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)

Genes of higher eucaryotic cells are considered to show only a limited response to nutritional stress. Here we show, however, that omission of a single essential amino acid from the medium caused a marked rise in the mRNA levels of c-myc, c-jun, junB and c-fos oncogenes and ornithine decarboxylase (ODC) in CHO cells. There was no general accumulation of mRNAs in amino acid-starved cells, since the gamma-actin, beta-tubulin, protein kinase C, RNA polymerase II, and glyceraldehyde-3-phosphate dehydrogenase mRNAs and the total poly(A)+ mRNA were not increased. The levels of c-myc, ODC, and c-jun mRNAs were elevated more by amino acid starvation than by inhibition of protein synthesis with cycloheximide, which is known to increase the levels of these mRNAs. Importantly, however, cycloheximide present during amino acid starvation reduced the rise in the levels of the mRNAs down to the level obtained with cycloheximide alone. This implies that protein synthesis is required for the accumulation of c-myc, ODC, and c-jun mRNAs in amino acid-deprived cells. The junB and c-fos mRNAs, instead, were increased to the same extent or less by amino acid starvation than by cycloheximide treatment. The accumulation of the c-myc mRNA in amino acid-starved cells was due to both stabilization of the mRNA and increase of its transcription. The rise in the c-jun mRNA level seemed to be caused merely by stabilization of the mRNA. Further, despite the inhibition of general protein synthesis, amino acid starvation led to an increase in the synthesis of c-myc polypeptide. The results suggest that mammalian cells have a specific mechanism for registering shortages of amino acids in order to make adjustments compatible with cellular growth.
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PMID:Deprivation of a single amino acid induces protein synthesis-dependent increases in c-jun, c-myc, and ornithine decarboxylase mRNAs in Chinese hamster ovary cells. 212 33

A single-stranded DNA-binding protein of Mr 35,000 (35K protein) was isolated from calf cerebral cortex by affinity chromatography on immobilized double-stranded and single-stranded DNA. Its localization in the nuclear compartment was demonstrated by immunohistochemistry. Previous studies had uncovered a homologous nonhistone chromosomal protein in the nuclei of rat cerebral cortex neurons, cerebellar neurons, oligodendrocytes, and liver cells. The rat protein accumulated in the nuclear compartment of neurons in exact temporal coincidence with the arrest of cell division and the initiation of terminal differentiation. Therefore, in the present work, the 35K protein was tested for an activating role in RNA transcription. During the course of this study we became aware that the 35K protein was identical to a glycolytic enzyme, glyceraldehyde-3-phosphate dehydrogenase (GAPDH, EC 1.2.1.12). When authentic GAPDH from rabbit skeletal muscle was injected into Xenopus laevis oocytes, it greatly stimulated RNA polymerase II transcription, whereas the 35K protein from calf brain did not. This apparent discrepancy was partially resolved by the finding that rabbit muscle GAPDH could be fractionated into two components by affinity chromatography on single-stranded DNA cellulose. Only 5% of the applied protein was retained on the column and could be eluted with a shallow salt gradient identical to the one used for the isolation of the 35K protein. This single-stranded DNA-binding component of rabbit muscle GAPDH did not stimulate transcription. Apparently, the 35K protein from calf brain corresponded to this single-stranded DNA-binding subfraction, which explained its failure to activate transcription.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Glyceraldehyde-3-phosphate dehydrogenase is a nonhistone protein and a possible activator of transcription in neurons. 242 47

