Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:2.7.7.6 (RNA polymerase)
34,946 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Regulation of the membrane cofactor protein (MCP: CD46) was examined. While the expression of MCP in mice carrying MCP(BC2) cDNA with 125 bp of 3' untranslated region (3'UT) was minimal, that in mice carrying MCP cDNA without total 3' UT was evident in many organs. Reverse transcriptase polymerase chain reaction (RT-PCR) analysis clearly showed the presence of mRNA even in transgenic mice with 3' UT, suggesting that the expression was regulated at the post-transcriptional stage. The in vitro expression data of MCP molecules on the stable Chinese hamster ovary (CHO) cell clone corresponded to that in transgenic mice. The first 125 bp downregulated the expression of MCP molecules in combination with not only beta-actin, but also SR alpha, promoter. Also, this region inhibited expression of decay accelerating factor (DAF: CD55) molecules when it was inserted into cDNA of DAF. Furthermore, the first 32 bp of the 3' UT revealed the same downregulation effect as 125 bp on MCP molecules. These findings indicated that the first 125 bp (and the first 32 bp in particular) of 3' UT regulate the expression of MCP molecules in transgenic mice.
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PMID:The regulation of membrane cofactor protein (CD46) expression by the 3' untranslated region in transgenic mice. 916 42

The association of increased metallothionein (MT) gene expression in breast cancer with metastasis and poor prognosis has led us to investigate the hypothesis that inhibition of MT gene expression may elicit antiproliferative effects in breast carcinoma MCF7 cells. To monitor the effect of downregulation of MT protein on growth, MCF7 cells were transiently transfected by electroporation with an 18-mer MT antisense phosphorothioate oligomer (AO) or an 18-mer random oligomer (RO). The MT-AO is complementary to the region 7 bases downstream from the AUG translational start site of the hMT-IIA gene. Transfection of MCE7 cells with the AO inhibited cell growth by 50-60% at 72 hours when compared to control cells or the cells transfected with RO. The AO-induced growth inhibition was associated with alterations in morphology suggestive of apoptotic cell death. This was further confirmed by DNA linker cleavage into oligonucleosomal fragments and decreased bcl-2 protein levels in AO-transfected cells as opposed to the RO-transfected cells. Reverse transcriptase polymerase chain reaction analysis showed that AO induced a 2-fold increase in the levels of c-fos and p53 transcripts in comparison to RO which had no significant effect. Conversely, c-myc transcripts were decreased by 2.5-fold in the AO-transfected cells when compared to the controls. Furthermore, MCF7 cells transfected with an expression plasmid pBAcNEO-sMT-IIA encompassing human MT-IIA cDNA, constitutively driven by beta-actin promotor, caused a 2.5-fold increase in intracellular levels of MT, as judged by PCR and western blot analysis, in comparison to the cells transfected with pBAcNEO plasmid. In contrast to the AO-induced growth inhibition, overexpression of cytoplasmic MT increased the cell multiplication by 2-fold compared with control cells or the cells transfected with the control plasmid 72 hours post-transfection. Moreover, the effects of AO on oncogene expression were reversed on increased expression of MT. These data suggest that overexpression of MT potentiates the growth of MCF7 cells, whereas downregulation of MT elicits antiproliferative effects.
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PMID:Antisense down-regulation of metallothionein induces growth arrest and apoptosis in human breast carcinoma cells. 917 39

The present study was designed to determine whether changes in DNA methyltransferase (DNA MTase) expression are involved in hepatocarcinogenesis. We examined DNA MTase expression in normal liver tissue (with no remarkable histological findings), liver tissue showing chronic hepatitis or cirrhosis, which are generally thought to be precancerous conditions, and hepatocellular carcinomas (HCCs) using the reverse-transcriptase polymerase chain reaction assay. DNA MTase mRNA levels were significantly higher in liver tissue showing chronic hepatitis and cirrhosis (DNA MTase mRNA/beta-actin mRNA ratio = 0.30 +/- 0.22, n = 24, P < 0.01) than in normal liver tissue either from patients with liver metastatic lesions of colonic cancer (0.14 +/- 0.05, n = 6) or from patients with HCCs (0.16 +/- 0.07, n = 3). DNA MTase mRNA levels were even higher in HCC tissue (0.34 +/- 0.18, n = 29). These results suggest that increased DNA MTase expression may be an early event during hepatocarcinogenesis. DNA MTase is a potential target for HCC preventive therapy.
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PMID:Increased DNA methyltransferase expression is associated with an early stage of human hepatocarcinogenesis. 947 34

