Biomedical subjects

Y Capetanaki

Publications and source records attributed to Y Capetanaki.

23 records · Page 2Linked to original sources

PubMed1993

Vimentin expression is differentially regulated by IL-2 and IL-4 in murine T cells.

IL-2 and IL-4 are T cell growth factors that are produced by different T cell subsets and have distinct roles in lymphocyte biology. Despite their importance in the immune system, little is known about the genes that these lymphokines may specifically control and the interaction of these lymphokines in regulating the expression of their target genes. In this paper, we use the factor-dependent murine T cell line (CT.4R) to investigate the interaction of IL-2 and IL-4 in regulating gene expression. We report that the intermediate filament protein vimentin is differentially regulated by these lymphokines. Cells grown in IL-2 typically express 10- to 20-fold more vimentin and vimentin RNA than those grown in IL-4, but express similar levels of other cytoskeletal proteins including actin and tubulin. Vimentin was specifically induced by IL-2 and apparently suppressed by IL-4 in normal lymph node T cells, suggesting that its differential regulation by these lymphokines is physiologically relevant. We investigated the synergy between IL-2 and IL-4 in regulating the expression of vimentin RNA and compared it to that of two other lymphokine-responsive genes, pancreatic lipase and the IL-2R alpha subunit. Complex regulatory interactions were revealed: IL-4 suppressed the ability of IL-2 to induce vimentin RNA but not IL-2R alpha RNA, whereas IL-2 inhibited the ability of IL-4 to induce lipase RNA. These results indicate that IL-2 and IL-4 can cross-regulate lymphokine-responsive genes and can simultaneously exert both positive and negative regulation of different genes within the same cell.

Animals↗

PubMed1993

Regulation of the mouse desmin gene: transactivated by MyoD, myogenin, MRF4 and Myf5.

Desmin, the muscle specific intermediate filament (IF) protein, is expressed at low levels in myoblasts and at the onset of differentiation its expression increases several fold. In an effort to explore the mechanism involved in the tissue-specific and developmentally regulated expression of desmin, we have isolated the mouse desmin gene. Sequence analysis of 976 bp 5' flanking region revealed several potential cis-acting elements: 1) Three E boxes (MyoD binding sites), namely, E1, E2 and E3, located at -79, -832 and -936, respectively; 2) one MEF2 binding site at -864; 3) a region with homology to M-CAT motif at -587; 4) several GC boxes. Transient transfections with various 5' flank deletion mutants into C2C12 muscle cells have revealed both positive and negative elements that seem to be involved in the expression of desmin. The first 81 bp upstream of the transcription initiation site, including E1 box, were sufficient to confer muscle specific expression of the desmin gene. The maximal level of expression was achieved by the construct containing up to -897 base pairs. The region between -578 to -976 behaves as a classical enhancer in the absence of which the region between -578 and -81 suppresses CAT activity. Gel electrophoretic mobility shift assays using both C2C12 muscle cell nuclear extracts as well as in vitro translated myoD/E12 and myogenin/E12 heterodimers, showed that both myoD and myogenin bind to the proximal E1 and the distal E2 boxes of the desmin promoter and enhancer respectively. Co-transfection of myoD, myogenin, MRF4 and Myf5, with the desmin-CAT construct into 10T-1/2 cells demonstrated that all these factors could transactivate desmin gene expression.

Animals↗

PubMed1990

Mouse vimentin: structural relationship to fos, jun, CREB and tpr.

We have isolated and characterized mouse cDNA clones representing the entire coding region of vimentin. RNA blot analysis of different stages during development has revealed differential control in the expression of vimentin mRNA in the different tissues studied. The nucleotide sequence extends 1800 base pairs and contains the 466 amino acid mouse vimentin polypeptide chain, flanked by 90 base pairs 5' and 312 base pairs 3' untranslated region. Conformational analysis of the deduced amino acid sequence was used to localize the three known structural domains: a non-alpha-helical N-terminal head of 81 residues, a rod-like domain of 330 residues arising from three alpha-helices, and a non-alpha-helical C-terminal domain of 55 residues. Amino acid sequence comparisons with other species revealed high sequence conservation of mouse vimentin to hamster (98.7%), human (96%), and chicken (88%) protein. Computer sequence analysis also revealed domains of significant homology between different alpha helical regions of vimentin and the DNA binding-leucine zipper domain of several proto-oncogenes and transcription regulators. Specifically, 50-70% structural similarity was observed between the basic domain of the DNA binding region of the nuclear proto-oncogene products c-fos and its related antigen fra-1, c-jun and the cAMP-responsive DNA binding protein CREB, with part of the N-terminal half region of helix 1b of vimentin. When the leucine zipper domains of all these proteins were compared to vimentin, at least two different regions of similarity in the vimentin molecule were found reaching up to 53% for jun, 60% for fos, and 76% for CREB. Further analysis revealed several domains of significant similarity (50%) between all alpha-helices of the rod domain of vimentin and the N-terminal (approximately 210 residues) activation domain tpr of the oncogenic raf.

Amino Acid Sequence↗

PubMed1989

Expression of the chicken vimentin gene in transgenic mice: efficient assembly of the avian protein into the cytoskeleton.

To study expression and function of the vimentin gene, transgenic mice were generated by microinjecting the entire chicken gene plus 2.4 kilobases of 5' and 2.6 kilobases of 3' flanking sequences. All the transgenic mice obtained had incorporated multiple copies of the gene. RNA analyses demonstrated that the chicken vimentin gene was efficiently expressed in an appropriate tissue-specific pattern and that the transcripts were properly processed, as in chicken, giving rise to two RNAs. The vimentin transgene was predominantly expressed in lens at levels of up to 10-fold the endogenous level in every transgenic line studied. The chicken vimentin transcripts were efficiently translated into polypeptides that were modified posttranslationally and could assemble into the mouse cytoskeleton. Overexpression of the chicken vimentin gene did not obviously affect the expression of the endogenous gene at the RNA or the protein level. Immunofluorescence microscopy further demonstrated that the chicken protein was properly expressed spatially in lens. However, the levels were much higher in the transgenic animals.

Animals↗

PubMed1989

Overexpression of the vimentin gene in transgenic mice inhibits normal lens cell differentiation.

To investigate the role of the intermediate filament protein vimentin in the normal differentiation and morphogenesis of the eye lens fiber cells, we generated transgenic mice bearing multiple copies of the chicken vimentin gene. In most cases, the vimentin transgene was overexpressed in the lenses of these animals, reaching up to 10 times the endogenous levels. This high expression of vimentin interfered very strongly with the normal differentiation of the lens fibers. The normal fiber cell denucleation and elongation processes were impaired and the animals developed pronounced cataracts, followed by extensive lens degeneration. The age of appearance and extent of these abnormalities in the different transgenic lines were directly related to the vimentin level. Electron microscopic analysis revealed that the accumulated transgenic protein forms normal intermediate filaments.

Animals↗