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I Vietor

Publications and source records attributed to I Vietor.

13 recordsLinked to original sources
PubMed2004

Inhibitory effect of TIS7 on Sp1-C/EBPalpha transcription factor module activity.

The transcription factors C/EBPalpha and Sp1 functionally interact to induce expression of specific genes during myeloid and epithelial cell differentiation. The C/EBPalpha-Sp1 transcription factor "module" binds to enhancer elements within the upstream regulatory sequences of target genes. In our previous study we identified mouse TPA inducible sequence 7 (TIS7) as a novel co-repressor in epithelial cells undergoing loss of polarity. Increased levels of TIS7 down-regulate the transcription of a specific set of genes. Using bioinformatic analysis we identified a common binding site for the C/EBPalpha-Spl transcription factor module within the upstream regulatory regions of TIS7-regulated genes. The inhibitory effect of TIS7 on C/EBPalpha-Sp1-mediated transcription was confirmed by reporter assays. Our data showed that the TIS7 effect was mediated through specific interference with Sp1 transcriptional activity. Furthermore, TIS7 prevented formation of a complex between Sp1 protein and its consensus DNA binding site. Data presented here further specify the mechanism of action of the transcriptional co-repressor TIS7 as well as document the strength of a bioinformatic approach for the prediction and analysis of transcription factor modules.

Animals↗
PubMed2004

Catecholamine synthesizing enzymes and their modulation by immobilization stress in knockout mice.

The c-fos knockout mice (c-fos KO) and corticotropin-releasing hormone knockout mice (CRH KO) can serve as interesting models for studying mechanisms involved in response of the organism to stress, focused mainly on the hypothalamic-pituitary-adrenal (HPA) axis and sympathoadrenal system (SAS). The present study focused on the investigation of changes in gene expression of catecholamine biosynthesizing enzymes tyrosine hydroxylase (TH), dopamine-beta-hydroxylase (DBH), and phenylethanolamine N-methyltransferase (PNMT) in adrenal medulla of c-fos KO and CRH KO mice stressed by immobilization. Levels of TH, DBH, and PNMT mRNA were determined by reverse transcription-polymerase chain reaction (RT-PCR). Single immobilization for 2 h significantly increased adrenomedullary TH, DBH, and PNMT mRNA levels in both c-fos KO and wild-type (WT) mice compared to unstressed controls. In CRH KO mice, PNMT gene expression was not increased to the same extent after single, but especially after repeated immobilization as in WT mice, in contrast to TH and DBH mRNA levels. Thus, our data indicate that CRH deficiency can influence the PNMT mRNA level in adrenal medulla during stress, confirming the idea that the HPA axis plays the crucial role in PNMT gene regulation in mice. On the other hand, c-Fos protein probably does not play a crucial role in TH, DBH, and PNMT gene expression in adrenal medulla under stress conditions.

Adrenal Medulla↗
PubMed2001

Perturbation of the tight junction permeability barrier by occludin loop peptides activates beta-catenin/TCF/LEF-mediated transcription.

Here we show that interference with the integrity of the transepithelial permeability barrier of mouse mammary epithelial cells by treatment with synthetic peptides, homologous to the second extracellular domain of occludin, decreased the amount of occludin protein present at tight junctions and led to the formation of multilayered, unpolarized cell clusters. In addition, transcription of the adherens junction protein beta-catenin was induced. Following accumulation of soluble beta-catenin protein, transcription by beta-catenin/TCF/LEF was increased, as revealed by transcriptional assays following transient transfection of the reporter construct. Furthermore, treatment with occludin-II peptides up-regulated RNA levels of the known beta-catenin/TCF/LEF downstream target gene c-myc. The data presented imply a functional cross-talk between tight and adherens junctions that possibly contributes to the stepwise transformation during oncogenesis.

Animals↗
PubMed1999

Transient transfection of mammary epithelial cells with a PEI/DNA/adenovirus system.

