SEARCH · PubMed Health

Results for “Transient expression”

Explore indexed PubMed citations for clinical trials, systematic reviews and public health research. Read source abstracts and follow each citation to its original PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 163 records · Page 9Linked to original sources

PubMed1986

[Stimulation of human lymphocytes by phorbol acetate and a calcium ionophore induces an early transient expression of the c-fos oncogene followed by c-myc].

After stimulation of human lymphocytes by 12-0 tetradecanoyl phorbol acetate (TPA) and by the calcium ionophore A23187 an early transient expression of proto-oncogene fos followed by an expression of c-myc is observed, as has been described in rodent fibroblast stimulated by growth factors. This observation suggests a direct role of protein Kinase C and of calcium flux an the induction of cellular oncogenes fos and myc that may be associated with the early steps of activation and proliferation of lymphocytes.

Calcimycin↗

PubMed1998

Identification of UV/blue light-response elements in the Arabidopsis thaliana chalcone synthase promoter using a homologous protoplast transient expression system.

To identify DNA sequences of the Arabidopsis thaliana chalcone synthase gene (CHS) concerned with induction by UV-B and UV-A/blue light, AtCHS promoter constructions were assayed by transient expression in protoplasts prepared from two different lines of cultured A. thaliana cells. The protoplasts responded similarly to A. thaliana leaf tissue in light-dependent CHS transcript accumulation. The reporter enzyme beta-glucuronidase (GUS) was used to monitor light-responsive promoter activity. A 1972 bp promoter conferred UV-B and UV-A/blue light induction of GUS activity. Deletion to 164 bp resulted in reduced promoter strength but retention of responsiveness to UV-B and UV-A/blue light. Further deletion abolished transcriptional activity. The 164 bp promoter contains sequences closely resembling LRUPcCHS, (light-responsive unit of the Petroselinum crispum CHS promoter). This A. thaliana CHS promoter region, designated LRUAtCHS, was sufficient to confer UV-B and UV-A/blue light responsiveness to a heterologous core promoter. Mutation of sequences in LRUAtCHS corresponding to the ACGT element and the MYB recognition element of LRUPcCHS resulted in inactivation of the 164 bp and 335 bp promoter deletions. However, the mutant 668 bp promoter retained residual UV-B and UV-A/blue light-induced expression, indicating the presence of additional functional sequences upstream of -335. Mutation of a single G-box-like sequence around -442 had no effect on light responsiveness, indicating that it does not function in light regulation of this promoter. Since no difference in responsiveness to UV-B and UV-A/blue light was observed with any promoter variant, we conclude that the two phototransduction pathways regulate transcription factors which interact with common promoter elements. The results from-our analysis of a A. thaliana light-responsive promoter will facilitate the study of light-dependent gene regulation by genetic means in Arabidopsis thaliana.

Acyltransferases↗

PubMed1992

Binding of typical and atypical antipsychotic agents to transiently expressed 5-HT1C receptors.

We determined the affinities of clozapine and 21 other typical and atypical antipsychotic agents for the cloned 5-hydroxytryptamine-1C (5-HT1C) receptor. For these studies, 5-HT1C receptors were transiently expressed in COS-7 cells using the vector pSVK3-5HT1C. We discovered that clozapine and several other putative typical and atypical antipsychotic agents (loxapine greater than tiosperone greater than SCH23390 greater than fluperlapine greater than rilapine greater than chlorpromazine) had relatively high affinities (7-30 nM) for the cloned 5-HT1C receptor. Other antipsychotic agents (risperidone greater than tenilapine greater than mesoridazine greater than thioridazine greater than cis-fluphenthixol) had intermediate affinities (30-100 nM), whereas many other antipsychotics (fluphenazine greater than spiperone greater than amperozide greater than melperone greater than thiothixene greater than haloperidol, metoclopramide, pimozide, domperidone, sulpiride) had low affinities (greater than 500 nM) for the cloned 5-HT1C receptor. The results indicate that although several putative atypical antipsychotic agents have high affinities for the cloned rat 5-HT1C receptor, the spectrum of drug binding does not correlate with the atypical nature of these compounds.

Antipsychotic Agents↗

PubMed1997

Transient expression of the serotonin transporter in the developing mouse thalamocortical system.

