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At least 739 records · Page 41Linked to original sources
PubMed1983

Size, location and polarity of T-DNA-encoded transcripts in nopaline crown gall tumors; common transcripts in octopine and nopaline tumors.

Up to thirteen T-DNA-encoded, polyadenylated transcripts of different relative abundance were detected by Northern blot hybridization in the tobacco nopaline BT37 crown gall teratoma tissue. Their sizes range from 900 to 2,700 bases. The polarity of eight of the thirteen transcripts was assigned by hybridization of labeled RNA to single-stranded DNA fragments of the T-region obtained by cloning in an M13 vector. Both strands of the T-DNA are transcribed. Our data indicate that most, if not all, transcripts are generated via independent promoter and poly(A)-addition sites on the T-DNA. Comparison of T-DNA-encoded transcripts present in crown gall tumors showing teratoma-like growth (BT37) with those from an unorganized tumor line (W38C58) reveals that this difference in phenotype is accompanied by a difference in the expression of the T-DNA. T-DNA sequences common to both octopine and nopaline tumors encode at least five, and probably six, cross-hybridizing transcripts of the same size, location, polarity and function. These transcripts are involved in the process of plant tumor formation and maintenance.

Arginine↗
PubMed1984

The transcriptional regulation of Xenopus 5s RNA genes in chromatin: the roles of active stable transcription complexes and histone H1.

The properties of active and repressed 5S rna genes in somatic cell chromatin from Xenopus laevis cultured cells were studied by transcription in vitro. The somatic 5S RNA genes, which are active in vivo, are packaged in chromatin as active stable transcription complexes lacking only RNA polymerase III for transcription activity. The oocyte 5S RNA genes, which are inactive in somatic tissues, do not have transcription factors bound to them in purified chromatin and are prevented from binding these factors by a structure dependent on histone H1. In chromatin active stable transcription complexes protect 5S RNA genes from histone H1-mediated repression and H1 binding prevents the formation of stable active transcription complexes.

Animals↗
PubMed1983

Electron microscopic studies of transcriptional complexes released from vaccinia cores during RNA-synthesis in vitro: methods for fractionation of transcriptional complexes.

Electron microscopic (EM) and biochemical methods were employed to study the transcriptional complexes present in detergent lysates of vaccinia virus cores actively synthesizing RNA in vitro. When processed and examined in the EM, 14 'transcriptional sites' could be observed on full-length DNA templates. Fractionation of lysates by equilibrium density centrifugation in CsSO4, chromatography on hydroxyapatite columns or by sedimentation in sucrose gradients, allowed isolation of DNA templates associated with transcripts but these manipulations often resulted in fragmentation of the DNA template or promoted the release of transcripts from the template. It is suggested that RNA transcripts remain associated with the template in regions of supercoiling. These regions, in turn, may be maintained by DNA-protein interactions which are compromised as the transcriptional complexes are fractionated and purified.

Centrifugation, Density Gradient↗
PubMed1987

Effects of ppGpp on transcription by DNA-dependent RNA polymerase from Escherichia coli: circular dichroism, absorption and specific transcription studies.

Concrete evidence is presented for conformational changes elicited in RNA polymerase upon binding ppGpp by circular dichroism measurements. In the presence of 100 microM ppGpp, the molar ellipticity of RNA polymerase at 220 nm is reduced by 14% from the initial value of - 11,100 deg X cm2 X dmol-1 at 25 degrees C. In vitro transcription on templates containing the beta-lactamase promoter and colicin E1 promoter on poly[d(A-T)] is inhibited by ppGpp. None of these templates had GC-rich nucleotide sequence near the transcription initiation site, and yet they were influenced by ppGpp. Comparison of the effect on the synthesis of mRNAs for beta-lactamase and colicin E1 and the synthesis of the proteins themselves indicates that the effect of ppGpp is at the level of transcription for the former case and involves coupled transcription-translation for the latter case. Difference absorption, polyacrylamide gel electrophoresis, and nitrocellulose filter-binding studies show that the binding of ppGpp to RNA polymerase does not impair the extent of the interaction between enzyme and DNA. Kinetic studies suggest that ppGpp affects transcription initiation on beta-lactamase promoter. On poly[d(A-T)], ppGpp affects the rate of open complex formation and is a mixed inhibitor with respect to the incorporation of nucleotides. Our results are consistent with the idea that ppGpp acts as a regulator by binding at a site different from the active site and changes the RNA polymerase conformation, causing altered transcriptional behavior on different DNA templates.