Two Drosophila genes that code for the enzyme glyceraldehyde-3-phosphate dehydrogenase (Gapdh) have been isolated and their structures determined by DNA sequence analysis. The two genes, Gadph-1 and Gapdh-2, are homologous to each other in their coding regions but differ entirely in the 5' and 3' flanking regions. Both genes are functionally expressed in adult flies as determined by Northern blot analysis using gene-specific probes. Gapdh-1 is mapped by in situ hybridization at position 43E-F on the right arm of the second chromosome and Gapdh-2 at position 13F on the left arm of the X chromosome. Transcription initiation sites as well as polyadenylation sites for both Gapdh transcripts have also been determined. Gapdh-1 lacks a sequence homologous to the TATA box in its -30-base pair region that is characteristic of many RNA polymerase II transcribed promoters. In contrast, Gapdh-2 contains a consensus TATA box sequence as well as a CAAT box in its promoter region. Furthermore, a sequence element ATTTGCAT (dc) and nontandem multiple direct repeats have been found in the -35 to -155-base pair 5' flanking region. Other than the intron located in the 5' noncoding region of Gapdh-2, both genes lack intervening sequences.
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PMID:Structure of two unlinked Drosophila melanogaster glyceraldehyde-3-phosphate dehydrogenase genes. 298 82

Collagen deposition and myofibroblast proliferation beneath the epithelial basement membrane in patients with asthma is now increasingly recognized, although the molecular pathogenesis remains obscure. We have evaluated messenger ribonucleic acid (mRNA) expression of the profibrotic cytokine, platelet-derived growth factor-beta (PDGF-beta), in alveolar macrophages obtained following fibreoptic bronchoscopy and bronchoalveolar lavage in patients with asthma. Three subject groups were studied: 1) asthmatics using regular inhaled glucocorticoid medication (ASTST, n = 9), 2) asthmatics using intermittent inhaled beta 2-agonist therapy only (ASTBR, n = 10); 3) nonasthmatic control volunteers (n = 10). Alveolar macrophage mRNA was extracted and PDGF-beta mRNA quantified by reverse-transcriptase polymerase chain reaction (PCR) (RT-PCR) and expressed as the ratio to that of a control gene glyceraldehyde-3-phosphate dehydrogenase (GAPDH). There were no significant differences in PDGF-beta mRNA expression between the groups, or between all asthmatic (n = 19) and control subjects. Furthermore, there was no correlation between alveolar macrophage PDGF-beta mRNA expression and airway spirometry, or duration of glucocorticoid usage or dose. Thus, in contrast to other fibrotic lung diseases, we found little evidence of enhanced expression of PDGF-beta mRNA in alveolar macrophages in clinically stable bronchial asthma.
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PMID:Platelet-derived growth factor-beta mRNA in human alveolar macrophages in vivo in asthma. 787 66

Escherichia coli D-glyceraldehyde-3-phosphate dehydrogenase (GAPDH) is produced by the gapA gene and is structurally related to eukaryotic GAPDHs. These facts led to the proposal that the gapA gene originated by a horizontal transfer of genetic information. The yields and start sites of gapA mRNAs produced in various fermentation conditions and genetic contexts were analyzed by primer extension. The transcriptional regulatory region of the gapA gene was found to contain four promoter sequences, three recognized by the vegetative RNA polymerase E sigma 70 and one recognized by the heat shock RNA polymerase E sigma 32. Transcription of gapA by E sigma 32 is activated in the logarithmic phase under conditions of starvation and of heat shock. Using a GAPDH- strain, we found that GAPDH production has a positive effect on cell growth at 43 degrees C. Thus, E. coli GAPDH displays some features of heat shock proteins. One of the gapA promoter sequences transcribed by E sigma 70 is subject to catabolic repression. Another one has growth phase-dependent efficiency. This complex area of differentially regulated promoters allows the production of large amounts of gapA transcripts in a wide variety of environmental conditions. On the basis of these data, the present view of E sigma 32 RNA polymerase function has to be enlarged, and the various hypotheses on E. coli gapA gene origin have to be reexamined.
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PMID:The Escherichia coli gapA gene is transcribed by the vegetative RNA polymerase holoenzyme E sigma 70 and by the heat shock RNA polymerase E sigma 32. 830 May 36

The effect of vitamin B6 deficiency on the activity of RNA polymerase and expression of several mRNAs in rat liver was investigated. The activities of RNA polymerase I and II in the liver of vitamin B6-deficient rats were found to be higher than the control rats by 30%. The expression of several mRNAs, including mRNAs for beta-actin and glyceraldehyde-3-phosphate dehydrogenase, and the content of poly(A)+ RNA were also increased in vitamin deficiency. These observations suggest that vitamin B6 influences gene expression in the liver, at least in part, by modulating the activity of RNA polymerase.
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PMID:Vitamin B6 deficiency causes activation of RNA polymerase and general enhancement of gene expression in rat liver. 840 98

