scholarly journals The Retinoblastoma Protein Binds the Promoter of the Survival Gene bcl-2 and Regulates Its Transcription in Epithelial Cells through Transcription Factor AP-2

2002 ◽  
Vol 22 (22) ◽  
pp. 7877-7888 ◽  
Author(s):  
Stephanie Decary ◽  
Julien T. Decesse ◽  
Vasily Ogryzko ◽  
John C. Reed ◽  
Irina Naguibneva ◽  
...  
Keyword(s):  
Transcription Factor ◽  
Epithelial Cells ◽  
Gene Promoter ◽  
Promoter Sequence ◽  
Promoter Sequences ◽  
Survival Gene ◽  
Nih 3T3 ◽  
First Time ◽  
E Cadherin

ABSTRACT The retinoblastoma (RB) gene product has been shown to restrict cell proliferation, promote cell differentiation, and inhibit apoptosis. Loss of RB function can induce both p53-dependent apoptosis and p53-independent apoptosis; little is known about the mechanisms of RB-regulated p53-independent apoptosis. Here we show that RB specifically activates transcription of the survival gene bcl-2 in epithelial cells but not in NIH 3T3 mesenchymal cells. This transcriptional activity is mediated by the transcription factor AP-2. By monitoring protein-DNA interactions in living cells using formaldehyde cross-linking and chromatin immunoprecipitation, we show that endogenous RB and AP-2 both bind to the same bcl-2 promoter sequence. In addition, we demonstrate that RB and AP-2 also bind to the E-cadherin gene promoter in vivo, consistent with regulation of this promoter by both AP-2 and RB in epithelial cells. This study provides evidence that RB activates bcl-2 and E-cadherin by binding directly to the respective promoter sequences and not indirectly by repressing an inhibitor. This recruitment is mediated by a transcription factor, in this case AP-2. For the first time, our results suggest a direct molecular mechanism by which RB might inhibit apoptosis independently of p53. The results are discussed in a context where RB and Bcl-2 contribute under nonpathological conditions to the maintenance of cell viability in association with a differentiated phenotype, contributing to the tumor suppressor function of RB and playing important roles in normal development.

scholarly journals Integrin α3β1 Promotes Invasive and Metastatic Properties of Breast Cancer Cells through Induction of the Brn-2 Transcription Factor

Cancers ◽  
2021 ◽  
Vol 13 (3) ◽  
pp. 480
Author(s):  
Rakshitha Pandulal Miskin ◽  
Janine S. A. Warren ◽  
Abibatou Ndoye ◽  
Lei Wu ◽  
John M. Lamar ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Transcription Factor ◽  
Cancer Cells ◽  
Cell Invasion ◽  
Breast Cancer Cells ◽  
Gene Promoter ◽  
Akt Inhibitor ◽  
Integrin Α3β1

In the current study, we demonstrate that integrin α3β1 promotes invasive and metastatic traits of triple-negative breast cancer (TNBC) cells through induction of the transcription factor, Brain-2 (Brn-2). We show that RNAi-mediated suppression of α3β1 in MDA-MB-231 cells caused reduced expression of Brn-2 mRNA and protein and reduced activity of the BRN2 gene promoter. In addition, RNAi-targeting of Brn-2 in MDA-MB-231 cells decreased invasion in vitro and lung colonization in vivo, and exogenous Brn-2 expression partially restored invasion to cells in which α3β1 was suppressed. α3β1 promoted phosphorylation of Akt in MDA-MB-231 cells, and treatment of these cells with a pharmacological Akt inhibitor (MK-2206) reduced both Brn-2 expression and cell invasion, indicating that α3β1-Akt signaling contributes to Brn-2 induction. Analysis of RNAseq data from patients with invasive breast carcinoma revealed that high BRN2 expression correlates with poor survival. Moreover, high BRN2 expression positively correlates with high ITGA3 expression in basal-like breast cancer, which is consistent with our experimental findings that α3β1 induces Brn-2 in TNBC cells. Together, our study demonstrates a pro-invasive/pro-metastatic role for Brn-2 in breast cancer cells and identifies a role for integrin α3β1 in regulating Brn-2 expression, thereby revealing a novel mechanism of integrin-dependent breast cancer cell invasion.