Circular Dichroism↗
PubMed1990

Cell cycle-dependent transcriptional and post-transcriptional regulation of chloroplast gene expression in Chlamydomonas reinhardtii.

The regulated expression of five chloroplast genes in Chlamydomonas reinhardtii during a 24 hour cell cycle (12 hours light, 12 hours dark) was analyzed. Transcription rates of the genes encoding the two reaction center proteins of Photosystem I (psaA, psaB), the subunits alpha and beta (atpA, atpB) of chloroplast ATP synthase and for chloroplast elongation factor tu (EF-tu) were measured during the cell cycle. All genes are maximally transcribed at the beginning of the light period. Transcription was induced before the onset of illumination by a light-independent mechanism. Transcript abundance of the same genes during the cell cycle was determined by quantification of Northern blots hybridized with gene-specific probes. The atpA, atpB and psaB mRNAs were most abundant in the first 6 hours of the light period and decreased to about 15% of maximum in the dark. The abundance of psaA mRNA showed less variation and was maximal around the middle of the cell cycle. The EF-tu mRNA showed a maximum early in the light period, but decreased to almost undetectable levels in the second half of the light period. Because of the similar transcriptional patterns observed, the differential steady state levels of these chloroplast transcripts appeared to be regulated at the post-transcriptional level.

Blotting, Northern↗
PubMed1987

Transcriptional analysis of the cyanobacterial gvpABC operon in differentiated cells: occurrence of an antisense RNA complementary to three overlapping transcripts.

Cyanobacteria are photosynthetic prokaryotes able to colonize almost all kinds of ecosystems. Some of them exhibit differentiation processes and/or may establish tight symbiotic associations. Upon changes in the environmental conditions, the cyanobacterium Calothrix 7601 differentiates hormogonia which are short filaments of small cells resulting from cellular division and fragmentation of the long filaments of vegetative cells. In Calothrix, hormogonia are characterized by their gliding motility and by a massive production of gas vesicles which confer buoyancy. At least four genes are involved in the formation of gas vesicles, three of which are organized in one operon (gvpABC). Four different RNA species, only present after induction of hormogonia differentiation, result from transcription of this operon. Mapping of the 5' and 3' ends of these transcripts demonstrates the presence of gvpA, gvpAB and gvpABC transcripts, all three having the same 5' end. Each of the three transcripts terminates a few bases downstream from stem-and-loop structures. Most interestingly, the fourth transcript is an antisense RNA starting from the 3' end of the gvpB gene and ending within the gvpA gene. This antisense RNA can thus form an homologous duplex with the three other transcripts, thereby being able to impair translation and/or modify mRNA stability.

Chromosome Mapping↗
PubMed1996

Detection of feline immunodeficiency virus transcripts by quantitative reverse transcription-polymerase chain reaction.

We developed a quantitative reverse transcription-polymerase chain reaction (RT-PCR) procedure to estimate replication status of feline immunodeficiency virus (FIV) in peripheral blood mononuclear cells (PBMCs) of cats. Primers used for the RT-PCR were designed to detect only multiple spliced transcripts of FIV and allowed us to detect the specific transcripts with high specificity. By using limiting-cell-dilution RT-PCR, we demonstrated that the specific transcripts were quantitatively detected in a single infected cell in a background of 1 x 10(6) uninfected cells without Southern blot hybridization. Furthermore, the transcripts were observed efficiently in all PBMCs of the chronically FIV-infected cats when examined by this RT-PCR technique. There results demonstrated that this RT-PCR method is applicable for specific detection of the FIV-specific transcripts in the PBMCs and for estimation of the viral replication status in vivo.

Animals↗
PubMed1994

The human progesterone receptor A-form functions as a transcriptional modulator of mineralocorticoid receptor transcriptional activity.

The human progesterone receptor (hPR) exists as two distinct molecular forms in most cells, hPR-A and -B. These receptor isoforms display distinct biological functions and demonstrate a cell and promoter specific ability to regulate gene transcription. In cellular contexts where hPR-A is transcriptionally inactive it can function as a ligand dependent inhibitor of mineralocorticoid receptor (MR) transcriptional activity. Inhibition occurs by a non-competitive mechanism as direct binding to MR is not required. Interestingly, PR agonists differ in their ability to facilitate the inhibitory function of hPR-A, suggesting that a specific receptor conformation may be preferred for this activity. Those compounds derived from 19-nor-testosterone are the most effective. The antiprogestins RU486, ZK98299 and ZK112993 are effective MR antagonists in the presence of coexpressed hPR-A. The mechanism of hPR-A mediated inhibition of MR transcriptional activity is unknown. We propose that inhibition results from a competition of hPR-A with MR for a common transcription factor and that the association of hPR-A with this factor is not transcriptionally productive.

