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F E Johansen

Publications and source records attributed to F E Johansen.

28 records · Page 2Linked to original sources
PubMed1998

A composite DNA element in the promoter of the polymeric immunoglobulin receptor regulates its constitutive expression.

The polymeric immunoglobulin receptor (pIgR), which is constitutively expressed on the basolateral surface of secretory epithelial cells, mediates external translocation of polymeric IgA and pentameric IgM (collectively called pIg) to exocrine secretions. A high level of synthesis must be maintained because the receptor is continuously cleaved to release bound secretory component (SC) in secretory IgA and secretory IgM, as well as free SC from unoccupied receptor. We have isolated the promoter of the pIgR gene and identified a short activating region that is required for the expression of pIgR promoter-driven reporter genes. This region contained an E-box and an inverted repeat sequence (IRS). Gel electrophoresis mobility shift assays with nuclear extracts from different pIgR-expressing epithelial cell lines demonstrated proteins that bind independently to both the E-box and the IRS sequence of the pIgR promoter. In addition, a DNA probe that contained both the E-box and the IRS gave rise to a larger complex that could not be competed by either element on its own. Binding was confirmed by DNase I footprinting of the E-box and IRS sequences with nuclear extracts, and by dimethyl sulfide footprinting in living HT-29 epithelial cells. Finally, a mutation in the pIgR promoter that inhibited protein binding to the E-box and the formation of the larger complex, abolished activated transcription from the reporter gene.

Base Sequence↗
PubMed1998

Gluten induces an intestinal cytokine response strongly dominated by interferon gamma in patients with celiac disease.

BACKGROUND & AIMS: Celiac disease appears to be a T cell-mediated enteropathy induced by gluten in genetically predisposed individuals. Duodenal biopsy specimens from patients with celiac disease and histologically normal controls were investigated to see if cytokine expression is related to disease activity. METHODS: Cytokine messenger RNA (mRNA) expression was determined by quantitative reverse-transcription polymerase chain reaction and in situ expression by immunohistochemistry. RESULTS: In normal controls, mRNA levels were usually below the quantitative limit, even after in vitro gluten stimulation. By contrast, interferon (IFN)-gamma mRNA was increased more than 1000-fold in untreated disease. In vitro gluten stimulation of specimens from treated patients (gluten-free diet) increased IFN-gamma mRNA to the levels of untreated patients. In addition, increased mRNA levels for interleukin (IL)-2, IL-4, IL-6, and tumor necrosis factor alpha were found after such stimulation, whereas mRNA for IL-5, IL-10, and IL-12p40 was usually below the quantitative level. Biopsy specimens from untreated patients contained on average 10-fold more lamina propria cells positive for IFN-gamma than normal controls, whereas cells containing IL-4 were rare in both subject groups. CONCLUSIONS: The results show that mucosal gluten exposure in patients with celiac disease rapidly elicits high levels of IFN-gamma expression and lower levels of IL-2, IL-4, IL-6, and tumor necrosis factor alpha even in the virtual absence of IL-12.

Adolescent↗
PubMed1998

Cytokine profiles of cultured microvascular endothelial cells from the human intestine.

BACKGROUND AND AIMS: Cytokine production by endothelial cells, has, for practical reasons, been chiefly studied in human umbilical vein endothelial cells (HUVEC) but, because tissue-specific differences apparently exist, the role of human intestinal microvascular endothelial cells (HIMEC) as a source of mucosal cytokines was also assessed. METHODS: The expression of cytokine transcripts in HIMEC was screened by means of reverse transcription polymerase chain reaction (RT-PCR) and compared with cytokine profiles of HUVEC. Production of cytokines was investigated by bioassay and enzyme linked immunosorbent assay (ELISA). RESULTS: In the basal unstimulated state, HIMEC and HUVEC cultures contained detectable mRNA for interleukin (IL)-3, IL-7, IL-8, IL-11, IL-14, IL-15, tumour necrosis factor (TNF)-alpha, transforming growth factor (TGF)-beta, and granulocytemacrophage colony stimulating factor (GM-CSF). However, message was undetectable for IL-2, IL-4, IL-5, IL-9, IL-10, IL-12p40, IL-13, and interferon (IFN)-gamma in the resting as well as the stimulated state. Stimulation of HIMEC and HUVEC with recombinant human (rh) IL-1 beta or rhTNF-alpha induced cell associated bioactive IL-1 alpha but not IL-1 beta, as well as enhanced secretion of both IL-6 and IL-8. Furthermore, transcript levels for GM-CSF and TNF-alpha were enhanced by rhIL-1 beta or rhTNF-alpha in both cell types. Supernatants from Th1-like or Th0-like gluten reactive intestinal T cell clones derived from patients with coeliac disease elicited cytokine profiles in both HIMEC and HUVEC similar to those revealed after rhIL-1 beta or rhTNF-alpha stimulation. CONCLUSIONS: These data demonstrate that the intestinal microvascular endothelium may contribute to the cytokine network of the intestinal mucosa with the ability to respond to locally generated cytokines and to produce potent inflammatory mediators.

