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S Leppä

Publications and source records attributed to S Leppä.

29 records · Page 2Linked to original sources
PubMed1997

Overexpression of HSF2-beta inhibits hemin-induced heat shock gene expression and erythroid differentiation in K562 cells.

Acquisition of heat shock factor 2 (HSF2) DNA binding activity is accompanied by induced transcription of heat shock genes in hemin-treated K562 cells undergoing erythroid differentiation. Previous studies revealed that HSF2 consists of two alternatively spliced isoforms, HSF2-alpha and HSF2-beta, whose relative abundance is developmentally regulated and varies between different tissues. To investigate whether the molar ratio of HSF2-alpha and HSF2-beta isoforms is crucial for the activation of HSF2 and whether the HSF2 isoforms play functionally distinct roles during the hemin-mediated erythroid differentiation, we generated cell clones expressing different levels of HSF2-alpha and HSF2-beta. We show that in parental K562 cells, the HSF2-alpha isoform is predominantly expressed and HSF2 can be activated upon hemin treatment. In contrast, when HSF2-beta is expressed at levels exceeding those of endogenous HSF2-alpha, the hemin-induced DNA binding activity and transcription of heat shock genes are repressed, whereas overexpression of HSF2-alpha results in an enhanced hemin response. Furthermore, the hemin-induced accumulation of globin, known as a marker of erythroid differentiation, is decreased in cells overexpressing HSF2-beta. We suggest that HSF2-beta acts as a negative regulator of HSF2 activity during hemin-mediated erythroid differentiation of K562 cells.

Alternative Splicing↗
PubMed1997

The phorbol ester 12-O-tetradecanoylphorbol 13-acetate enhances the heat-induced stress response.

Induction of heat shock gene expression is mediated by specific heat shock transcription factors (HSFs), but the signaling pathways leading to activation of HSFs are poorly understood. To elucidate whether protein kinase C-responsive signaling pathways could be involved in the regulation of heat shock gene expression, we have examined the effects of the protein kinase C activator 12-O-tetradecanoylphorbol 13-acetate (TPA) on the heat-induced stress response in K562 cells. We demonstrate that TPA treatment markedly enhances heat shock gene expression during heat stress, although TPA alone does not induce the heat shock response. This TPA-mediated enhancement can initially be detected as an accelerated acquisition of DNA binding and transcriptional activity of HSF1 resulting in elevated Hsp70 protein concentrations. In the presence of TPA, the attenuation of HSF1 DNA binding activity during continuous exposure to heat shock occurs more rapidly and in concert with the appearance of newly synthesized Hsp70, which supports earlier studies on the autoregulatory role of Hsp70 in deactivation of HSF1. During heat stress, a correlation between the hyperphosphorylation of HSF1 and its transcriptional activity was observed, in both the presence and the absence of TPA. Our results show that the heat-induced stress response can be significantly modulated by activation of protein kinase C-responsive signaling pathways.

DNA↗
PubMed1997

Heat shock response--pathophysiological implications.

All organisms exposed to environmental stress conditions share a common molecular response characterized by a dramatic change in the pattern of gene expression followed by an elevated synthesis of heat shock or stress proteins. These proteins function as molecular chaperones to protect cells from environmental stress damage by binding to partially denatured proteins, dissociating protein aggregates, and regulating the correct folding and intracellular translocation of newly synthesized polypeptides. Accumulating evidence supports a role for heat shock proteins in a number of disease states of which inflammatory reactions and ischaema provide the best studied examples. The inducible heat shock response involves transcriptional gene activation mediated by specific regulatory proteins called heat shock transcription factors, which bind to the promoter of heat shock genes in a sequence-specific manner. However, the signalling pathways leading to the activation of these transcription factors need to be characterized in more detail to be able to understand the role, cause, or consequence, of heat shock proteins in human diseases. This review presents recent progress in unravelling the regulation of heat shock gene expression in cells subjected to heat or other forms of stress. By using inflammatory responses and myocardial ischaema as examples, the putative use of heat shock proteins are discussed as targets for future therapeutic applications.

Animals↗
PubMed1997

Collagenase 3 (matrix metalloproteinase 13) gene expression by HaCaT keratinocytes is enhanced by tumor necrosis factor alpha and transforming growth factor beta.

