species Specific

AbstractPlants recognize surrounding microbes by sensing microbe-associated molecular patterns (MAMPs) to activate pattern-triggered immunity (PTI). Despite their significance for microbial control, the evolution of PTI responses remains largely uncharacterized. Employing comparative transcriptomics of six Arabidopsis thaliana accessions and three additional Brassicaceae species for PTI responses to the MAMP flg22, we identified a set of genes with expression changes under purifying selection in the Brassicaceae species and genes exhibiting species-specific expression signatures. Variation in flg22-triggered transcriptome and metabolome responses across Brassicaceae species was incongruent with their phylogeny while expression changes were strongly conserved within A. thaliana, suggesting directional selection for some species-specific gene expression. We found the enrichment of WRKY transcription factor binding sites in 5’-regulatory regions in conserved and species-specific responsive genes, linking the emergence of WRKY-binding sites with the evolution of gene responses in PTI. Our findings advance our understanding of transcriptome evolution during biotic stress.

scholarly journals target: an R package to predict combined function of transcription factors

F1000Research ◽  
2021 ◽  
Vol 10 ◽  
pp. 344
Author(s):  
Mahmoud Ahmed ◽  
Deok Ryong Kim
Keyword(s):  
Gene Expression ◽  
Transcription Factor ◽  
Transcription Factors ◽  
Binding Sites ◽  
R Package ◽  
Expression Data ◽  
Bioconductor Package ◽  
Chip Experiment ◽  
Binding Data ◽  
Two Factors

Researchers use ChIP binding data to identify potential transcription factor binding sites. Similarly, they use gene expression data from sequencing or microarrays to quantify the effect of the factor overexpression or knockdown on its targets. Therefore, the integration of the binding and expression data can be used to improve the understanding of a transcription factor function. Here, we implemented the binding and expression target analysis (BETA) in an R/Bioconductor package. This algorithm ranks the targets based on the distances of their assigned peaks from the factor ChIP experiment and the signed statistics from gene expression profiling with factor perturbation. We further extend BETA to integrate two sets of data from two factors to predict their targets and their combined functions. In this article, we briefly describe the workings of the algorithm and provide a workflow with a real dataset for using it. The gene targets and the aggregate functions of transcription factors YY1 and YY2 in HeLa cells were identified. Using the same datasets, we identified the shared targets of the two factors, which were found to be, on average, more cooperatively regulated.

scholarly journals Characterization of sequence determinants of enhancer function using natural genetic variation

2021 ◽  
Author(s):  
Marty G. Yang ◽  
Emi Ling ◽  
Christopher J. Cowley ◽  
Michael E. Greenberg ◽  
Thomas Vierbuchen
Keyword(s):  
Binding Sites ◽  
Sequence Variation ◽  
Regulatory Elements ◽  
Human Populations ◽  
Sequence Variant ◽  
Sequence Variants ◽  
Enhancer Activity ◽  
F1 Hybrid ◽  
Enhancer Function ◽  
The Impact

Sequence variation in enhancers, a class of cis-regulatory elements that control cell type-specific gene transcription, contributes significantly to phenotypic variation within human populations. Enhancers are short DNA sequences (~200 bp) composed of multiple binding sites (4-10 bp) for transcription factors (TFs). The transcriptional regulatory activity of an enhancer is encoded by the type, number, and distribution of TF binding sites that it contains. However, the sequence determinants of TF binding to enhancers and the relationship between TF binding and enhancer activity are complex, and thus it remains difficult to predict the effect of any given sequence variant on enhancer function. Here, we generate allele-specific maps of TF binding and enhancer activity in fibroblasts from a panel of F1 hybrid mice that have a high frequency of sequence variants. We identified thousands of enhancers that exhibit differences in TF binding and/or activity between alleles and use these data to define features of sequence variants that are most likely to impact enhancer function. Our data demonstrate a critical role for AP-1 TFs at many fibroblast enhancers, reveal a hierarchical relationship between AP-1 and TEAD TF binding at enhancers, and delineate the nature of sequence variants that contribute to AP-1 TF binding. These data represent one of the most comprehensive assessments to date of the impact of sequence variation on enhancer function in chromatin, with implications for identifying functional cis-regulatory variation in human populations.

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