The experiments described herein were designed to determine whether tumor necrosis factor alpha (TNF-alpha) displays a diurnal variation in various areas of the normal rat brain. TNF-alpha mRNA transcripts were detected by reverse-transcriptase polymerase chain reaction. To monitor diurnal changes in TNF-alpha and alpha-tubulin expression, rats were sacrificed every 4 h for 24 h starting 1 h after light onset; relative mRNA levels were determined for the cerebellum, cortex, hippocampus, hypothalamus and brainstem. TNF-alpha mRNA was higher during the light than in the dark phase in the hypothalamus and hippocampus. alpha-Tubulin mRNA exhibited a similar diurnal variation in the hypothalamus, hippocampus and cortex. In contrast, beta-actin mRNA was lower during the light phase than the dark phase in the hippocampus and cortex. The observed diurnal variations in TNF-alpha mRNA are consistent with the hypothesis that TNF has a physiological role in the brain.
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PMID:Diurnal variations of tumor necrosis factor alpha mRNA and alpha-tubulin mRNA in rat brain. 948 99

Inflammatory cells were obtained from the spinal cords of rats with acute experimental autoimmune encephalomyelitis (EAE) induced by inoculation with myelin basic protein (MBP) and adjuvants. Reverse transcriptase-polymerase chain reaction (RT-PCR) was used to investigate the expression of mRNA for interleukin-2 (IL-2), IL-4, IL-10 and interferon-gamma (IFN-gamma) by cells from groups of rats studied 10-21 days after inoculation. On all days of study, the inflammatory cells, which were predominantly lymphocytes, expressed mRNA for IL-2, IL-4, IL-10 and IFN-gamma. In the mRNA from normal rat spinal cord tissue, there was little expression of cytokine mRNA. Cells from a short-term MBP-reactive T cell line expressed all the cytokines. Densitometry was used to measure the products of PCR, to assess the expression of each cytokine relative to that of beta-actin. IL-2 mRNA was expressed throughout the course of disease and reached a peak on day 18, during late clinical recovery. IFN-gamma was expressed throughout the course of the disease and was also high during late recovery. IL-4 mRNA was present in the spinal cord throughout the course of the disease, with a slight rise during late recovery. Relative expression of IL-10 rose to a peak on days 17-19, during late recovery from clinical disease. This study indicates that IL-2, IL-4, IL-10 and IFN-gamma are expressed by inflammatory cells in the spinal cord in EAE, with the relative expression of all cytokines being high during late clinical recovery.
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PMID:Cytokine expression by inflammatory cells obtained from the spinal cords of Lewis rats with experimental autoimmune encephalomyelitis induced by inoculation with myelin basic protein and adjuvants. 968 21

Total parenteral nutrition (TPN) may cause increased rates of bacterial translocation (BT), possibly due to a loss of epithelial integrity. Cultured epithelial cells have been shown to lose tight junction integrity with interferon gamma (INF-gamma) an action which may be blocked by transforming growth factor beta (TGF-beta). Because intraepithelial lymphocytes (IEL) are a rich source of these cytokines in the epithelium, we hypothesized that changes in the IEL, while mice were receiving TPN, may be responsible for the mediation of such cytokine responses. C57BL/6 mice were randomized to a Control group which received intravenous saline and mouse chow, or a TPN group which received intravenous TPN with no oral feeding. At 7 days mice were assessed for BT. Isolated IEL were stained for CD4, CD8, and CD44 (as a marker for memory T-cells) and flow cytometry was performed. mRNA was extracted from remaining IEL for cytokine expression. Reverse transcriptase polymerase chain reaction was performed to detect TGF-beta1 and INF-gamma mRNA expression. Densities were standardized to beta-actin expression. The incidence of BT to mesenteric lymph nodes was 40 and 12.5%, for the TPN and Control groups, respectively. TPN led to statistically significant decreases in the CD4+, CD8-; CD4+, CD8+; and the CD8+, CD44+ IEL subpopulations (P < 0.05). mRNA expression for INF-gamma was increased by 53% (P < 0.05), and TGF-beta1 mRNA expression was decreased by 75% (P = 0.1) in the IEL of TPN mice when compared with Controls. TPN led to significant changes in the IEL. Such alterations of the IEL phenotype and function may be a critical mechanism by which epithelial integrity is lost.
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PMID:Alteration of the intestinal intraepithelial lymphocytes during total parenteral nutrition. 975 21

