Missing Links: DNA‐Binding and Target Gene Specificity of Floral Homeotic Proteins

2006 ◽  
pp. 209-236 ◽  
Author(s):  
Rainer Melzer ◽  
Kerstin Kaufmann ◽  
Günter Theißen
Keyword(s):  
Dna Binding ◽  
Target Gene ◽  
Floral Homeotic

scholarly journals Novel DNA Motif Binding Activity Observed In Vivo With an Estrogen Receptor α Mutant Mouse

2014 ◽  
Vol 28 (6) ◽  
pp. 899-911 ◽  
Author(s):  
Sylvia C. Hewitt ◽  
Leping Li ◽  
Sara A. Grimm ◽  
Wipawee Winuthayanon ◽  
Katherine J. Hamilton ◽  
...  
Keyword(s):  
Dna Binding ◽  
Target Gene ◽  
Mutant Mouse ◽  
Estrogen Receptor Α ◽  
Binding Activity ◽  
Estrogen Response Element ◽  
Dna Binding Domain ◽  
Estrogen Response ◽  
Uterine Growth

Abstract Estrogen receptor α (ERα) interacts with DNA directly or indirectly via other transcription factors, referred to as “tethering.” Evidence for tethering is based on in vitro studies and a widely used “KIKO” mouse model containing mutations that prevent direct estrogen response element DNA- binding. KIKO mice are infertile, due in part to the inability of estradiol (E2) to induce uterine epithelial proliferation. To elucidate the molecular events that prevent KIKO uterine growth, regulation of the pro-proliferative E2 target gene Klf4 and of Klf15, a progesterone (P4) target gene that opposes the pro-proliferative activity of KLF4, was evaluated. Klf4 induction was impaired in KIKO uteri; however, Klf15 was induced by E2 rather than by P4. Whole uterine chromatin immunoprecipitation-sequencing revealed enrichment of KIKO ERα binding to hormone response elements (HREs) motifs. KIKO binding to HRE motifs was verified using reporter gene and DNA-binding assays. Because the KIKO ERα has HRE DNA-binding activity, we evaluated the “EAAE” ERα, which has more severe DNA-binding domain mutations, and demonstrated a lack of estrogen response element or HRE reporter gene induction or DNA-binding. The EAAE mouse has an ERα null–like phenotype, with impaired uterine growth and transcriptional activity. Our findings demonstrate that the KIKO mouse model, which has been used by numerous investigators, cannot be used to establish biological functions for ERα tethering, because KIKO ERα effectively stimulates transcription using HRE motifs. The EAAE-ERα DNA-binding domain mutant mouse demonstrates that ERα DNA-binding is crucial for biological and transcriptional processes in reproductive tissues and that ERα tethering may not contribute to estrogen responsiveness in vivo.

scholarly journals p53 dynamically directs TFIID assembly on target gene promoters

2016 ◽  
Author(s):  
R. A. Coleman ◽  
Z. Qiao ◽  
S. K. Singh ◽  
C. S. Peng ◽  
M. Cianfrocco ◽  
...  
Keyword(s):  
Dna Binding ◽  
Single Molecule ◽  
Transcription Initiation ◽  
Gene Networks ◽  
Target Gene ◽  
Core Promoter ◽  
Gene Promoters ◽  
Binding Modes ◽  
Dissociation Kinetics ◽  
Target Promoters

AbstractThe p53 tumor suppressor protein is a central regulator that turns on vast gene networks to maintain cellular integrity upon various stimuli. p53 activates transcription initiation in part by aiding recruitment of TFIID to the promoter. However, the precise means by which p53 dynamically interacts with TFIID to facilitate assembly on target gene promoters remains elusive. To address this key question, we have undertaken an integrated approach involving single molecule fluorescence microscopy, single particle cryo-electron microscopy, and biochemistry. Our real-time single molecule imaging demonstrates that TFIID alone binds poorly to native p53 target promoters. p53 unlocks TFIID’s ability to bind DNA by increasing TFIID contacts with both the core promoter and a region surrounding p53’s response element (RE). Analysis of single molecule dissociation kinetics reveals that TFIID interacts with promoters via transient and prolonged DNA binding modes that are each regulated by p53. Importantly, our structural work reveals that TFIID’s conversion from a canonical form to a rearranged DNA-binding conformation is enhanced in the presence of DNA and p53. Notably, TFIID’s interaction with DNA induces p53 to rapidly dissociate, effectively liberating the RE on the promoter. Collectively, these findings indicate that p53 dynamically escorts and loads the basal transcription machinery onto its target promoters.