Collagenase-3 (matrix metalloproteinase 13; MMP-13) is a novel matrix metalloproteinase, the expression of which to date has only been detected in human breast carcinoma tissue and osteoarthritic cartilage. Here, we show that MMP-13 transcripts are expressed by human HaCaT keratinocytes but not by primary human epidermal keratinocytes. The levels of MMP-13 mRNAs in HaCaT cells were enhanced up to 130- and 45-fold by tumor necrosis factor alpha (TNF-alpha) and transforming growth factor beta (TGF-beta), respectively. The maximal induction of MMP-13 mRNAs by TNF-alpha was noted after a 6-h incubation, whereas with TGF-beta, the maximal stimulation was observed after 24 h. The up-regulation of MMP-13 mRNA abundance by TNF-alpha and TGF-beta was dependent on protein synthesis and was prevented partially by dexamethasone and retinoic acid. Nuclear run-on assays demonstrated activation of MMP-13 gene transcription by TNF-alpha maximally at the 2-h time point and by TGF-beta after 12 h of treatment. Incubation of HaCaT keratinocytes with TNF-alpha and TGF-beta also increased production of proMMP-13 into the culture media, as detected by Western blotting. Our data indicate that the MMP-13 gene is expressed by transformed epidermal keratinocytes, suggesting a role for MMP-13 in the invasive capacity of human epidermal malignancies.

Blotting, Northern↗
PubMed1996

Syndecan-1 expression in mammary epithelial tumor cells is E-cadherin-dependent.

E-cadherin is a Ca(2+)-dependent cell-cell adhesion molecule, which is mainly expressed in epithelial cells. Recent studies have shown that E-cadherin has an important role as an invasion suppressor molecule in epithelial tumor cells. Syndecan-1 is a cell surface proteoglycan that has been implicated in a number of cellular functions including cell-cell adhesion, cell-matrix anchorage and growth factor presentation for signalling receptors. Its suppression has also been shown to be associated with malignant transformation of epithelial cells. In order to better understand the coordinated regulation of cell-cell and cell-matrix interactions during malignant transformation, we have studied the expression of syndecan-1 in malignant mammary tumor cells genetically manipulated for E-cadherin expression. In invasive NM-e-ras-MAC1 cells, where E-cadherin was partially downregulated by specific antisense RNA, syndecan-1 expression was suppressed. Furthermore, transfection of E-cadherin cDNA into invasive NM-f-ras-TD cells resulted in the upregulation of syndecan-1 expression in association with decreased invasiveness. In both cases, regulation of syndecan-1 occurred post-transcriptionally, since syndecan-1 mRNA levels remained unchanged. Instead, a translational regulation is suggested, since syndecan-1 core protein synthesis was E-cadherin dependent. Another cell adhesion protein, beta 1-integrin was not affected by E-cadherin expression. The data provide an example of coordinated changes in the expression of two cell adhesion molecules, syndecan-1 and E-cadherin during epithelial cell transformation.

Animals↗
PubMed1995

Increased glycosylation of beta 1 integrins affects the interaction of transformed S115 mammary epithelial cells with laminin-1.

The effect of transformation on the expression and the functions of beta 1 integrins was studied using an in vitro cell transformation model. S115 mammary epithelial tumor cells undergo transformation into tumorigenic fibroblastoid cells in the presence of steroids. Transformation was found to reduce the attachment and the spreading of S115 cells on laminin-1 but not fibronectin. Adhesion of S115 cells to laminin-1 was inhibited in the presence of an antibody against the beta 1 integrin subunit. Both nontreated and transformed S115 cells expressed at least two putative laminin-1-binding beta 1 integrins at the same level. In transformed cells, however, the mature integrin subunits appeared to be structurally altered, showing a slower electrophoretic mobility. Treatment with N-glycosidase-F and tunicamycin abolished this mobility difference, suggesting that the presence of complex-type N-linked oligosaccharides was responsible. Detailed enzymatic analysis of the oligosaccharides present on the beta 1 subunits revealed that the difference in glycosylation is, at least partially, due to poly-N-lactosaminoglycan chains on beta 1 integrin from transformed cells. Removal of this difference in glycosylation by either cleavage of the polylactosaminoglycan chains with endo-beta-galactosidase or inhibiton of complex-type glycan formation with swainsonine repeatedly enhanced the spreading of transformed cells on laminin-1. Thus, the increased size of complex-type oligosaccharides on beta 1 integrin may affect cell-laminin-1 interactions. Similar changes may contribute to the altered adhesion of cancer cells during the invasion and metastasis.