Celiac Disease↗
PubMed1997

Mechanism for enhanced external transfer of dimeric IgA over pentameric IgM: studies of diffusion, binding to the human polymeric Ig receptor, and epithelial transcytosis.

Transport of polymeric Igs (pIgA and pIgM) across secretory epithelia is mediated by the polymeric Ig receptor (pIgR), also known as the transmembrane secretory component. Compared with local production, external transfer of pIgA is favored 6- to 12-fold over that of pIgM on a molar basis. This transfer may be modulated at several levels: diffusion through matrix and basement membranes, ligand affinity for pIgR, and efficiency of epithelial transcytosis. To investigate these possibilities, we compared the ability of Madin-Darby canine kidney epithelial cells transfected with human pIgR to transport pIgA vs pIgM from the basolateral to the apical face, and examined the inhibitory effect of various filter types used for mounting of the monolayer. Binding data showed that pIgR bound pIgA and pIgM with similar affinity. Internalization of both ligands was fast and took place at similar rates; transcytosis was also found to be equally efficient at the molar level. Thus, the overall rate of transport across the epithelial monolayer was comparable for pIgA and pIgM, and was not further enhanced by ligand stimulation over a 20-fold increased concentration level. Conversely, pIgA had a considerable advantage over pIgM in passive diffusion assays performed in vitro. Moreover, in situ immunofluorescence staining showed retention of IgM over IgA and IgG in mucosal basement membrane zones, in contrast to the preferential epithelial uptake of IgA and, less so, IgM. The biologic consequences of the highly efficient epithelial pIg transport, and the diffusion advantage of pIgA over pIgM, are discussed in relation to the evolution and function of secretory Abs.

Animals↗
PubMed1997

Interaction of ATF6 and serum response factor.

Serum response factor (SRF) is a transcription factor which binds to the serum response element (SRE) in the c-fos promoter. It is required for regulated expression of the c-fos gene as well as other immediate-early genes and some tissue-specific genes. To better understand the regulation of SRF, we used a yeast interaction assay to screen a human HeLa cell cDNA library for SRF-interacting proteins. ATF6, a basic-leucine zipper protein, was isolated by binding to SRF and in particular to its transcriptional activation domain. The binding of ATF6 to SRF was also detected in vitro. An ATF6-VP16 chimera activated expression of an SRE reporter gene in HeLa cells, suggesting that ATF6 can interact with endogenous SRF. More strikingly, an antisense ATF6 construct reduced serum induction of a c-fos reporter gene, suggesting that ATF6 is involved in activation of transcription by SRF. ATF6 was previously partially cloned as a member of the ATF family. The complete cDNA of ATF6 was isolated, and its expression pattern was described.

Activating Transcription Factor 6↗
PubMed1997

Mucosal immunity--a major adaptive defence mechanism.

