Vas deferens epithelial cells in subculture: a model to study androgen regulation of gene expression

1995 ◽  
Vol 15 (2) ◽  
pp. 129-141 ◽  
Author(s):  
A Dassouli ◽  
Ch Darne ◽  
S Fabre ◽  
M Manin ◽  
G Veyssière ◽  
...  
Keyword(s):  
Gene Expression ◽  
Androgen Receptor ◽  
Epithelial Cells ◽  
Vas Deferens ◽  
Regulatory Elements ◽  
Receptor Expression ◽  
Time Dependent ◽  
Dependent Manner ◽  
Regulated Gene Expression ◽  
Cat Gene

ABSTRACT The understanding of androgen-regulated gene expression requires a cell culture system that mimics the functions of cells in vivo. In the present paper we have examined a vas deferens epithelial cell subculture system. Cultured vas deferens epithelial cells have been shown to exhibit polarized properties characteristic of functioning epithelia and to display a high level of androgen receptors. Incubation of cells with androgen caused a decrease in cellular androgen receptor mRNA that was time-dependent. Total suppression was observed after 24 h of exposure to androgen. By contrast, incubation of vas deferens epithelial cells with androgen resulted in a threefold increase in the cellular content of androgen receptor protein, as assayed by ligand binding. In response to androgens, vas deferens epithelial cells expressed mouse vas deferens protein mRNA (MVDP mRNA). Maximum expression of the MVDP gene, at both mRNA and protein levels, was observed after 24 h of androgen induction. DEAE-dextran transfection conditions were defined using the MMTV-CAT vector. Dihydrotestosterone stimulated the transcription activation of MMTV-CAT gene in vas deferens epithelial cells in a dose- and time-dependent manner. No induction was seen when fragments of the MVDP promoter region were cloned directly in front of the CAT gene and transiently transfected into vas deferens epithelial cells. It was found that cotransfection of cells with MVDP-CAT constructs and with an androgen receptor expression vector resulted in a small but consistent androgen-dependent increase in reporter gene activity. Transiently transfected vas deferens epithelial cells are a suitable model with which to study the effect of androgen on gene regulatory elements.

scholarly journals Androgen receptor expression is regulated by the phosphoinositide 3-kinase/Akt pathway in normal and tumoral epithelial cells

2002 ◽  
Vol 366 (3) ◽  
pp. 729-736 ◽  
Author(s):  
Michèle MANIN ◽  
Silvère BARON ◽  
Karine GOOSSENS ◽  
Claude BEAUDOIN ◽  
Claude JEAN ◽  
...  
Keyword(s):  
Gene Expression ◽  
Prostate Cancer ◽  
Androgen Receptor ◽  
Epithelial Cells ◽  
Regulation Of Gene Expression ◽  
Receptor Expression ◽  
Akt Pathway ◽  
Suitable Model ◽  
Metastatic Prostate Carcinoma ◽  
Phosphoinositide 3 Kinase

The androgen receptor (AR) is a ligand-responsive transcription factor known to play a central role in the pathogenesis of prostate cancer. However, the regulation of AR gene expression in the normal and pathological prostate remains poorly understood. This study focuses on the effect of the phosphoinositide 3-kinase (PI 3-kinase)/Akt axis on AR expression in vas deferens epithelial cells (VDEC), a suitable model to study androgen regulation of gene expression, and LNCaP cells (derived from a metastasis at the left supraclavicular lymph node from a 50-year-old patient with a confirmed diagnosis of metastatic prostate carcinoma). Taken together, our data show for the first time that the PI 3-kinase/Akt pathway is required for basal and dihydrotestosterone-induced AR protein expression in both VDEC and LNCaP. Inhibition of the PI 3-kinase/Akt pathway reduced AR expression and the decline in AR protein level correlated with a decrease in AR mRNA in VDEC but not in LNCaP. Since PI 3-kinase/Akt axis is active in prostate cancer, cross-talk between PI 3-kinase/Akt and AR signalling pathways may have implications for endocrine therapy.

scholarly journals Moxifloxacin Activates the SOS Response in Mycobacterium tuberculosis in a Dose- and Time-Dependent Manner

2021 ◽  
Vol 9 (2) ◽  
pp. 255
Author(s):  
Angelo Iacobino ◽  
Giovanni Piccaro ◽  
Manuela Pardini ◽  
Lanfranco Fattorini ◽  
Federico Giannoni
Keyword(s):  
Gene Expression ◽  
Mycobacterium Tuberculosis ◽  
Physiological State ◽  
Transcriptional Response ◽  
Sos Response ◽  
Resistant Mutant ◽  
Time Dependent ◽  
Dependent Manner ◽  
Gyra Gene ◽  
Drug Concentrations