The serotonin transporter was visualized in sections through the developing mouse thalamus by autoradiography of [3H]citalopram binding. In late gestation, a high density of transporter expression appeared in the ventrobasal thalamic complex and medial geniculate body. During the first postnatal week, binding in these areas decreased to low levels. A similar pattern of transient [3H]citalopram binding was observed in the somatosensory cortex, although the rise and decline of labeling occurred some days later. The density of the serotonergic innervation in the ventrobasal thalamic complex is known to be very low during the entire developmental period. Therefore, these data suggest that the serotonin transporter may be expressed transiently by thalamic neurons projecting to the cerebral cortex (as a "heterocarrier") which are capable of taking up serotonin in the somatosensory cortex. We propose that serotonin may act temporarily as a "false" transmitter in thalamocortical axons.

Animals↗

PubMed1993

Transient expression of the intermediate filament nestin during skeletal muscle development.

It has previously been established that skeletal muscle development is accompanied by changes in the composition of intermediate filaments: vimentin is expressed predominantly in myoblasts and desmin in adult myotubes. We show that the intermediate filament transitions during muscle development are more complex, and involve a transient expression of the recently discovered intermediate filament nestin. Nestin RNA is expressed predominantly early, in a biphasic pattern, and is markedly downregulated in adult rat muscle, whereas desmin RNA becomes more abundant throughout development. Nestin protein was found up to the postnatal myotube stage, where it colocalized with desmin in Z bands. The intracellular distribution of nestin, vimentin and desmin was analysed in the human myogenic cell line G6 before and after in vitro differentiation. Despite its more distant evolutionary and structural relationship to the other two intermediate filaments, nestin formed a cytoplasmic filamentous network indistinguishable from that of desmin and vimentin, both in undifferentiated myoblasts and after differentiation to multinuclear myotubes. In conclusion, our data suggest that nestin is an integrated component of the dynamic intermediate filament network during muscle development and that nestin copolymerizes with desmin and vimentin at stages of coexpression.

Animals↗

PubMed1991

The transient expression of type II collagen at tissue interfaces during mammalian craniofacial development.

Using immunocytochemical techniques, the spatiotemporal distribution of the major collagen isoform of cartilage, type II collagen, has been investigated during early craniofacial development in the mouse embryo. Early and transient expression was associated with the otic and optic vesicles, the ventrolateral surfaces of the developing brain, olfactory conchi, endocardial and mesocardial tissues, the lateral and basal surfaces of the pharyngeal endoderm and beneath the ectoderm of the branchial arches. A number of these locations are sites of epithelial-mesenchymal tissue interaction believed to generate the component parts of the chondrocranium; here, type II collagen appears transiently in advance of overt chondrogenesis in the mesenchyme. At such sites, immunofluorescence is typically localised along the basal surface of the epithelial partner, with the strongest reaction detected between the basal aspects of the otic and rhombencephalic epithelia. Immunoelectron microscopy, using pre-embedding immunostaining and a protein G-gold technique, reveals that the type II collagen is adjacent to, but not integral with, the basal laminae. Gold particles are clearly associated with 10-15 nm fibrils of the extracellular matrix in the reticulate lamina region. The pattern of type II collagen expression in the mouse closely correlates with that demonstrated previously in the quail, indicating a high degree of phylogenetic conservation between these two vertebrate species. These findings are consistent with the hypothesis that the pattern of epithelial secretion of type II collagen, or a coexpressed matrix molecule, constitutes a morphogenetic signal, realised as a matrix-mediated tissue interaction, and specifying the form of the vertebrate chondrocranium. Three-dimensional reconstruction of early type II collagen distribution, and of the subsequent chondrocranial cartilages, reveals that chondrocranial form can be derived from a 'pre-pattern' of epithelially derived type II collagen expressed at epithelial-mesenchymal tissue interfaces.

Animals↗

PubMed1991

Transient expression analyses of DNA extending 2.4 kb upstream of the human renin gene.