The epithelial glycoprotein called secretory component (SC) is quantitatively the most important receptor of the immune system because it is responsible for external transport of locally produced polymeric IgA (pIgA) to generate remarkably large amounts of secretory IgA. Antibodies of this type constitute the major mediators of specific humoral immunity. Transmembrane SC belongs to the Ig supergene family and functions as a common pIg receptor, also translocating pentameric IgM externally to form secretory IgM. The B cells responsible for mucosal pIg production are initially stimulated in organized mucosa-associated lymphoepithelial structures, particularly the Peyer's patches in the distal small intestine; from these inductive site they migrate as memory cells to exocrine tissues all over the body. Mucous membranes are thus furnished with secretory antibodies in an integrated way, ensuring a variety of specificities at every secretory effector site. There is currently great interest in exploiting this integrated or "common" mucosal immune system for oral vaccination against pathogenic infectious agents and also to induce tolerance in T cell-mediated autoimmune diseases. However, much remains to be learned about mechanisms for antigen uptake and processing necessary to elicit stimulatory or suppressive mucosal immune responses. Moreover, evidence is emerging for the existence of considerable regionalization with regard to functional links between inductive sites and effecter sites of mucosal immunity.

Animals↗
PubMed1994

Suppression of Ras transformation by serum response factor.

Serum response factor (SRF) is a nuclear transcription factor that binds to the serum response element (SRE) found in the promoter regions of a number of growth factor-inducible genes, as well as muscle-specific genes. The smooth muscle alpha-actin promoter contains two SRE sequences that can bind to SRF. Its expression is repressed in Ras-transformed fibroblast cells and derepressed in revertant cells. In this study, we demonstrate that SRF can activate alpha-actin expression in Ras-transformed cells and that overexpression of SRF in Ras-transformed cells can revert their transformed phenotype. The ability of SRF to bind to the SRE was required for this effect, since mutations that inhibit DNA binding abolish SRF's ability to activate alpha-actin expression and suppress transformation by the ras oncogene. These results show that SRF, thought to be involved in stimulation of cell growth through activation of growth factor-inducible genes, can actually have the opposite effect and suggest a novel mechanism for suppression of transformation by Ras.

Actins↗
PubMed1994

Two pathways for serum regulation of the c-fos serum response element require specific sequence elements and a minimal domain of serum response factor.

The c-fos serum response element (SRE) is necessary and sufficient for induction of the c-fos gene in response to serum and growth factors. This activation is dependent upon serum response factor (SRF), a transcriptional activator which binds the SRE. A factor, p62TCF, which binds in conjunction with SRF to the SRE and which is activated by mitogen-activated protein kinase, has also been implicated in c-fos regulation. By using a reporter gene system with weak SRE mutations that is dependent upon overexpression of SRF for serum induction, we have found that there are at least two pathways for serum induction that converge on the SRE. Loss of TCF binding by mutations in SRF and the SRE did not reduce serum induction of the reporter genes. We have found a pathway for serum induction that is sensitive to mutations in the A/T-containing central sequence of the SRE and which is independent of TCF. When this pathway was mutated, activation was dependent upon TCF binding, demonstrating that TCF can also function in serum induction. Both of the signalling pathways required a minimal domain of SRF. This domain, spanning SRF's DNA binding domain, was sufficient for serum induction when fused to a heterologous transcriptional activation domain.

Amino Acid Sequence↗
PubMed1993

Identification of transcriptional activation and inhibitory domains in serum response factor (SRF) by using GAL4-SRF constructs.

The binding of serum response factor (SRF) to the c-fos serum response element has been shown to be essential for serum and growth factor activation of c-Fos. Since SRF is ubiquitously expressed, it has been difficult to measure the activity of SRF introduced into cells. To assay for functions of SRF in cells, we have changed its DNA binding specificity by fusing it to the DNA binding domain of GAL4. Transfection of GAL4-SRF constructs into cells has allowed us to identify SRF's transcriptional activation domain as well as domains which inhibit this activity. First, we found that the transcriptional activation domain maps to between amino acids 339 and 508 in HeLa cells and to between amino acids 414 and 508 in NIH 3T3 cells. Second, we show that in the context of GAL4-SRF constructs, there are two separate domains of SRF that can inhibit its activation domain. Although these domains overlap the DNA binding and dimerization domains of SRF, these functions were not required for inhibition. Finally, we show that one of the inhibitory domains is modular in that it can also inhibit activation when it is moved amino terminal to GAL4's DNA binding domain in an SRF-GAL4-SRF construct. The implications of these inhibitory domains for SRF regulation are discussed.

Base Sequence↗