Studies on epithelial cells often require the transient expression of exogenous proteins in polarized epithelial cells. However, the major limitation of this approach has been the difficulty of obtaining transient gene expression in polarized epithelial cell cultures. We report here on the application of a polyethylenimine (PEI)/DNA/adenovirus system for efficient transient gene expression in mammary epithelial cells. Based on luciferase assay and FACScan analysis the PEI/DNA/adenovirus system is shown to be an effective and simple method for transfecting epithelial cells.

Adenoviridae↗
PubMed1998

Different effects of insulin and 2-deoxy-D-glucose administration on tyrosine hydroxylase gene expression in the locus coeruleus and the adrenal medulla in rats.

The major brain norepinephrinergic nucleus, locus coeruleus, is an important integrating element of extero- and interoceptive stimuli in organisms facing different physiological challenges. We investigated the effects of single and repeated (seven times) exposure to immobilization stress (120 min daily), insulin (5 IU/kg, i.p. daily) or 2-deoxy-D-glucose (500 mg/kg, i.p. daily) administration on tyrosine hydroxylase (TH) mRNA levels, the rate-limiting enzyme in catecholamine biosynthesis, by in situ hybridization in locus coeruleus and by Northern blot analysis in the adrenal medulla of rats. Both the single and repeated immobilization caused a significant increase in TH mRNA levels in the locus coeruleus (1.5-2-fold; p < 0.05) and in the adrenal medulla (about 4-fold; p < 0.05) when compared with unstressed controls. Hypoglycemia induced by a single or repeated insulin administration led to about fourfold (p < 0.01) elevation in adrenal medullary TH mRNA levels, whereas TH mRNA in locus coeruleus remained unchanged when compared with saline-treated controls. In contrast to the effect of insulin-induced hypoglycemia, cellular glucoprivation caused by a single or repeated 2-deoxy-D-glucose administration significantly elevated TH mRNA levels in both the adrenal medulla (fourfold; p < 0.01) and the locus coeruleus (twofold; p < 0.01). Our data suggest that in contrast to immobilization or cellular glucoprivation caused by 2-deoxy-D-glucose administration, insulin-induced hypoglycemia is not a specific or quantitatively sufficient stimulus for induction of TH gene expression in the locus coeruleus, although all these stressors strongly activate the process in the adrenal medulla.

Adrenal Medulla↗
PubMed1997

In search of differentially expressed genes and proteins.

A great challenge for modern cell biology is the successful examination of the co-expression of thousands of genes under physiological or pathological conditions and how the expression patterns define the different states of a single cell, tissue or a microorganism. Gene expression can be analyzed today on a large scale by advanced technical approaches for differential screening of proteins and mRNAs. The identification of differentially expressed mRNAs has been successfully applied to understand gene function and the underlying molecular mechanism(-s) of differentiation, development and disease state. Analysis of gene expression by the systematic mapping of thousands of proteins present in a cell or tissue can be achieved by the use of two-dimensional (2D) gel electrophoresis, quantitative computer image analysis, and protein identification techniques. In this article, we comment on some of these techniques and try to stress their advantages and drawbacks. We show how data from RNA/DNA mapping, sequence information from genome projects and protein pattern profiling can be linked with each other and annotated. These comprehensive approaches permit the study of differential gene and protein expressions in cells or tissues.

Animals↗
PubMed1997

Sodium salicylate induces apoptosis via p38 mitogen-activated protein kinase but inhibits tumor necrosis factor-induced c-Jun N-terminal kinase/stress-activated protein kinase activation.