Contributions of extracellular and intracellular domains of full length and chimeric cadherin molecules to junction assembly in epithelial cells

1998 ◽  
Vol 111 (9) ◽  
pp. 1305-1318 ◽  
Author(s):  
S.M. Norvell ◽  
K.J. Green
Keyword(s):  
Epithelial Cells ◽  
Cell Surface ◽  
Extracellular Domain ◽  
Full Length ◽  
Intracellular Domain ◽  
Epithelial Junctions ◽  
Intracellular Domains ◽  
E Cadherin ◽  
Cell Cell

The integrity of cell-cell junctions in epithelial cells depends on functional interactions of both extracellular and intracellular domains of cadherins with other junction proteins. To examine the roles of the different domains of E-cadherin and desmoglein in epithelial junctions, we stably expressed full length desmoglein 1 and chimeras of E-cadherin and desmoglein 1 in A431 epithelial cells. Full length desmoglein 1 was able to incorporate into or disrupt endogenous desmosomes depending on expression level. Each of the chimeric cadherin molecules exhibited distinct localization patterns at the cell surface. A chimera of the desmoglein 1 extracellular domain and the E-cadherin intracellular domain was distributed diffusely at the cell surface while the reverse chimera, comprising the E-cadherin extracellular domain and the desmoglein 1 intracellular domain, localized in large, sometimes contiguous patches at cell-cell interfaces. Nevertheless, both constructs disrupted desmosome assembly. Expression of constructs containing the desmoglein 1 cytoplasmic domain resulted in approximately a 3-fold decrease in E-cadherin bound to plakoglobin and a 5- to 10-fold reduction in the steady-state levels of the endogenous desmosomal cadherins, desmoglein 2 and desmocollin 2, possibly contributing to the dominant negative effect of the desmoglein 1 tail. In addition, biochemical analysis of protein complexes in the stable lines revealed novel in vivo protein interactions. Complexes containing beta-catenin and desmoglein 1 were identified in cells expressing constructs containing the desmoglein 1 tail. Furthermore, interactions were identified between endogenous E-cadherin and the chimera containing the E-cadherin extracellular domain and the desmoglein 1 intracellular domain providing in vivo evidence for previously predicted lateral interactions of E-cadherin extracellular domains.

Activation of NF-κB in intestinal epithelial cells by enteropathogenicEscherichia coli

1997 ◽  
Vol 273 (4) ◽  
pp. C1160-C1167 ◽  
Author(s):  
Suzana D. Savkovic ◽  
Athanasia Koutsouris ◽  
Gail Hecht
Keyword(s):  
Transcription Factor ◽  
Epithelial Cells ◽  
Intestinal Epithelial Cells ◽  
Bacterial Flora ◽  
Inflammatory Cells ◽  
Initial Response ◽  
Intestinal Epithelial ◽  
E Coli ◽  
Normal Bacterial Flora ◽  
First Time