scholarly journals Differential Contribution of N- and C-Terminal Regions of HIF1α and HIF2α to Their Target Gene Selectivity

2020 ◽  
Vol 21 (24) ◽  
pp. 9401
Author(s):  
Antonio Bouthelier ◽  
Florinda Meléndez-Rodríguez ◽  
Andrés A. Urrutia ◽  
Julián Aragonés
Keyword(s):  
Transcription Factors ◽  
Dna Binding ◽  
Cellular Response ◽  
Target Gene ◽  
Target Genes ◽  
Transactivation Domain ◽  
Transactivation Domains ◽  
Relative Contribution

Cellular response to hypoxia is controlled by the hypoxia-inducible transcription factors HIF1α and HIF2α. Some genes are preferentially induced by HIF1α or HIF2α, as has been explored in some cell models and for particular sets of genes. Here we have extended this analysis to other HIF-dependent genes using in vitro WT8 renal carcinoma cells and in vivo conditional Vhl-deficient mice models. Moreover, we generated chimeric HIF1/2 transcription factors to study the contribution of the HIF1α and HIF2α DNA binding/heterodimerization and transactivation domains to HIF target specificity. We show that the induction of HIF1α-dependent genes in WT8 cells, such as CAIX (CAR9) and BNIP3, requires both halves of HIF, whereas the HIF2α transactivation domain is more relevant for the induction of HIF2 target genes like the amino acid carrier SLC7A5. The HIF selectivity for some genes in WT8 cells is conserved in Vhl-deficient lung and liver tissue, whereas other genes like Glut1 (Slc2a1) behave distinctly in these tissues. Therefore the relative contribution of the DNA binding/heterodimerization and transactivation domains for HIF target selectivity can be different when comparing HIF1α or HIF2α isoforms, and that HIF target gene specificity is conserved in human and mouse cells for some of the genes analyzed.

Copper affects the binding of HIF-1α to the critical motifs of its target genes

Metallomics ◽  
2019 ◽  
Vol 11 (2) ◽  
pp. 429-438 ◽  
Author(s):  
Zhijuan Wu ◽  
Wenjing Zhang ◽  
Y. James Kang
Keyword(s):  
Dna Binding ◽  
Target Gene ◽  
Target Genes ◽  
Gene Selection ◽  
Hypoxic Conditions ◽  
Selection Of

Copper regulates the target gene selection of HIF-1α under hypoxic conditions by affecting HIF-1α-DNA binding patterns across the genome.

Smad4 Inhibits Nup98-Hoxa9 Transcriptional Activity by Interacting at Hox Homeodomain.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 2570-2570
Author(s):  
Ning Wang ◽  
Mei Wan ◽  
Yi Tang ◽  
Hyung G. Kim ◽  
Claudia Cotta ◽  
...  
Keyword(s):  
Dna Binding ◽  
Fusion Protein ◽  
Progenitor Cells ◽  
Myeloid Leukemia ◽  
Target Gene ◽  
Luciferase Reporter ◽  
Hox Proteins ◽  
Hematopoietic Stem ◽  
Hox Gene ◽  
Nih 3T3