Previous studies on Escherichia coli demonstrated that sub-minimum inhibitory concentration (MIC) of fluoroquinolones induced the SOS response, increasing drug tolerance. We characterized the transcriptional response to moxifloxacin in Mycobacterium tuberculosis. Reference strain H37Rv was treated with moxifloxacin and gene expression studied by qRT-PCR. Five SOS regulon genes, recA, lexA, dnaE2, Rv3074 and Rv3776, were induced in a dose- and time-dependent manner. A range of moxifloxacin concentrations induced recA, with a peak observed at 2 × MIC (0.25 μg/mL) after 16 h. Another seven SOS responses and three DNA repair genes were significantly induced by moxifloxacin. Induction of recA by moxifloxacin was higher in log-phase than in early- and stationary-phase cells, and absent in dormant bacilli. Furthermore, in an H37Rv fluoroquinolone-resistant mutant carrying the D94G mutation in the gyrA gene, the SOS response was induced at drug concentrations higher than the mutant MIC value. The 2 × MIC of moxifloxacin determined no significant changes in gene expression in a panel of 32 genes, except for up-regulation of the relK toxin and of Rv3290c and Rv2517c, two persistence-related genes. Overall, our data show that activation of the SOS response by moxifloxacin, a likely link to increased mutation rate and persister formation, is time, dose, physiological state and, possibly, MIC dependent.

scholarly journals Cigarette Smoke Stimulates SARS-CoV-2 Internalization by Activating AhR and Increasing ACE2 Expression in Human Gingival Epithelial Cells

2021 ◽  
Vol 22 (14) ◽  
pp. 7669
Author(s):  
Cassio Luiz Coutinho Almeida-da-Silva ◽  
Harmony Matshik Dakafay ◽  
Kaitlyn Liu ◽  
David M. Ojcius
Keyword(s):  
Epithelial Cells ◽  
Oral Cavity ◽  
Cigarette Smoke ◽  
Large Body ◽  
Receptor Expression ◽  
Host Cells ◽  
Dependent Manner ◽  
Gingival Epithelial Cells ◽  
Human Gingival Epithelial Cells ◽  
Oral Cells

A large body of evidence shows the harmful effects of cigarette smoke to oral and systemic health. More recently, a link between smoking and susceptibility to coronavirus disease 2019 (COVID-19) was proposed. COVID-19 is due to infection with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), which uses the receptor ACE2 and the protease TMPRSS2 for entry into host cells, thereby infecting cells of the respiratory tract and the oral cavity. Here, we examined the effects of cigarette smoke on the expression of SARS-CoV-2 receptors and infection in human gingival epithelial cells (GECs). We found that cigarette smoke condensates (CSC) upregulated ACE2 and TMPRSS2 expression in GECs, and that CSC activated aryl hydrocarbon receptor (AhR) signaling in the oral cells. ACE2 was known to mediate SARS-CoV-2 internalization, and we demonstrate that CSC treatment potentiated the internalization of SARS-CoV-2 pseudovirus in GECs in an AhR-dependent manner. AhR depletion using small interference RNA decreased SARS-CoV-2 pseudovirus internalization in CSC-treated GECs compared with control GECs. Our study reveals that cigarette smoke upregulates SARS-CoV-2 receptor expression and infection in oral cells. Understanding the mechanisms involved in SARS-CoV-2 infection in cells of the oral cavity may suggest therapeutic interventions for preventing viral infection and transmission.

scholarly journals A gain-of-function single nucleotide variant creates a new promoter which acts as an orientation-dependent enhancer-blocker

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Yavor K. Bozhilov ◽  
Damien J. Downes ◽  
Jelena Telenius ◽  
A. Marieke Oudelaar ◽  
Emmanuel N. Olivier ◽  
...  
Keyword(s):  
Gene Expression ◽  
Genetic Diseases ◽  
Regulatory Elements ◽  
Base Change ◽  
Dependent Manner ◽  
Globin Genes ◽  
Single Nucleotide Variants ◽  
Single Nucleotide ◽  
Super Enhancer ◽  
Single Base Change