The NaCl-reabsorbing collecting duct epithelium develops by budding and branching of the embryonic ureter. The expression of Na+ channels during this branching morphogenesis was studied in the outermost branches of rat ureteric buds (UB; embryonic day E15 to postnatal day P6) and in cortical collecting ducts (CCD; days P7-P28) in primary monolayer culture. Expression of both Na+ channel mRNA and of Na+-selective membrane conductance were estimated by quantitative reverse-transcriptase polymerase chain reaction (RT-PCR) and by patch-clamp recording, respectively. UB and CCD uniformly represented a principal-like cell type in culture. Messenger RNA encoding the alpha-ENaC subunit was detected in oligo-dT primed cDNA (5 ng) of embryonic UB cells (E15-17) after 30 PCR cycles. The abundance of alpha-ENaC mRNA, when normalized by reference to beta-actin, was higher by a factor of 2 in postnatal (P1-6) UB and by a factor of 5 in CCD cells (P7-14) compared with the embryonic stage. Highly Na+-selective, low-conductance channels were identified in apical patches from both UB and CCD monolayers, but only CCD cells exhibited macroscopic, amiloride-sensitive Na+ currents in whole-cell patch-clamp recordings. We conclude that alpha-ENaC mRNA and functional Na+ channel protein are expressed already before morphogenesis of the CCD is completed and prior to the onset of epithelial NaCl reabsorption.
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PMID:Expression of the epithelial sodium channel (ENaC) during ontogenic differentiation of the renal cortical collecting duct epithelium. 991 8

One of the underlying mechanisms of multidrug resistance (MDR) is cellular over-production of P-glycoprotein (P-gp), which acts as a drug efflux pump. P-gp is encoded by a small group of related genes termed MDR; only MDR1 is known to confer drug resistance. To overcome P-gp-mediated drug resistance, we have developed two anti-MDR1 hammerhead ribozymes driven by the beta-actin promoter. Upon transduction of the ribozymes into MDR cells, vincristine resistance was decreased. These two ribozymes were constructed, which showed different cleavage activities. In this study, to determine suitable target sites for the anti-MDR1 ribozyme, the exon 1b-intron 1 boundary, the translation-initiation site, the intron 1-exon 2 boundary and the exon 2-intron 2 boundary, codons 179 and 196 of the MDR1 gene were selected as candidates. To improve the ribozyme activity, a retroviral vector containing RNA polymerase III promoter was used. Stable retrovirus producer cells were generated by transfecting the retroviral vector plasmids carrying the ribozyme into the packaging cell line. Retroviral vector transduction of human leukemia cell lines expressing MDR1 was accomplished by co-culturing these with virus producer cells. Stably transduced cells were selected by G418 and pooled to determine the efficacy of each ribozyme. These ribozyme-transduced cells became vincristine-sensitive concomitant with the decreases in MDR1 expression, P-gp amount and drug efflux pump function. Among the ribozymes tested, the anti-MDR1 ribozyme against the translation-initiation site exhibited the strongest efficacy. This retrovirus-mediated transfer of anti-MDR1 ribozyme may be applicable to the treatment of MDR cells as a specific means to reverse resistance.
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PMID:Retrovirus-mediated transfer of anti-MDR1 hammerhead ribozymes into multidrug-resistant human leukemia cells: screening for effective target sites. 1036 43

This study was designed to determine whether changes in p27Kip1 expression are involved in human hepatocarcinogenesis. We examined the p27Kip1 mRNA expression in hepatocellular carcinomas and surrounding non-cancerous liver tissue samples from 21 patients using a reverse-transcriptase polymerase chain reaction assay. The mean p27Kip1 mRNA expression levels (ratio of p27Kip1/beta-actin mRNA) were significantly lower in hepatocellular carcinomas than in non-cancerous liver tissues (0.49 +/- 0.24 versus 0.57 +/- 0.22, P < 0.05). p27Kip1 mRNA expression was reduced in 11 (52%) of the 21 hepatocellular carcinomas when compared with the surrounding non-cancerous liver tissues. These findings suggest that reduced p27Kip1 expression may be at least partly responsible for human hepatocarcinogenesis.
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PMID:Reduced p27Kip1 expression in hepatocellular carcinomas. 1039 55

Oligosaccharide-plasmid DNA conjugates were synthesized simply and effectively via the diazocoupling method. Plasmids (pUC19, pTRI-beta-actin, and pEGFP-C1) were treated with an N-beta-lactoside-substituted diazonium salt to yield diazocoupling products with degree of substitutions of 2.5-3.1 mol% of overall nucleobases. The lactose-pUC19 conjugate was found to resist restriction enzymes more strongly than the nonconjugated plasmid DNA and to acquire a strong binding affinity to galactose-specific lectin RCA(120). The diazocoupling modification of pTRI-beta-actin plasmid DNA little influenced in vitro transcription with T7 RNA polymerase. When lactose-pEGFP-C1 conjugate was transfected to baby hamster kidney (BHK) cells by means of cationic lipids, transduced gene was expressed in BHK cells similarly with the nonconjugated pEGFP-C1. The modification of plasmid DNA with carbohydrate enhanced the resistance to restriction enzymes and developed a strong binding affinity to galactose-specific lectin.
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PMID:Conjugation of plasmid DNAs with lactose via diazocoupling enhances resistance to restriction enzymes and acquires binding affinity to galactose-specific lectin. 1040 69


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