Abstract Homeobox (Hox) genes play crucial roles in hematopoiesis. Alterations of Hox gene expressions are found in patients with hematological malignancy. Nup98-Hoxa9, a fusion protein combining the N-terminus of Nup98 and the C-terminus of Hoxa9 resulting from t (7; 11) chromosomal translocation, is the first evidence of Hox gene involvement in human leukemia. We have shown that BMP specific R-Smads and Smad4 interact with Hox proteins including Hoxa9, and the interaction blocks binding of Hox proteins to their DNA elements. Here, we show that Smad4, the common signal transducer of both TGF-β and BMP, interacts with the homeodomain of Nup98-Hoxa9 in a coimmunoprecipitation assay. Furthermore, this interaction abolished the DNA binding ability of Nup98-Hoxa9 to DNA in both EMSA using purified GST fusion protein and in in vivo chromatin immunoprecipitation assays. Subsequent mapping data revealed the MH1 domain of Smad4 mediated the interaction. These findings suggest that Smad4 could be used to inhibit Hox-induced leukemia. We then attempted to examine whether Smad4 could inhibit the downstream target gene by Nup98-Hoxa9. Although Nup98-Hoxa9 is believed to function as an aberrant DNA-binding transcriptional activator, downstream genes that contribute to leukemogenesis are largely unknown. It has been reported that Nup98-Hoxa9 induces Hoxa9 expression in murine primary bone marrow cells. Accumulating evidence suggests that Hoxa9 plays important roles in normal and malignant hematopoiesis. Enforced expression of Hoxa9 causes proliferative expansion of primitive hematopoietic stem/progenitor cells and subsequently inhibits their differentiation. Aberrant expression of HOXA9 due to retroviral integration induced acute myeloid leukemia (AML) in mice. By visually scanning the sequence of the Hoxa9 promoter, we found many TGAT and TTA (T/C) consensus sequences, which are preferentially recognized by the heterodimer of PBX protein and the proteins of the Abd-B-like gene family, including Hoxa9, respectively. To test whether Hoxa9 could be a direct transcriptional target for Nup98-Hoxa9, we used a Hoxa9 promoter/luciferase reporter construct along with the expression plasmid pcDNA-Nup98/Hoxa9 to transiently transfect NIH/3T3 fibroblast cells and Ba/F3 murine hematopoietic progenitor cells. Nup98-Hoxa9 showed substantial transcription activation on Hoxa9 promoter in these cell lines. This activation is further enhanced by cotransfection of PBX1a, which stabilizes the binding of Nup98-Hoxa9 to DNA. Nup98-Hoxa9 also bound to radiolabelled DNA oligo from the Hoxa9 promoter in vitro by EMSA assay. These data indicate that Nup98-Hoxa9 directly activates the Hoxa9 promoter. Interestingly, Smad4 removed the binding of Nup98-Hoxa9 to the Hoxa9 promoter. Co-overexpression of Smad4 inhibited the transactivation activity of Nup98-Hoxa9 on the Hoxa9 promoter regardless of the presence of PBX1a. In NIH/3T3 cells, Smad4 inhibited Nup98-Hoxa9-induced Hoxa9 expression both at the mRNA and protein levels. In summary, we show that Smad4 inhibited DNA binding activity of Nup98-Hoxa9 by interacting at Hox homeodomain. This interaction abolished the transcription activity of Nup98-Hoxa9 on its target gene, thus preventing Nup98-Hoxa9 from working as a transcription factor, which could lead to a new therapeutic target for the treatment of myeloid leukemia caused by Nup98-Hoxa9.

Arsenite inhibits p53 phosphorylation, DNA binding activity, and p53 target gene p21 expression in mouse epidermal JB6 cells,

2006 ◽  
Vol 45 (11) ◽  
pp. 861-870 ◽  
Author(s):  
Faqing Tang ◽  
Guangming Liu ◽  
Zhiwei He ◽  
Wei-Ya Ma ◽  
Ann M. Bode ◽  
...  
Keyword(s):  
Dna Binding ◽  
Target Gene ◽  
Binding Activity ◽  
Dna Binding Activity ◽  
P53 Target Gene ◽  
Jb6 Cells

scholarly journals p53 Dynamically Directs TFIID Assembly on Target Gene Promoters