Animals↗
PubMed2006

Actin-based modeling of a transcriptionally competent nuclear substructure induced by transcription inhibition.

During transcription inactivation, the nuclear bodies in the mammalian cells often undergo reorganization. In particular, the interchromatin granule clusters, or IGCs, become colocalized with RNA polymerase II (RNAP II) upon treatment with transcription inhibitors. This colocalization has also been observed in untreated but transcriptionally inactive cells. We report here that the reorganized IGC domains are unique substructure consisting of outer shells made of SC35, ERK2, SF2/ASF, and actin. The apparently hollow holes of these domains contain clusters of RNAP II, mostly phosphorylated, and the splicing regulator SMN. This class of complexes are also the sites where prominent transcription activities are detected once the inhibitors are removed. Furthermore, actin polymerization is required for reorganization of the IGCs. In connection with this, immunoprecipitation and immunostaining experiments showed that nuclear actin is associated with IGCs and the reorganized IGC domains. The study thus provides further evidence for the existence of an actin-based nuclear skeleton structure in association with the dynamic reorganization processes in the nucleus. Overall, our data suggest that mammalian cells have adapted to utilize the reorganized, uniquely shaped IGC domains as the temporary storage sites of RNAP II transcription machineries in response to certain transient states of transcription inactivation.

Actins↗
PubMed1984

Induced transcription of the mouse beta-globin transcription unit in erythroleukemia cells. Time-course of induction and of changes in chromatin structure.

The transcription of the beta-globin genes in mouse erythroleukemia cells has been examined by hybridizing labeled RNA obtained from isolated nuclei after chain elongation in the presence of [alpha-32P]UTP. There is induction of at least 30-fold of beta maj globin transcription after cells are treated with either dimethylsulfoxide or hexamethylene bisacetamide. The induction requires 36 to 48 hours to be maximal, during which time the cells double about three to four times. During this time, a site in the beta maj DNA region becomes hypersensitive to DNase. The development of this hypersensitive site is co-ordinate with the transcriptional increase. The induced transcripts in the beta-globin region are alpha-amanitin-sensitive (and therefore are RNA polymerase II products). An examination of weak transcriptional signals to DNA fragments upstream of the beta maj globin gene in uninduced mouse erythroleukemia cells and in cells that do not make globin is also reported. The low level of hybridization to the upstream regions in uninduced erythroleukemia cells, in L cells (a fibroblast) and in a strain of erythroleukemia cells that no longer make globin are not equally sensitive to alpha-amanitin as in the induced signal. These experiments help define the inducible transcription unit for beta maj globin mRNA production.

Animals↗
PubMed1997

Cytochrome P4501A1 and cytochrome P4501A2 are downregulated at both transcriptional and post-transcriptional levels by conditions resulting in interferon-alpha/beta induction.

The interferon mediated downregulation of constitutive and inducible cytochrome P450 enzymes occurs through a pretranslational mechanism which depresses the mRNA encoding cytochrome P450s. We measured the transcription rates of CYP1A genes and the turnover of CYP1A mRNA in rats treated with the interferon-alpha/beta inducer polyinosinic acid-polycytidylic acid. The rate of transcription of CYP1A1 and CYP1A2 genes was significantly decreased in hepatic nuclei isolated from male rats treated with polyinosinic acid-polycytidylic acid (10 mg/kg). In addition the rate of degradation of hepatic CYP1A1 and CYP1A2 mRNA was examined following the inhibition of de novo transcription by actinomycin D (1 mg/kg). Messenger RNA levels were analysed by Northern and slot blotting with a 1.2 kb murine CYP1A1 cDNA probe. Interferon significantly augmented the rate of loss of CYP1A1 and CYP1A2 mRNAs suggesting that post-transcriptional degradation of mRNA contributes to the pre-translational events that cause cytochrome P450 downregulation. These results support the involvement of both transcriptional and post-transcriptional mechanisms in the loss of cytochrome P450s mediated by interferon inducers.