The initial response to infection is recruitment of acute inflammatory cells to the involved site. Interleukin (IL)-8 is the prototypical effector molecule for this process. Transcription of the IL-8 gene is primarily governed by the nuclear transcription factor (NF)-κB. Intestinal epithelial cells produce IL-8 in response to infection by enteric pathogens yet remain quiescent in a milieu where they are literally bathed in normal bacterial flora. We therefore sought to investigate NF-κB activation in response to enteropathogenic Escherichia coli (EPEC), nonpathogenic E. coli, and bacterial lipopolysaccharide in an intestinal epithelial cell (T84) model and to determine whether EPEC-induced activation of NF-κB factor is causally linked to IL-8 production. We report herein that NF-κB is activated by EPEC, yet such a response is not extended to nonpathogenic organisms or purified E. coli lipopolysaccharide. Transcription factor decoys significantly diminished IL-8 production in response to EPEC, demonstrating a causal relationship. Furthermore, deletion of specific EPEC virulence genes abrogates the NF-κB-activating property of this pathogen, suggesting that specific bacterial factors are crucial for inducing this response. These studies show for the first time that infection of intestinal epithelial cells with EPEC activates NF-κB, which in turn initiates IL-8 transcription, and highlight the differential response of these cells to bacterial pathogens vs. nonpathogens.

scholarly journals Leukemia-Related Transcription Factor TEL Is Negatively Regulated through Extracellular Signal-Regulated Kinase-Induced Phosphorylation

2004 ◽  
Vol 24 (8) ◽  
pp. 3227-3237 ◽  
Author(s):  
Kazuhiro Maki ◽  
Honoka Arai ◽  
Kazuo Waga ◽  
Ko Sasaki ◽  
Fumihiko Nakamura ◽  
...  
Keyword(s):  
Transcription Factor ◽  
Dna Binding ◽  
Dominant Negative ◽  
Mitogen Activated Protein Kinase ◽  
Dominant Negative Effect ◽  
Phosphorylation Sites ◽  
3T3 Cells ◽  
Nih 3T3 ◽  
Nih 3T3 Cells

ABSTRACT TEL is an ETS family transcription factor that possesses multiple putative mitogen-activated protein kinase phosphorylation sites. We here describe the functional regulation of TEL via ERK pathways. Overexpressed TEL becomes phosphorylated in vivo by activated ERK. TEL is also directly phosphorylated in vitro by ERK. The inducible phosphorylation sites are Ser213 and Ser257. TEL binds to a common docking domain in ERK. In vivo ERK-dependent phosphorylation reduces trans-repressional and DNA-binding abilities of TEL for ETS-binding sites. A mutant carrying substituted glutamates on both Ser213 and Ser257 functionally mimics hyperphosphorylated TEL and also shows a dominant-negative effect on TEL-induced transcriptional suppression. Losing DNA-binding affinity through phosphorylation but heterodimerizing with unmodified TEL could be an underlying mechanism. Moreover, the glutamate mutant dominantly interferes with TEL-induced erythroid differentiation in MEL cells and growth suppression in NIH 3T3 cells. Finally, endogenous TEL is dephosphorylated in parallel with ERK inactivation in differentiating MEL cells and is phosphorylated through ERK activation in Ras-transformed NIH 3T3 cells. These data indicate that TEL is a constituent downstream of ERK in signal transduction systems and is physiologically regulated by ERK in molecular and biological features.

Inhibition of Hes1 activity in gall bladder epithelial cells promotes insulin expression and glucose responsiveness

2009 ◽  
Vol 87 (6) ◽  
pp. 975-987 ◽  
Author(s):  
R. A. Coad ◽  
J. R. Dutton ◽  
D. Tosh ◽  
J. M.W. Slack
Keyword(s):  
Transcription Factor ◽  
Epithelial Cells ◽  
Gall Bladder ◽  
Cell Formation ◽  
Pancreatic Tissue ◽  
Dominant Negative ◽  
Biliary System ◽  
Natural Occurrence ◽  
Β Cell