2017 ◽  
Vol 37 (13) ◽  
Author(s):  
R. A. Coleman ◽  
Z. Qiao ◽  
S. K. Singh ◽  
C. S. Peng ◽  
M. Cianfrocco ◽  
...  
Keyword(s):  
Dna Binding ◽  
Single Molecule ◽  
Transcription Initiation ◽  
Gene Networks ◽  
Target Gene ◽  
Core Promoter ◽  
Gene Promoters ◽  
Imaging Data ◽  
Element Analysis ◽  
Target Promoters

ABSTRACT p53 is a central regulator that turns on vast gene networks to maintain cellular integrity in the presence of various stimuli. p53 activates transcription initiation in part by aiding recruitment of TFIID to the promoter. However, the precise means by which p53 dynamically interacts with TFIID to facilitate assembly on target gene promoters remains elusive. To address this key issue, we have undertaken an integrated approach involving single-molecule fluorescence microscopy, single-particle cryo-electron microscopy, and biochemistry. Our real-time single-molecule imaging data demonstrate that TFIID alone binds poorly to native p53 target promoters. p53 unlocks TFIID's ability to bind DNA by stabilizing TFIID contacts with both the core promoter and a region within p53's response element. Analysis of single-molecule dissociation kinetics reveals that TFIID interacts with promoters via transient and prolonged DNA binding modes that are each regulated by p53. Importantly, our structural work reveals that TFIID's conversion to a rearranged DNA binding conformation is enhanced in the presence of DNA and p53. Notably, TFIID's interaction with DNA induces p53 to rapidly dissociate, which likely leads to additional rounds of p53-mediated recruitment of other basal factors. Collectively, these findings indicate that p53 dynamically escorts and loads TFIID onto its target promoters.

scholarly journals Pbx-Hox Heterodimers Recruit Coactivator-Corepressor Complexes in an Isoform-Specific Manner

1999 ◽  
Vol 19 (12) ◽  
pp. 8219-8225 ◽  
Author(s):  
Hiroshi Asahara ◽  
Sanjoy Dutta ◽  
Hung-Ying Kao ◽  
Ronald M. Evans ◽  
Marc Montminy
Keyword(s):  
Gene Expression ◽  
Dna Binding ◽  
Cell Fate ◽  
Target Gene ◽  
Biological Action ◽  
Binding Activity ◽  
Creb Binding Protein ◽  
Target Gene Expression

ABSTRACT Homeobox (hox) proteins have been shown to regulate cell fate and segment identity by promoting the expression of specific genetic programs. In contrast to their restricted biological action in vivo, however, most homeodomain factors exhibit promiscuous DNA binding properties in vitro, suggesting a requirement for additional cofactors that enhance target site selectivity. In this regard, thepbx family of homeobox genes has been found to heterodimerize with and thereby augment the DNA binding activity of certain hox proteins on a subset of potential target sites. Here we examine the transcriptional properties of a forcedhox-pbx heterodimer containing the pancreas-specific orphan homeobox factor pdx fused to pbx-1a. Compared to the pdx monomer, the forced pdx-pbx1a dimer, displayed 10- to 20-fold-higher affinity for a consensushox-pbx binding site but was completely unable to bind ahox monomer recognition site. The pdx-pbx dimer stimulated target gene expression via an N-terminaltrans-activation domain in pdx that interacts with the coactivator CREB binding protein. The pdx-pbxdimer was also found to repress transcription via a C-terminal domain in pbx-1a that associates with the corepressors SMRT and NCoR. The transcriptional properties of the pdx-pbx1complex appear to be regulated at the level of alternative splicing; apdx-pbx polypeptide containing the pbx1bisoform, which lacks the C-terminal extension in pbx1a, was unable to repress target gene expression via NCoR-SMRT. Sincepbx1a and pbx1b are differentially expressed in endocrine versus exocrine compartments of the adult pancreas, our results illustrate a novel mechanism by which pbx proteins may modulate the expression of specific genetic programs, either positively or negatively, during development.

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