Animals↗
PubMed1993

Translational suppression of syndecan-1 expression in Ha-ras transformed mouse mammary epithelial cells.

A cell surface proteoglycan, syndecan-1, has been shown to participate in the maintenance of the epithelial cell morphology. A point mutated activated c-Ha-ras gene under the control of the glucocorticoid inducible MMTV-LTR promoter was transfected into the mouse mammary epithelial cell line, NOG-8. The NOG-8 ras cells were used to study changes in syndecan-1 expression during epithelial transformation. NOG-8 ras cells, when induced to express Ha-ras, transformed and formed foci in monolayer cultures and colonies in suspension cultures. Expression of syndecan-1 at the cell surface was markedly reduced in cells showing the transformed phenotype. The accumulation of newly synthesized core protein of syndecan-1 was suppressed in these cells, whereas mRNA levels remained unchanged. This novel finding indicates that syndecan-1 expression is translationally suppressed in the Ha-ras-transformed epithelial cells. Hence, syndecan-1 loss during epithelial transformation could take place without altering syndecan gene transcription and, on the other hand, could be one of the critical events involved in malignant transformation.

Animals↗
PubMed1992

Syndecan expression regulates cell morphology and growth of mouse mammary epithelial tumor cells.

S115 mouse mammary epithelial cells lose their epithelial morphology and become tumorigenic when exposed to steroids. We have recently reported that testosterone exposure results in the suppression of syndecan expression, suggesting that this cell surface proteoglycan may influence S115 cell phenotype. We now report that a similar suppression and morphological response of S115 cells can be achieved by glucocorticoid exposure. We introduced into S115 cells an exogenous gene construct containing the full-length human syndecan cDNA under the control of a glucocorticoid-inducible retroviral promoter, in order to study the effect of syndecan expression on S115 cell behavior. Glucocorticoid-induced re-expression of syndecan in S115 cells restored an epithelial phenotype, while control transfectants and parental S115 cells exhibited an altered, nonepithelial phenotype. Moreover, the S115 cells expressing exogenous syndecan revealed a reduced ability to form colonies in soft agar. Therefore, the maintenance of epithelial morphology and normal growth of S115 cells are dependent on syndecan expression.

Actin Cytoskeleton↗
PubMed1991

Steroid-induced epithelial-fibroblastic conversion associated with syndecan suppression in S115 mouse mammary tumor cells.

Cell-matrix interactions play an important role in the maintenance of cell shape, supposed to be mediated by the anchorage of cellular cytoskeleton to extracellular matrix via matrix receptors. In this work the expression of one of the known matrix receptors, syndecan, was studied during the hormone-induced change in the phenotype of Shionogi 115 (S115) mouse mammary tumor cells. In the presence of testosterone, when S115 cells express fibroblastic phenotype, they increased their growth rate and became gradually anchorage independent. These cells, however, revealed strong RGDS-dependent binding to fibronectin (FN) but not binding to the heparin-binding domain of FN. Instead, S115 cells growth without testosterone showed epithelial morphology and binding to the heparin-binding domain of FN, suggesting an alteration of syndecan expression in hormone-treated S115 cells. As quantitated by radioimmunoassay and by Western blot, the amounts of both matrix-binding ectodomain of syndecan and syndecan mRNA (2.6 kb) declined in hormone-treated S115 cells. The addition of antiandrogen cyproterone acetate to culture medium opposed the effect of testosterone on syndecan mRNA. We thus propose that the inactivation of syndecan gene and the consequent suppression of syndecan expression is related to the altered adhesion properties, the disappearance of epithelial phenotype, and, on the other hand, to the appearance of transformed-like phenotype in hormone-treated S115 cells.

Animals↗