The biliary system has a close developmental relationship with the pancreas, evidenced by the natural occurrence of small numbers of biliary-derived β-cells in the biliary system and by the replacement of biliary epithelium with pancreatic tissue in mice lacking the transcription factor Hes1. In normal pancreatic development, Hes1 is known to repress endocrine cell formation. Here we show that glucose-responsive insulin secretion can be induced in biliary epithelial cells when activity of the transcription factor Hes1 is antagonised. We describe a new culture system for adult murine gall bladder epithelial cells (GBECs), free from fibroblast contamination. We show that Hes1 is expressed both in adult murine gall bladder and in cultured GBECs. We have created a new dominant negative Hes1 (ΔHes1) by removal of the DNA-binding domain, and show that it antagonises Hes1 function in vivo. When ΔHes1 is introduced into the GBEC it causes expression of insulin RNA and protein. Furthermore, it confers upon the cells the ability to secrete insulin following exposure to increased external glucose. GBEC cultures are induced to express a wider range of mature β cell markers when co-transduced with ΔHes1 and the pancreatic transcription factor Pdx1. Introduction of ΔHes1 and Pdx1 can therefore initiate a partial respecification of phenotype from biliary epithelial cell towards the pancreatic β cell.

scholarly journals Genomic Promoter Analysis Predicts Functional Transcription Factor Binding

2008 ◽  
Vol 2008 ◽  
pp. 1-9 ◽  
Author(s):  
J. Sunil Rao ◽  
Suresh Karanam ◽  
Colleen D. McCabe ◽  
Carlos S. Moreno
Keyword(s):  
Transcription Factor ◽  
Binding Sites ◽  
Transcription Factor Binding Sites ◽  
Transcription Factor Binding ◽  
Proximal Promoter ◽  
Marginal Effect ◽  
Promoter Sequences ◽  
Factor Binding ◽  
Functional Transcription Factor

Background. The computational identification of functional transcription factor binding sites (TFBSs) remains a major challenge of computational biology. Results. We have analyzed the conserved promoter sequences for the complete set of human RefSeq genes using our conserved transcription factor binding site (CONFAC) software. CONFAC identified 16296 human-mouse ortholog gene pairs, and of those pairs, 9107 genes contained conserved TFBS in the 3 kb proximal promoter and first intron. To attempt to predict in vivo occupancy of transcription factor binding sites, we developed a novel marginal effect isolator algorithm that builds upon Bayesian methods for multigroup TFBS filtering and predicted the in vivo occupancy of two transcription factors with an overall accuracy of 84%. Conclusion. Our analyses show that integration of chromatin immunoprecipitation data with conserved TFBS analysis can be used to generate accurate predictions of functional TFBS. They also show that TFBS cooccurrence can be used to predict transcription factor binding to promoters in vivo.

scholarly journals Biogenesis of Polarized Epithelial Cells During Kidney Development In Situ: Roles of E-Cadherin–mediated Cell–Cell Adhesion and Membrane Cytoskeleton Organization

1998 ◽  
Vol 9 (11) ◽  
pp. 3161-3177 ◽  
Author(s):  
Peter A. Piepenhagen ◽  
W. James Nelson
Keyword(s):  
Cell Adhesion ◽  
Epithelial Cells ◽  
Kidney Development ◽  
Lateral Membrane ◽  
Membrane Cytoskeleton ◽  
Triton X ◽  
E Cadherin ◽  
Cell Cell

Organization of proteins into structurally and functionally distinct plasma membrane domains is an essential characteristic of polarized epithelial cells. Based on studies with cultured kidney cells, we have hypothesized that a mechanism for restricting Na/K-ATPase to the basal-lateral membrane involves E-cadherin–mediated cell–cell adhesion and integration of Na/K-ATPase into the Triton X-100–insoluble ankyrin- and spectrin-based membrane cytoskeleton. In this study, we examined the relevance of these in vitro observations to the generation of epithelial cell polarity in vivo during mouse kidney development. Using differential detergent extraction, immunoblotting, and immunofluorescence histochemistry, we demonstrate the following. First, expression of the 220-kDa splice variant of ankyrin-3 correlates with the development of resistance to Triton X-100 extraction for Na/K-ATPase, E-cadherin, and catenins and precedes maximal accumulation of Na/K-ATPase. Second, expression of the 190-kDa slice variant of ankyrin-3 correlates with maximal accumulation of Na/K-ATPase. Third, Na/K-ATPase, ankyrin-3, and fodrin specifically colocalize at the basal-lateral plasma membrane of all epithelial cells in which they are expressed and during all stages of nephrogenesis. Fourth, the relative immunofluorescence staining intensities of Na/K-ATPase, ankyrin-3, and fodrin become more similar during development until they are essentially identical in adult kidney. Thus, renal epithelial cells in vivo regulate the accumulation of E-cadherin–mediated adherens junctions, the membrane cytoskeleton, and Na/K-ATPase through sequential protein expression and assembly on the basal-lateral membrane. These results are consistent with a mechanism in which generation and maintenance of polarized distributions of these proteins in vivo and in vitro involve cell–cell adhesion, assembly of the membrane cytoskeleton complex, and concomitant integration and retention of Na/K-ATPase in this complex.