AMP nucleosidase (EC 3.2.2.4) from Escherichia coli and AMP deaminase (EC 3.5.4.6) from bakers' yeast are proposed to regulate cellular AMP levels under allosteric control of the activator ATP and the inhibitor, PO4. Both enzymes contain catalytic sites which bind AMP and regulatory sites which bind ATP. The deduced amino acid sequences of the proteins revealed only one region of homology in which six of eight amino acids are identical. A similar sequence is found in glyceraldehyde-3-phosphate dehydrogenase, phoE, ras proteins, RNA polymerase, K(+)-ATPase, nucleolin, and other proteins expected to have nucleotide or phosphate binding properties. In the crystal structure of glyceraldehyde-3-phosphate dehydrogenase, this sequence is part of the NAD(+)-binding site. The function of these amino acids was explored with a deletion mutant of AMP nucleosidase. The protein was over-produced in a pTZ construct using the AMP nucleosidase promoter which resulted in approximately 30% of the total protein as the desired enzyme. The mutation was characterized by DNA sequence analysis and by direct analysis of the peptides using high performance liquid chromatography-mass spectrometry. Deletion of amino acids 128-135, corresponding to DGSELTLD, produced an enzyme with a 20-fold decrease in Vmax but with smaller changes in substrate saturation kinetics, activation by MgATP, inhibition by inorganic phosphate, and inhibition by the tight-binding inhibitor, formycin 5-phosphate. The deletion mutant of AMP nucleosidase exhibits hysteresis in establishing a steady-state rate of product formation which is most pronounced in the absence of MgATP. These results establish that the sequence DGSELTLD in E. coli AMP nucleosidase is not required for binding of AMP, MgATP, or inorganic phosphate. However, the mutant enzyme has a structural defect related to the polymerization state which delays the onset of catalysis and decreases the catalytic efficiency.
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PMID:Mutagenic analysis of AMP nucleosidase from Escherichia coli. Deletion of a region similar to AMP deaminase and peptide characterization by mass spectrometry. 847 16

1. Induction of nitric oxide synthase (iNOS) results in overproduction of nitric oxide (NO), which may be a principal cause of the massive vasodilatation and hypotension observed in septic shock. Since NO-induced vasorelaxation is mediated via the soluble isoform of guanylate cyclase (sGC), the regulation of sGC activity during shock is of obvious importance, but yet poorly understood. The aim of the present study was to investigate the activation of sGC by sodium nitroprusside (SNP) before and after exposure of rat aortic smooth muscle cells to endotoxin (LPS) or interleukin-1 beta (IL-1 beta). 2. Exposure of rat aortic smooth muscle cells to SNP (10 microM) elicited up to 200 fold increases in cyclic GMP. This effect was attenuated by 30-70% in IL-1 beta- or LPS-pretreated cells, in a pretreatment time-and IL-1 beta- or LPS-concentration-dependent manner. When, however, cells were exposed to IL-1 beta or LPS and then stimulated with the particulate guanylate cyclase activator, atriopeptin II, no reduction in cyclic GMP accumulation was observed. 3. Pretreatment of rats with LPS (5 mg kg-1, i.v.) for 6 h led to a decrease in aortic ring SNP-induced cyclic GMP accumulation. 4. The IL-1 beta-induced reduction in SNP-stimulated cyclic GMP accumulation in cultured cells was dependent on NO production, as arginine depletion abolished the downregulation of cyclic GMP accumulation in response to SNP. 5. Reverse-transcriptase-polymerase chain reaction analysis revealed that the ratio of steady state mRNA for the alpha, subunit of sGC to glyceraldehyde phosphate dehydrogenase was decreased in LPS- or IL-1 beta-treated cells, as compared to vehicle-treated cells. 6. Protein levels of the alpha 1 sGC subunit remained unaltered upon exposure to LPS or IL-1 beta, suggesting that the early decreased cyclic GMP accumulation in IL-1 beta- or LPS-pretreated cells was probably due to reduced sGC activation. Thus, the observed decreased responsiveness of sGC to NO stimulation following cytokine or LPS challenge may represent an important homeostatic mechanism to offset the extensive vasodilatation seen in sepsis.
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PMID:Downregulation of nitrovasodilator-induced cyclic GMP accumulation in cells exposed to endotoxin or interleukin-1 beta. 883 57