Animals↗
PubMed1997

Thymocytes and cultured thymic epithelial cells express transcripts encoding alpha-3, alpha-5 and beta-4 subunits of neuronal nicotinic acetylcholine receptors: preferential transcription of the alpha-3 and beta-4 genes by immature CD4 + 8 + thymocytes.

Thymic tissues express transcripts encoding the alpha-3, alpha-5 and beta-4 subunits of nicotinic neuronal acetylcholine receptors (AcChRs) suggesting that neuronal AcChRs similar to those expressed in ganglia are expressed in the thymus. Transcription occurs in both isolated thymocytes and thymic epithelial cells. RT-PCR analyses of thymocyte subsets indicate that immature CD4 + 8 + thymocytes express higher levels of the alpha-3 and beta-4 transcripts than more mature thymocytes. Compared to freshly isolated thymocytes, peripheral blood lymphocytes do not express alpha-3 and beta-4 AcChR subunit transcripts. Cultured thymocytes rapidly down-regulate transcription of the alpha-3 and beta-4 AcChR subunit genes by a process that is not reversed by stimulation with phytohemagglutinin and IL-2. Thus our results indicate that there is transcriptional regulation of neuronal AcChR subunit genes during the process of thymocyte maturation and that factors within the thymic microenvironment influence expression of the alpha-3 and beta-4 AcChR subunit genes by developing T cells.

Amyloid beta-Protein Precursor↗
PubMed1997

Transcription of the vasoactive intestinal peptide gene in response to glucocorticoids: differential regulation of alternative transcripts is modulated by a labile protein in rat anterior pituitary.

Expression of the vasoactive intestinal peptide (VIP) gene is controlled by glucocorticoids in a tissue- and endocrine status-specific manner. We have investigated the molecular mechanisms that determine glucocorticoid regulation of VIP gene expression in the rat pituitary. In initial experiments, using explant cultures of rat pituitary glands, we have demonstrated that treatment with the glucocorticoid agonist dexamethasone leads to a marked increase in VIP mRNA levels. This effect was found to be selective for the larger of two alternatively polyadenylated VIP transcripts, and in addition, protein synthesis inhibitors markedly enhanced the magnitude of this response indicating that a labile pituitary protein acts to attenuate the transcript-selective response to glucocorticoids. Nuclear run-on analysis of transcription demonstrated that the effects of dexamethasone in vitro are mediated largely, if not completely, at the level of transcription. In order to investigate the role of VIP promoter sequence in the glucocorticoid response, we then demonstrated that the activity of rat VIP gene promoter/reporter constructs in GH3 pituitary cells are up-regulated by dexamethasone. This up-regulation is virtually abolished following removal of promoter sequence between -162 and -89 of the start of transcription. Using an in vitro electrophoretic mobility shift assay, we have also demonstrated that this region of the promoter binds recombinant glucocorticoid receptor protein. The results of our study therefore indicate a direct mechanism of action for the modulation of VIP gene expression by glucocorticoids, and furthermore provide evidence of a mechanism that permits selective glucocorticoid regulation of alternative VIP transcripts.

Animals↗
PubMed1996

Point mutations in a transcription terminator, lambda tI, that affect both transcription termination and RNA stability.

The terminator tI is located approx. 280 nucleotides beyond the int gene of bacteriophage lambda. Besides its role as a transcription terminator, tI may confer stability to the int message by protecting it from 3' exonucleolytic degradation. In order to study the role of the tI sequence in transcription termination and RNA stability, three different point mutations tI1, tI2, and tI3 were isolated and characterized. All the tI mutations map in the G + C-rich region of dyad symmetry in the terminator and decrease the transcriptional termination of tI in vivo from 99% for the wild type terminator to 81-93% as determined by galactokinase activity and in vitro from 80% for the wild type terminator to 8-12% using the E. coli RNA polymerase. Additionally, the tI mutations cause upstream transcript instability in vivo. This instability defect caused by tI mutations is compensated by the host mutant deficient in polynucleotide phosphorylase resulting in increased steady state levels of these mutant transcripts. The results show that the intact hairpin of tI is essential for efficient transcription termination and for maintaining mRNA stability by blocking the 3' to 5' exonucleolytic activity of polynucleotide phosphorylase.

Bacteriophage lambda↗
PubMed1989

Transcription of human type I collagen genes. Variation in the relative rates of transcription of the pro alpha 1 and pro alpha 2 genes.