scholarly journals Studies on the Role of the Transcription Factor Tcf21 in the Transdifferentiation of Parietal Epithelial Cells into Podocyte-Like Cells

2021 ◽  
Vol 55 (S4) ◽  
pp. 48-67
Keyword(s):  
Cell Cycle ◽  
Transcription Factor ◽  
Transcription Factors ◽  
Epithelial Cells ◽  
Bhlh Transcription Factor ◽  
Binding Motif ◽  
A Genome ◽  
Parietal Epithelial Cells ◽  
Transcription Factor Yy1

Background/Aims: Podocyte differentiation is essential for proper blood filtration in the kidney. It is well known that transcription factors play an essential role to maintain the differentiation of podocytes. The present study is focused on the basic helix-loop-helix (bHLH) transcription factor Tcf21 (Pod1) which is essential for the development of podocytes in vivo. Since parietal epithelial cells (PECs) are still under debate to be progenitor cells which can differentiate into podocytes, we wanted to find out whether the expression of Tcf21 induces a transition of PECs into podocytes. Methods: We transfected PECs with Tcf21-GFP and analyzed the expression of PEC- and podocyte-specific markers. Furthermore, we performed ChIP-Seq analysis to identify new putative interaction partners and target genes of Tcf21. Results: By gene arrays analysis, we found that podocytes express high levels of Tcf21 in vivo in contrast to cultured podocytes and parietal epithelial cells (PECs) in vitro. After the expression of Tcf21 in PECs, we observed a downregulation of specific PEC markers like caveolin‑1, β-catenin and Pax2. Additionally, we found that the upregulation of Tcf21 induced multi-lobulation of cell nuclei, budding and a formation of micronuclei (MBM). Furthermore, a high number of PECs showed a tetraploid set of chromosomes. By qRT-PCR and Western blot analysis, we revealed that the transcription factor YY1 is downregulated by Tcf21. Interestingly, co-expression of YY1 and Tcf21 rescues MBM and reduced tetraploidy. By ChIP-Seq analysis, we identified a genome-wide Tcf21-binding site (CAGCTG), which matched the CANNTG sequence, a common E-box binding motif used by bHLH transcription factors. Using this technique, we identified additional Tcf21 targets genes that are involved in the regulation of the cell cycle (e.g. Mdm2, Cdc45, Cyclin D1, Cyclin D2), on the stability of microtubules (e.g. Mapt) as well as chromosome segregation. Conclusion: Taken together, we demonstrate that Tcf21 inhibits the expression of PEC-specific markers and of the transcription factor YY1, induces MBM as well as regulates the cell cycle suggesting that Tcf21 might be important for PEC differentiation into podocyte-like cells.