In a previous study, we demonstrated that sodium salicylate (NaSal) selectively inhibits tumor necrosis factor (TNF)-induced activation of the p42 and p44 mitogen-activated protein kinases (MAPKs) (known as extracellular signal-regulated kinases). Here we show that in normal human FS-4 fibroblasts NaSal inhibits TNF-induced activation of another member of the MAPK family, the c-Jun N-terminal kinase/stress-activated protein kinase. c-Jun N-terminal kinase activation induced by interleukin 1 or epidermal growth factor was less strongly inhibited by NaSal. Unexpectedly, treatment of FS-4 cells with NaSal alone produced a strong activation of p38 MAPK and cell death by apoptosis. NaSal-induced apoptosis was blocked by the selective p38 MAPK inhibitor SB-203580, indicating that p38 MAPK serves as a mediator of NaSal-induced apoptosis in human fibroblasts. Activation of p38 MAPK and the resulting induction of apoptosis may be important in the demonstrated antineoplastic actions of nonsteroidal anti-inflammatory drugs.

Apoptosis↗
PubMed1996

Glucoprivation by insulin leads to trans-synaptic increase in rat adrenal tyrosine hydroxylase mRNA levels.

Effects of single or repeated insulin or 2-deoxy-D-glucose administration on adrenal tyrosine hydroxylase mRNA and protein levels were examined in rats. Insulin produced hypoglycemia and an elevation in plasma epinephrine and norepinephrine levels. A significant increase (3-5-fold) in tyrosine hydroxylase mRNA levels was found at 5 h, decreasing to near basal levels at 24 h following the first and also the sixth consecutive injection of insulin or 2-deoxy-D-glucose. Whereas insulin treatment raised tyrosine hydroxylase mRNA levels in intact adrenals, no increase in tyrosine hydroxylase mRNA levels occurred following adrenal denervation by splanchnic nerve transection. Western blot analysis showed that although a single insulin treatment did not affect tyrosine hydroxylase protein levels, a significant increase was observed following the seventh administration. This study shows that insulin-induced hypoglycemia increases tyrosine hydroxylase gene expression and that this process is regulated by a central mechanism via the splanchnic nerve.

Adrenal Medulla↗
PubMed1996

CCAAT box enhancer binding protein alpha (C/EBP-alpha) stimulates kappaB element-mediated transcription in transfected cells.

A construct comprising three tandemly repeated copies of the kappaB element from the interleukin-8 gene linked to chloramphenicol acetyltransferase (CAT) (3xNF-kappaBCAT) was transcriptionally activated in normal human FS-4 fibroblasts by co-transfection with expression vectors for NF-kappaB p50, p65, or p52. Unexpectedly, a significant activation of 3xNF-kappaBCAT was also seen upon its co-transfection with the expression vector for CCAAT box enhancer binding protein alpha (C/EBP-alpha) (but not C/EBP-beta or C/EBP-delta). Stimulation by C/EBP-alpha required some other factor(s) present in FS-4 cells because no transcriptional activation of 3xNF-kappaBCAT was seen after co-transfection with C/EBP-alpha in F9 mouse embryonic carcinoma cells, known to be deficient in several transcription factors. To determine whether transcriptional activation was the result of interaction with one of the major NF-kappaB proteins, we co-transfected C/EBP-alpha with NF-kappaB p50, p65, p50 + p65, or p52 into F9 or FS-4 cells. No cooperative interaction was seen; in fact, C/EBP- alpha reduced p65-stimulated transcription, especially in F9 cells. Electrophoretic mobility shift assay with a kappaB probe revealed that the addition of recombinant C/EBP-alpha protein to nuclear extracts from untreated FS-4 cells resulted in the appearance of four bands. Only one of these bands was supershifted by antibody to p50, whereas antibodies to p65 or other NF-kappaB proteins had no effect. Our findings show that C/EBP-alpha may cause activation of some kappaB element-containing genes lacking C/EBP binding sites.

Animals↗
PubMed1996

Induction of adrenal tyrosine hydroxylase mRNA by single immobilization stress occurs even after splanchnic transection and in the presence of cholinergic antagonists.