The relative rates of transcription of the two genes for type I collagen have been measured in a runoff transcription assay using nuclei isolated from cultured human fibroblasts. Control experiments indicated that the ratio of pro alpha 1(I)/pro alpha 2(I) transcripts detected with a given nuclear preparation did not vary over a range of transcription times, nuclei concentrations, and amounts of filter-bound cDNA used in the assay. However, a significant difference in the ratio was observed when nuclei isolated from cells grown under different conditions were used. Nuclei from sub-confluent cultures generally transcribed the two genes in a ratio of 2:1 or lower, while nuclei from post-confluent cultures transcribed the genes in a much higher ratio of about 4:1. Analysis of the amount of pro alpha 1(I) and pro alpha 2(I) RNA transcribed suggests that it is the transcription of the pro alpha 1(I) gene, and not the pro alpha 2(I) gene, that varies with culture conditions. The steady state ratios of pro alpha 1(I)/pro alpha 2(I) RNA remained near 2:1 under all conditions. Thus, some post-transcriptional mechanism apparently results in maintenance of the steady-state pro alpha 1(I)/pro alpha 2(I) RNA ratios at approximately 2:1.

Cell Line↗
PubMed1997

Interferon-gamma regulates collagen and fibronectin gene expression by transcriptional and post-transcriptional mechanisms.

Interferon-gamma (IFN-gamma) regulates the expression of collagen and fibronectin genes by molecular mechanisms not completely understood. We investigated the effects of IFN-gamma on the expression of the genes encoding alpha 1 (I) procollagen (COL1A1), alpha 1 (III) procollagen (COL3A1), and fibronectin (FN) in cultured normal human lung fibroblasts. Labeled newly synthesized proteins were analysed by electrophoresis, mRNA levels and stability by Northern hybridizations, and transcription rates by in vitro assays. IFN-gamma caused a reduction in the production of alpha 1 (I) and alpha 1 (III) procollagens and of fibronectin. The reduction in the production of procollagen chains was shown to result from a combination of IFN-gamma-induced inhibition of the transcription rates of the COL1A1 and COL3A1 genes and destabilization of the corresponding transcripts. IFN-gamma increased the transcription rate of FN, but also decreased the stability of the corresponding transcripts. The net results indicate that the regulation of the expression of extracellular matrix genes by IFN-gamma is a complex process that involves changes in gene transcription rates, alterations in mRNAs stability, and possibly, modulation of the rates of translation.

Cells, Cultured↗
PubMed2000

Mapping of mRNA transcripts in the genome of molluscum contagiosum virus: transcriptional analysis of the viral slam gene family.

Molluscum contagiosum virus (MCV) is a member of the poxvirus family and causes benign skin tumors in children and immunocompromised individuals. The primary structure and coding capacity of MCV was previously determined by DNA nucleotide sequencing (Senkevich et al., Science 273, 813-816, 1996). Hypothetical genes were predicted based on (i) amino acid homologies with known genes, (ii) presence or absence of conserved transcription regulation signals, and (iii) algorithms based on learning sets of coding sequences. These methods provide a rational basis for the prediction of MCV coding sequences. However, the existence and exact size of MCV open reading frames and the precise position of transcription regulation signals can only be determined by MCV mRNA transcript mapping experiments. We developed methods for the characterization of the mRNA transcripts of MCV genes in infected skin tissue and abortively infected human fibroblast cell cultures. Using these methods the properties of the mRNA transcripts of the MCV SLAM (signaling lymphocytic activating molecule) gene family (mc002L, mc161R, and mc162R) were analyzed. The mRNA start site found for the mc161R transcript suggests that a second start codon is used leading to a mc161R open reading frame that is nine amino acid residues shorter than predicted.

Antigens, CD↗
PubMed2000

Does the mitochondrial transcription-termination complex play an essential role in controlling differential transcription of the mitochondrial DNA?

The mechanism of mitochondrial transcription is well documented although the method of regulation remains obscure. The mitochondrial transcription-termination complex, mTERF, holds a key position in determining the fate of heavy-strand promotor-initiated transcripts and has been suggested as a candidate in the regulation of mitochondrial DNA (mtDNA) transcription. We report here the first example of a modulation of mTERF-complex binding activity concomitant with a differential mtDNA transcription rate. We suggest that these observations are indicative of a method of intra-organellar transcriptional fine tuning.

Cytosol↗