The Role of MRPL23 Antisense RNA 1 (MRPL23-AS1) in the Pre-Metastatic Microenvironment of Malignancy During the Process of Epithelial-Mesenchymal Transition

2021 ◽  
Vol 11 (5) ◽  
pp. 864-871
Author(s):  
Chong Zhang ◽  
Huxia Gu ◽  
Dingrong Liu ◽  
Jing Fang ◽  
Yan Yang
Keyword(s):  
Endothelial Cells ◽  
Promoter Region ◽  
Epithelial Mesenchymal Transition ◽  
Vascular Endothelial Cells ◽  
Gene Promoter ◽  
Mesenchymal Transition ◽  
Metastatic Microenvironment ◽  
Promoter Dna ◽  
E Cadherin

We aimed to explore MRPL23-AS1’s role in the pre-metastatic microenvironment of malignancy during epithelial-mesenchymal transition (EMT). Identification and verification of lncRNA-interacting proteins in salivary adenoid cystic carcinoma (SACC) cells were conducted via RNA-pulldown, silver staining, and Western blotting. RIP and RIP-seq were sequentially administered to verify the binding partners of lncRNA. CHIRP was performed to detect the promoter DNA in the downstream of lncRNA-protein complex. Ultimately CHIP-qPCR detected the effects of lncRNA on the binding degree of its interacting protein to the promoter DNA in the downstream genes and the methyla-tion level of histones in the promoter region. The exosomes secreted by different SACC cells were extracted from culture supernatant to measure lncRNA expression via qPCR. MRPL23-AS1 interacted with EZH2 protein and promoted EZH2 binding to E-cadherin gene promoter region along with the H3K27 methylation. MRPL23-AS1 could promote EMT of SACC cells and increase pulmonary vascular endothelial cells permeability via exosomes secretion. MRPL23-AS1 up-regulated VEGFA, while down-regulated E-cadherin and VE-cadherin in endothelial cells. Exosomes rich in MRPL23-AS1 could boost lung metastasis in vivo. MRPL23-AS1 inhibits E-cadherin level and promotes EMT of SACC cells, suggesting that it might be a biomarker and therapeutic target for lung cancer.

scholarly journals Ksp-cadherin gene promoter. II. Kidney-specific activity in transgenic mice

1999 ◽  
Vol 277 (4) ◽  
pp. F599-F610 ◽  
Author(s):  
Peter Igarashi ◽  
Cooduvalli S. Shashikant ◽  
R. Brent Thomson ◽  
Dilys A. Whyte ◽  
Shuxian Liu-Chen ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Transgenic Mice ◽  
Collecting Duct ◽  
Gene Promoter ◽  
Regulatory Elements ◽  
Developmental Expression ◽  
Tubular Epithelial Cells ◽  
Specific Expression ◽  
Tissue Specific

Kidney-specific cadherin (Ksp-cadherin, cadherin 16) is a tissue-specific member of the cadherin superfamily that is expressed exclusively in the basolateral membrane of tubular epithelial cells in the kidney. To determine the basis for tissue-specific expression of Ksp-cadherin in vivo, we evaluated the activity of the promoter in transgenic mice. Transgenic mice containing 3.3 kb of the mouse Ksp-cadherin promoter and an Escherichia coli lacZ reporter gene were generated by pronuclear microinjection. Assays of β-galactosidase enzyme activity showed that the transgene was expressed exclusively in the kidney in both adult and developing mice. Within the kidney, the transgene was expressed in a subset of renal tubular epithelial cells that endogenously expressed Ksp-cadherin and that were identified as collecting ducts by colabeling with Dolichos biflorus agglutinin. In the developing metanephros, expression of the transgene in the branching ureteric bud correlated with the developmental expression of Ksp-cadherin. Identical patterns of expression were observed in multiple founder mice, indicating that kidney specificity was independent of transgene integration site. However, heterocellular expression was observed consistent with repeat-induced gene silencing. We conclude that the Ksp-cadherin gene promoter directs kidney-specific expression in vivo. Regulatory elements that are sufficient to recapitulate the tissue- and differentiation-specific expression of Ksp-cadherin in the renal collecting duct are located within 3.3 kb upstream to the transcriptional start site.

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