The influence on homologous and heterologous promoter activity of DNA extending 2.4 kb upstream of the human renin gene (REN) was examined by transient expression assay in JEG-3 cells, using the gene for chloramphenicol acetyl transferase (CAT) as reporter, and cotransfection with pCH110 to control for transfection efficiency. Analyses of constituent subfragments of the region 5' of residue -144, using the herpes simplex virus thymidine kinase (tk) promoter to drive transcription, provided no evidence for negative regulatory influences within the -2400 to -144 DNA. That distal 5'-flanking DNA may have little influence on promoter activity is further supported by a sharp decline in nucleotide homology between human, rat and mouse renin genes further upstream than human residue -604. Constructs containing renin DNA to residue +13, i.e., which retained the REN promoter, all displayed very low CAT activities, consistent with negative cis-acting control within the -149 to +13 region. This finding contrasts with results of similar studies for mouse, in which renin gene control was suggested to be mediated primarily via cell-specific trans-acting activator(s) acting on yet-to-be identified enhancer(s). Mouse renin genes have, however, a common DNA insertion that could have disrupted the negative element in this region, and which might contain enhancer target(s) for trans-acting factor(s). In conclusion, the present study involving JEG-3 cells has demonstrated that distal human renin 5'-flanking DNA has little cis-acting influence on promoter activity, whereas DNA located within 100 base pairs of the renin promoter may have a negative regulatory effect.(ABSTRACT TRUNCATED AT 250 WORDS)

Cloning, Molecular↗

PubMed1986

Transient expression of murine interferon-alpha genes in mouse and monkey cells.

The coding regions of murine interferon-alpha (IFN-alpha) genes were combined with promoter and 3'-noncoding sequences from other eukaryotic genes. Transient expression of these fusion genes was achieved in monkey COS cells and in a mouse cell line (TOP cells) expressing polyoma virus (Py) large T antigen constitutively. The efficiency of the different expression plasmids was determined by measuring the amount of IFN secreted into the medium. Replacement of the 3'-noncoding region of an IFN-alpha gene by that of the rabbit beta-globin gene resulted in a fourfold higher IFN-alpha production. The SV40 early promoter and the Moloney murine leukemia virus (MoMLV) long terminal repeat (LTR) produced similar amounts of IFN-alpha in COS cells. However, a tandem combination of the SV40 enhancer/early promoter and the mouse metallothionein-I promoter appeared fivefold more active than the SV40 early promoter. In TOP cells the MoMLV LTR was found to be threefold more active than the Py early promoter.

Animals↗

PubMed1992

Infectious Sindbis virus transient expression vectors for studying antigen processing and presentation.

Sindbis virus (SIN) is a small positive-strand enveloped RNA virus that infects a broad range of vertebrate and insect cells. A SIN vector (called dsSIN), designed for transient expression of heterologous RNAs and proteins, was engineered by inserting a second subgenomic mRNA promoter sequence into a nonessential region of the SIN genome. By using this vector, dsSIN recombinants have been constructed that express either bacterial chloramphenicol acetyltransferase, a truncated form of the influenza hemagglutinin (HA), or mini-genes encoding two distinct immunodominant cytotoxic T lymphocyte (CTL) HA epitopes. Infection of murine cell lines with these recombinants resulted in the expression of approximately 10(6)-10(7) chloramphenicol acetyltransferase polypeptides per cell and efficient sensitization of target cells for lysis by appropriate major histocompatibility complex-restricted HA-specific CTL clones in vitro. In addition, priming of an influenza-specific T-cell response was observed after immunizing mice with dsSIN recombinants expressing either a truncated form of HA or the immunodominant influenza CTL epitopes. This SIN expression system allows the generation of high-titered recombinant virus stocks in a matter of days and should facilitate mapping and mutational analysis of class I major histocompatibility complex-restricted T-cell epitopes expressed via the endogenous pathway of antigen processing and presentation.

Amino Acid Sequence↗

PubMed1984

High level transient expression of a chloramphenicol acetyl transferase gene by DEAE-dextran mediated DNA transfection coupled with a dimethyl sulfoxide or glycerol shock treatment.

Using a plasmid containing the bacterial chloramphenicol acetyl transferase gene, we have assayed for transient expression of DNA introduced into mouse L cells by a variety of transfection conditions. High efficiency uptake and expression of this foreign DNA have been achieved by modifying the DEAE dextran mediated transfection procedure of McCutchan and Pagano (1) to include a shock with either dimethyl sulfoxide or glycerol. Inclusion of the shock step can increase expression of the transfected gene a surprising approximately 50 fold. With plasmid constructs that do not replicate after transfection, we can readily detect CAT activity in an overnight autoradiographic exposure from less than 0.1% of an extract from a 60 mm dish of transfected cells. We have determined the amounts of DNA, the amount and time course of DEAE-dextran and dimethyl sulfoxide treatments, the effects of additional DNA, and the time after transfection which yield maximal expression. Overall, this transfection protocol using DEAE-dextran coupled to a shock treatment is simple, straightforward, and gives consistently high levels of expression of the input DNA.