Immobilization (IMO) stress elevates plasma catecholamines and increases tyrosine hydroxylase (TH) gene expression in rat adrenals. This study examined the mechanism(s) of IMO-induced changes in adrenal TH mRNA levels. Innervation of the adrenal medulla is predominantly cholinergic and splanchnicotomy as well as nicotinic receptor antagonists prevent the cold-induced rise in TH mRNA levels. In this study, the IMO-induced rise in plasma catecholamines, but not TH mRNA levels, was reduced by the antagonist chlorisondamine. Muscarinic antagonist atropine also did not prevent the IMO stress-elicited rise in TH mRNA. Furthermore, denervation of the adrenals by unilateral splanchnicotomy did not block the IMO-induced rise in TH mRNA but completely prevented the induction of neuropeptide Y mRNA. These results suggest that (1) the large increase in adrenal TH gene expression elicited by a single IMO stress is not regulated via cholinergic receptors or splanchnic innervation, and (2) there is a dissociation between regulatory mechanisms of catecholamine secretion and elevation of TH gene expression in the adrenal medulla of rats during IMO stress.

Adrenal Medulla↗
PubMed1994

Pathways of heat shock protein 28 phosphorylation by TNF in human fibroblasts.

Treatment of human diploid FS-4 fibroblasts with TNF or IL-1 led to a rapid increase in the phosphorylation of a approximately 28-kDa protein. Increased phosphorylation was seen after 5 min of TNF treatment, it reached a plateau between 10 and 30 min, and decreased thereafter. Immunoprecipitation with specific antibodies identified the 28-kDa protein as a member of the family of small heat shock proteins (Hsp28). Treatment of cells with different kinase inhibitors (staurosporine, H7, H8, HA-1004, or chelerythrine chloride) failed to inhibit TNF-induced Hsp28 phosphorylation, suggesting that neither protein kinase C nor other common protein kinases were involved. Treatment of FS-4 cells with sodium arsenite led to a very strong increase in the phosphorylation of Hsp28 demonstrable after 5 min and persisting for at least 4 h. Tyrosine phosphorylation of pp42 and pp44 MAP kinases was increased by TNF treatment, whereas arsenite produced a modest increase in tyrosine phosphorylation of pp44 while decreasing that of pp42 MAP kinase. The finding that sodium arsenite strongly increased Hsp28 phosphorylation, together with the resistance of TNF-induced phosphorylation to kinase inhibitors, supports the notion that increased serine phosphorylation of Hsp28 in this system involves inhibition of protein phosphatase activity.

1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine↗
PubMed1993

Tumor necrosis factor-induced activation and increased tyrosine phosphorylation of mitogen-activated protein (MAP) kinase in human fibroblasts.

Tumor necrosis factor (TNF) is a pleiotropic cytokine whose many demonstrated actions include effects on cell growth and differentiation. TNF treatment of cells is known to lead to a rapid increase in serine/threonine phosphorylation of many cellular proteins, but the kinases responsible remain largely unidentified. We show that TNF treatment induces a rapid and transient increase in mitogen-activated protein kinase (MAPK) activity in the human diploid FS-4 cell line, for which TNF is known to be mitogenic. TNF-induced activation of MAPK was demonstrated by its enhanced ability to phosphorylate myelin basic protein in vitro and by a characteristic shift in the electrophoretic mobility of MAPK proteins. MAPK activation was accompanied by a significant increase of MAPK phosphorylation on tyrosine residues, which was demonstrated by 32P labeling of cells and isolation of the labeled proteins after immunoprecipitation with antibodies to phosphotyrosine, and by direct immunoblotting of SDS-polyacrylamide gel electrophoresis-fractionated unlabeled cell lysates with antibodies to phosphotyrosine. The pp42 and pp44 MAPK were the only proteins whose tyrosine phosphorylation was demonstrably increased in FS-4 cells after TNF treatment. MAPK activation is likely to represent an important component in the cascade of signals that link TNF receptors to various TNF-elicited cellular responses.

Blotting, Western↗