AbstractMany single nucleotide variants (SNVs) associated with human traits and genetic diseases are thought to alter the activity of existing regulatory elements. Some SNVs may also create entirely new regulatory elements which change gene expression, but the mechanism by which they do so is largely unknown. Here we show that a single base change in an otherwise unremarkable region of the human α-globin cluster creates an entirely new promoter and an associated unidirectional transcript. This SNV downregulates α-globin expression causing α-thalassaemia. Of note, the new promoter lying between the α-globin genes and their associated super-enhancer disrupts their interaction in an orientation-dependent manner. Together these observations show how both the order and orientation of the fundamental elements of the genome determine patterns of gene expression and support the concept that active genes may act to disrupt enhancer-promoter interactions in mammals as in Drosophila. Finally, these findings should prompt others to fully evaluate SNVs lying outside of known regulatory elements as causing changes in gene expression by creating new regulatory elements.

scholarly journals Blastocystis ratti Contains Cysteine Proteases That Mediate Interleukin-8 Response from Human Intestinal Epithelial Cells in an NF-κB-Dependent Manner

2007 ◽  
Vol 7 (3) ◽  
pp. 435-443 ◽  
Author(s):  
Manoj K. Puthia ◽  
Jia Lu ◽  
Kevin S. W. Tan
Keyword(s):  
Gene Expression ◽  
Epithelial Cells ◽  
Intestinal Inflammation ◽  
Experimental Studies ◽  
Cysteine Proteases ◽  
Interleukin 8 ◽  
Dependent Manner ◽  
Public Health Importance ◽  
Wide Range ◽  
First Time

ABSTRACT Blastocystis is a ubiquitous enteric protozoan found in the intestinal tracts of humans and a wide range of animals. Evidence accumulated over the last decade suggests association of Blastocystis with gastrointestinal disorders involving diarrhea, abdominal pain, constipation, nausea, and fatigue. Clinical and experimental studies have associated Blastocystis with intestinal inflammation, and it has been shown that Blastocystis has potential to modulate the host immune response. Blastocystis is also reported to be an opportunistic pathogen in immunosuppressed patients, especially those suffering from AIDS. However, nothing is known about the parasitic virulence factors and early events following host-parasite interactions. In the present study, we investigated the molecular mechanism by which Blastocystis activates interleukin-8 (IL-8) gene expression in human colonic epithelial T84 cells. We demonstrate for the first time that cysteine proteases of Blastocystis ratti WR1, a zoonotic isolate, can activate IL-8 gene expression in human colonic epithelial cells. Furthermore, we show that NF-κB activation is involved in the production of IL-8. In addition, our findings show that treatment with the antiprotozoal drug metronidazole can avert IL-8 production induced by B. ratti WR1. We also show for the first time that the central vacuole of Blastocystis may function as a reservoir for cysteine proteases. Our findings will contribute to an understanding of the pathobiology of a poorly studied parasite whose public health importance is increasingly recognized.

Tenascin-C inhibits extracellular matrix-dependent gene expression in mammary epithelial cells. Localization of active regions using recombinant tenascin fragments

1995 ◽  
Vol 108 (2) ◽  
pp. 519-527 ◽  
Author(s):  
P.L. Jones ◽  
N. Boudreau ◽  
C.A. Myers ◽  
H.P. Erickson ◽  
M.J. Bissell
Keyword(s):  
Gene Expression ◽  
Extracellular Matrix ◽  
Protein Synthesis ◽  
Epithelial Cells ◽  
Mammary Epithelial Cells ◽  
Active Regions ◽  
Mammary Epithelial ◽  
Dependent Manner ◽  
Beta Casein ◽  
Casein Protein

The physiological role of tenascin in vivo has remained obscure. Although tenascin is regulated in a stage and tissue-dependent manner, knock-out mice appear normal. When tenascin expression was examined in the normal adult mouse mammary gland, little or none was present during lactation, when epithelial cells actively synthesize and secrete milk proteins in an extracellular matrix/lactogenic hormone-dependent manner. In contrast, tenascin was prominently expressed during involution, a stage characterized by the degradation of the extracellular matrix and the subsequent loss of milk production. Studies with mammary cell lines indicated that tenascin expression was high on plastic, but was suppressed in the presence of the laminin-rich, Engelbreth-Holm-Swarm (EHS) tumour biomatrix. When exogenous tenascin was added together with EHS to mammary epithelial cells, beta-casein protein synthesis and steady-state mRNA levels were inhibited in a concentration-dependent manner. Moreover, this inhibition by tenascin could be segregated from its effects on cell morphology. Using two beta-casein promoter constructs attached to the chloramphenicol acetyltransferase reporter gene we showed that tenascin selectively suppressed extracellular matrix/prolactin-dependent transcription of the beta-casein gene in three-dimensional cultures. Finally, we mapped the active regions within the fibronectin type III repeat region of the tenascin molecule that are capable of inhibiting beta-casein protein synthesis. Our data are consistent with a model where both the loss of a laminin-rich basement membrane by extracellular matrix-degrading enzymes and the induction of tenascin contribute to the loss of tissue-specific gene expression and thus the involuting process.