Acetyltransferases↗

PubMed1990

Transient expression of full-length and truncated forms of the human nerve growth factor receptor.

To facilitate the characterization of nerve growth factor (NGF) receptor and mutated forms of the receptor, we have set up a rapid, efficient transient expression system utilizing COS cells. The human NGF receptor is a 427 amino acid protein with a hydrophobic signal sequence, a 222 amino acid extracellular domain, a single transmembrane domain and a 155 amino acid intracellular domain. The NGF receptor and a truncated form lacking the cytoplasmic domain were expressed in a COS cell expression system. Both recombinant proteins were detected on the cell surface and at a perinuclear site. Specific binding of 125I-NGF to the recombinant proteins was detected by chemical cross-linking. The extracellular domain of the NGF receptor was also expressed in the same system and detected in the COS cell endoplasmic reticulum and in the culture supernatant. This recombinant protein also specifically binds NGF.

Animals↗

PubMed1992

Neuropeptide Y m-RNA and peptide are transiently expressed in the developing rat spinal cord.

The distribution of neuropeptide Y (NPY) m-RNA and peptide were studied during the ontogeny of the rat spinal cord, by means of in situ hybridization and immunohistochemistry. NPY expressing neurons were visualized by both techniques, although by in situ hybridization they appeared slightly more abundant than by immunohistochemistry. In the ventral horn, a few neurons transiently expressed NPY from the embryonic day 15 to birth, but not in the adult animals. In the embryonic dorsal horn, NPY expressing neurons were numerous and seen as early as embryonic day 15 (E15). In contrast, in the postnatal and the adult dorsal horn, the number of neurons expressing the NPY phenotype was dramatically lower.

Animals↗

PubMed1991

Patterns of transient expression of the arenavirus nucleocapsid protein gene in transfected cells.

The cloned genes for the nucleocapsid proteins N of Junín and LCM (lymphocytic choriomeningitis) arenaviruses were inserted into the SV40-derived expression vector designated pKG4. When BHK-21 (baby hamster kidney fibroblasts) and CV-1 (African green monkey kidney fibroblasts) cell lines were transfected using these constructions, the transient expression yielded a polypeptide that could not be distinguished either by size nor by immunoreactivity from the N protein synthesized during the viral infection. The immunofluorescence analysis showed a pattern of intracellular localization similar to that observed in virus infected cells, i.e. varying from a diffuse cytoplasmic staining to granules, either distributed throughout the cytoplasm or concentrated in the perinuclear region. The association of the N protein with basophilic granules is similar to that observed in the cytopathic effect caused by arenaviruses, and could be related to the physicochemical properties of this polypeptide containing numerous basic amino acid sequences, that would allow for the interaction with cellular RNAs.

Animals↗

PubMed1994

Transient expression of bile-duct-specific cytokeratin in fetal mouse hepatocytes.

The differentiation of hepatocytes and biliary epithelial cells has been histochemically analyzed with anti-calf cytokeratin antiserum in the fetal mouse liver. Almost all young fetal hepatocytes transiently express bile-duct-specific cytokeratin; subsequently, the strong staining of the cytokeratin is confined to progenitor cells of intrahepatic biliary epithelial cells around portal veins. These results suggest that all fetal hepatocytes are bi-potent in terms of the differentiation of mature hepatocytes and intrahepatic bile-duct cells, and that the microenvironment around portal veins plays an important role in bile-duct differentiation. Large periportal hepatocytes continue to stain weakly for cytokeratin until 2 weeks after birth, although the number of positive hepatocytes decreases with development. The differentiation of bile ducts from periportal hepatocytes may continue for 2 weeks after birth.

Albumins↗

PubMed1991

Transient expression of the cloned mouse c-Ha-ras 5' upstream region in transfected primary SENCAR mouse keratinocytes demonstrates its power as a promoter element.