It has been long known that neoplastic transformation is accompanied by a lowered requirement for extracellular Ca2+ for growth. The studies presented here demonstrate that human fibroblastic cell lines produce the two commonly found 'housekeeping' isoforms of the plasma membrane Ca(2+)-ATPase (PMCA), PMCA1b and 4b, and at the expression of both is demonstrably lower in cell lines neoplastically transformed by SV40 than in the corresponding parental cell lines. Western blot analyses of lysates from control (GM00037) and SV40-transformed (GM00637) skin fibroblasts revealed a 138 kDa PMCA whose level was significantly lower in the SV40-transformed cells relative to either total cellular protein or alpha-tubulin. Similar analyses of plasma membrane preparations from control WI-38) and SV40-transformed (WI-38VA13) lung fibroblasts revealed 3-4-fold lower levels of PMCA in the SV40-transformed cells. Competitive ELISAs performed on detergent solubilized plasma membrane preparations indicated at least 3-4-fold lower levels of PMCA in the SV40-transformed cell lines compared to controls. Reverse transcriptase coupled-PCR analyses showed that PMCA1b and PMCA4b were the only isoforms expressed in all four cell lines. The PMCA4b mRNA level detected by Northern analysis also was substantially lower in SV40 transformed skin fibroblasts than in non-transformed fibroblasts. Quantitative RT-PCR analyses showed levels of PMCA1b and 4b mRNAs to be 5 and 10-fold lower, respectively, in GM00637 than in GM00037 when the levels of PCR products were normalized to glyceraldehyde-3-phosphate dehydrogenase (G3PDH) mRNA. These results demonstrate that the expression of these distinct PMCA genes is substantially lower in SV40 transformed human skin and lung fibroblasts and may be coordinately regulated in these cells.
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PMID:Analysis of plasma membrane Ca(2+)-ATPase expression in control and SV40-transformed human fibroblasts. 905 77

Spontaneous tumor regression, which is observed clinically and histologically in some primary melanomas, occurs in the absence of any effective therapy. It is probably immunologically mediated, because regressing melanomas are infiltrated with larger numbers of activated T cells, primarily CD4+, than nonregressing melanomas. To investigate the hypothesis that spontaneous regression of melanomas is caused by T-cell cytokine production, cytokine mRNA expression in 20 primary melanomas was examined using a noncompetitive, quantitative reverse-transcriptase polymerase chain reaction method. DNA standards were used to generate known numbers of molecules in each sample. Results were standardized to the internal control, glyceraldehyde-3-phosphate dehydrogenase. mRNA for CD35, lymphotoxin (TNF-beta), and IL-2 were significantly elevated in the ten regressing melanomas compared to the ten nonregressing melanomas. IFN-gamma mRNA was also elevated in regressing melanomas but failed to reach statistical significance. The Th2 cytokines IL-10 and IL-13 did not show differences in the regressing melanomas compared to nonregressing melanomas; neither did the pro-inflammatory cytokines IL-1alpha, IL-1beta, IL-6, IL-8, and TNF-alpha, nor the growth factors, bFGF and TGF-beta or GM-CSF. This study shows an association between Th1 cytokines and spontaneously regressing melanomas. Although we have not shown that these cytokines cause regression, these findings support our hypothesis that activated CD4+ T cells may mediate melanoma regression by secretion of Th1 cytokines.
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PMID:T helper 1 cytokine mRNA is increased in spontaneously regressing primary melanomas. 918 21


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