Comparison of gene expression and activation of transcription factors in organoid-derived monolayer intestinal epithelial cells and organoids

2021 ◽  
Author(s):  
Yu Takahashi ◽  
Yu Inoue ◽  
Keitaro Kuze ◽  
Shintaro Sato ◽  
Makoto Shimizu ◽  
...  
Keyword(s):  
Gene Expression ◽  
Epithelial Cells ◽  
Gene Expression Regulation ◽  
Intestinal Epithelial Cells ◽  
Three Dimensional ◽  
Culture Conditions ◽  
Dependent Manner ◽  
Intestinal Epithelial

Abstract Intestinal organoids better represent in vivo intestinal properties than conventionally used established cell lines in vitro. However, they are maintained in three-dimensional culture conditions that may be accompanied by handling complexities. We characterized the properties of human organoid-derived two-dimensionally cultured intestinal epithelial cells (IECs) compared with those of their parental organoids. We found that the expression of several intestinal markers and functional genes were indistinguishable between monolayer IECs and organoids. We further confirmed that their specific ligands equally activate intestinal ligand-activated transcriptional regulators in a dose-dependent manner. The results suggest that culture conditions do not significantly influence the fundamental properties of monolayer IECs originating from organoids, at least from the perspective of gene expression regulation. This will enable their use as novel biological tools to investigate the physiological functions of the human intestine.

scholarly journals Epigenomic landscape of enhancer elements during Hydra head organizer formation

2020 ◽  
Vol 13 (1) ◽  
Author(s):  
Puli Chandramouli Reddy ◽  
Akhila Gungi ◽  
Suyog Ubhe ◽  
Sanjeev Galande
Keyword(s):  
Gene Expression ◽  
Wnt Signaling ◽  
Histone Modifications ◽  
Specific Inhibitor ◽  
Regulatory Elements ◽  
The Body ◽  
Luciferase Reporter ◽  
Dependent Manner ◽  
Chromatin Architecture ◽  
Ectopic Activation

Abstract Background Axis patterning during development is accompanied by large-scale gene expression changes. These are brought about by changes in the histone modifications leading to dynamic alterations in chromatin architecture. The cis regulatory DNA elements also play an important role towards modulating gene expression in a context-dependent manner. Hydra belongs to the phylum Cnidaria where the first asymmetry in the body plan was observed and the oral-aboral axis originated. Wnt signaling has been shown to determine the head organizer function in the basal metazoan Hydra. Results To gain insights into the evolution of cis regulatory elements and associated chromatin signatures, we ectopically activated the Wnt signaling pathway in Hydra and monitored the genome-wide alterations in key histone modifications. Motif analysis of putative intergenic enhancer elements from Hydra revealed the conservation of bilaterian cis regulatory elements that play critical roles in development. Differentially regulated enhancer elements were identified upon ectopic activation of Wnt signaling and found to regulate many head organizer specific genes. Enhancer activity of many of the identified cis regulatory elements was confirmed by luciferase reporter assay. Quantitative chromatin immunoprecipitation analysis upon activation of Wnt signaling further confirmed the enrichment of H3K27ac on the enhancer elements of Hv_Wnt5a, Hv_Wnt11 and head organizer genes Hv_Bra1, CnGsc and Hv_Pitx1. Additionally, perturbation of the putative H3K27me3 eraser activity using a specific inhibitor affected the ectopic activation of Wnt signaling indicating the importance of the dynamic changes in the H3K27 modifications towards regulation of the genes involved in the head organizer activity. Conclusions The activation-associated histone marks H3K4me3, H3K27ac and H3K9ac mark chromatin in a similar manner as seen in bilaterians. We identified intergenic cis regulatory elements which harbor sites for key transcription factors involved in developmental processes. Differentially regulated enhancers exhibited motifs for many zinc-finger, T-box and ETS related TFs whose homologs have a head specific expression in Hydra and could be a part of the pioneer TF network in the patterning of the head. The ability to differentially modify the H3K27 residue is critical for the patterning of Hydra axis revealing a dynamic acetylation/methylation switch to regulate gene expression and chromatin architecture.

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