The mouse Ha-ras oncogene is activated by point mutation and overexpressed in developing papillomas during two-stage skin carcinogenesis in SENCAR mice. One of our research aims is to characterize the factors regulating Ha-ras gene expression at the transcriptional level in SENCAR mouse epidermis. Towards this goal, we sequenced 1400 bp of the 5' upstream region of the mouse Ha-ras gene so as to characterize various cis-regulatory elements present in the gene. We identified seven sites with the proper consensus sequence for binding the SP1 transcription factor and three potential binding sites for the CTF-1 factor. In addition, we located a 13-base sequence with 92% homology to the consensus sequence for an estrogen response element and two hexamers with consensus sequences identical to the core sequence of the glucocorticoid response element. A series of transient gene expression vectors was constructed in which various regions of the mouse Ha-ras 5' upstream region were fused to the chloramphenicol acetyltransferase (CAT) gene. These expression plasmids were transfected into newborn and adult primary SENCAR epidermal cells, the epidermal cell population that presumably contains the stem cells involved in two-stage skin tumorigenesis. Transient gene expression assays carried out after 48-72 h indicated that a 2.3-kb Ha-ras 5' fragment produced CAT activity comparable to that produced by pSV2CAT and pdolCMVCAT, both of which are plasmids with strong viral promoters and enhancers driving CAT gene expression. Maintenance of transfected keratinocytes under both nondifferentiating (0.05 mM calcium) and differentiating (1.2 mM calcium) culture conditions demonstrated that the mouse Ha-ras upstream region was relatively unresponsive to changes in calcium concentration in transient expression assays carried out in either newborn or adult keratinocytes. Our results demonstrated the power of the cloned mouse Ha-ras promoter and upstream region in driving transient gene expression after transfection into primary keratinocytes.

Animals↗

PubMed1982

Transient expression of the chicken lysozyme gene after transfer into human cells.

The chicken lysozyme gene was inserted into an SV40-based plasmid vector, and the recombinants were transfected into the human cell lines HeLa and MCF-7. Correct and efficient transient expression directed by the lysozyme promoter was found in both of these cell lines, as determined by S1 nuclease mapping and Northern blot analysis of the RNAs made. SV40 sequences dramatically enhance the expression of the lysozyme gene. This enhancing effect is only acting in cis and is distance/orientation dependent, since clones containing the lysozyme gene in either orientation produce different amounts of correct lysozyme transcripts. The transfected lysozyme gene was not induced by steroid hormone treatment of the cells.

Breast Neoplasms↗

PubMed1988

Transient expression and sequence of the matrix (M1) gene of WSN influenza A virus in a vaccinia vector.

A cDNA encoding the entire amino acid sequence of the matrix (M1) protein of influenza A/WSN/33 virus was cloned, sequenced, and expressed in a vaccinia virus system consisting of the T7 bacteriophage RNA polymerase and a plasmid carrying the M1 gene flanked by T7 polymerase promoter and terminator sequences. The transiently expressed M1 gene product comigrated on SDS-polyacrylamide gels with the endogenous WSN virus M1 protein and was recognized in Western blot analysis by three epitope-specific monoclonal antibodies directed to the M1 protein. The nucleotide sequence and the predicted amino acid sequence of the cloned WSN virus M1 coding region was found to be more than 97% homologous to that of the M1 gene of influenza virus A/PR/8/34 reported by G. Winter and S. Fields (Nucleic Acids Res. 8, 1965-1974, 1980).

Amino Acid Sequence↗

PubMed1991

Mammalian achaete-scute homolog 1 is transiently expressed by spatially restricted subsets of early neuroepithelial and neural crest cells.

Using monoclonal antibodies, we have examined the expression pattern of MASH1, a basic helix-loop-helix protein that is a mammalian homolog of the Drosophila achaete-scute proteins. In Drosophila, achaete-scute genes are required for the determination of a subset of neurons. In the rat embryo, MASH1 expression is confined to subpopulations of neural precursor cells. The induction of MASH1 precedes, but is extinguished upon, overt neuronal differentiation. MASH1 is expressed in the forebrain by spatially restricted domains of neuroepithelium and in the peripheral nervous system exclusively by precursors of sympathetic and enteric neurons. The features of early and transient expression, in spatially restricted subpopulations of neural precursors, are similar to those observed for achaete-scute. Thus, the amino acid sequence conservation between MASH1 and achaete-scute is reflected in a parallel conservation of cell type specificity of expression, similar to the case of mammalian MyoD and Drosophila nautilus. These data support the idea that helix-loop-helix proteins may represent an evolutionarily conserved family of cell-type determination genes, of which MASH1 is the first neural-specific member identified in vertebrates.

Animals↗