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)

Pancreatic digestive enzymes have rarely been reported in human nonpancreatic organs. We examined their expression in the epithelial cells of the nonpancreatic gastrointestinal organs, looking for pancreatic alpha-amylase, trypsin, chymotrypsin and pancreatic lipase. Western blotting, enzyme assay and pancreatic alpha-amylase mRNA were also used in selected specimens. In normal tissues, immunoreactivity of one or more of these enzymes was frequently noted in cells of the salivary glands, stomach, duodenum, large pancreatic ducts, extrahepatic bile ducts and gall bladder. The epithelium of the normal oesophagus, small intestine and colon were consistently negative for these enzymes. In pathologic tissues, immunoreactivity for one or more enzymes was present in epithelial cells of pleomorphic adenomas of the salivary glands, oesophageal squamous cell carcinoma, gastric adenoma and adenocarcinoma, pancreatic adenocarcinoma, cholecystitis, adenocarcinoma of the gall bladder and extrahepatic bile duct, and colon adenoma and adenocarcinoma. Western blotting showed a specific band of each enzyme in some specimens of normal stomach. In situ hybridization for pancreatic alpha-amylase mRNA showed specific signals in the normal stomach, but not in the normal colon. Reverse transcriptase polymerase chain reaction analysis for pancreatic alpha-amylase mRNA revealed specific signals in the normal stomach. Enzyme assay revealed that the stomach and gall bladder showed these activities. The data suggest that pancreatic digestive enzymes are produced by several epithelial cell types of the nonpancreatic gastrointestinal organs, that the organs positive for pancreatic enzyme have a common cell lineage, and that neoplasms continue to express or neoexpress these enzymes after neoplastic transformation.
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PMID:Expression of pancreatic digestive enzymes in normal and pathologic epithelial cells of the human gastrointestinal system. 933 41

The GC-rich segment containing GGAGGC (Alu core) is conserved within the RNA polymerase III (pol III) promoters of Alu family sequences. We have shown that the GGAGGC motif functions as a modulator of DNA replication as well as of transcription, and identified the proteins binding to the motif in human HeLa cells. In this study, the Alu core binding proteins were partially purified from human Raji cells by using an Alu core DNA affinity column. Both the proteins thus purified were implied to be subunits of Ku antigen based on the following criteria: The molecular weights of the proteins estimated on gel electrophoreses were 70 and 85 kDa, respectively, under denaturing conditions, while under non-denaturing conditions only one band was observed for the same sample at 150 kDa, probably representing hetero-dimer formed between the 70 and 85 kDa proteins. The sizes and the hetero-dimer formation are reminiscent of the 70 and 80 kDa subunits of Ku antigen (Ku-p70 and Ku-p80). Moreover, the purified proteins were immunoreactive with anti-Ku antibodies, and the specific DNA-protein complex on the Alu core element was cancelled by the anti-Ku antibodies. The nucleoprotein complex showed the same clipping patterns as those of the complex between the Alu core element and an authentically purified Ku antigen after proteolytic cleavage with trypsin and chymotrypsin.
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PMID:Ku antigen binds to Alu family DNA. 950 18

Histone-DNA templates for bacteriophage T7 RNA polymerase were assembled from histone octamers and three different DNA species, two circular (pGEMEX-1 and pT207-18) and one linear (T7-207-18). pGEMEX is devoid of nucleosome positioning sequences, while in pT207-18 and T7-207-18 the region downstream of the promoter contains 18 tandem repeats of a 207 bp positioning sequence derived from the 5S RNA gene of the sea urchin Lytechinus variegatus. Elimination of the histone tails in the assembled oligonucleosomes by trypsin digestion is accompanied, in all three DNA species, by substantial increases in transcription efficiency, assayed at different KCl and MgCl2 concentrations, after allowing for the aggregation observed under certain conditions. In the absence of KCl and at low MgCl2 concentration, the presence of 2 mM spermidine causes substantial aggregation of the intact oligonucleosomes but has a much smaller effect on those trypsin digested. The untreated histone-DNA templates, assembled on pGEMEX-1 and T7-207-18, give transcription products significantly shorter than those obtained with the corresponding free DNA. With oligonucleosome templates lacking histone tails, the transcripts have an average length intermediate between those corresponding to free DNA and intact histone-DNA, which indicates a partial elimination of the elongation restrictions imposed by intact histone octamers. The absence of histone terminal domains facilitates both transcriptional initiation and elongation. Apparently, the interaction of the histone tails with DNA at the nucleosomal level is responsible, at least in part, for their repressive effect on transcription.
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PMID:Repressive effect on oligonucleosome transcription of the core histone tail domains. 958 38

T7 RNA polymerase shows an increase in processive transcription in the presence of low concentrations of guanidine hydrochloride (GdnCl) upto 60 mM, which is not observed when the enzyme is treated with urea. Higher concentrations of the denaturant lead to a progressive loss in the processive transcriptional activity of the enzyme. We have attempted to explain the above phenomenon in terms of the structural change in the enzyme. Fluorescence and CD studies suggest that the tertiary structure of the native enzyme undergoes an alteration upon addition of low concentration of guanidine hydrochloride. This is also indicated from the decreased susceptibility of the enzyme to limited proteolysis by trypsin.
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PMID:Enhancement of transcriptional activity of T7 RNA polymerase by guanidine hydrochloride. 963 52

Histone-DNA templates for bacteriophage T7 RNA polymerase were assembled from a plasmid containing a promoter and a terminator for this polymerase, (H3 x H4)2 tetramers deprived of their tail domains, and H2A x H2B dimers. Histone (H3 x H4)2 tetramers lacking their terminal domains were obtained from trypsin-digested nucleosomal cores. The oligonucleosomal templates containing (H3 x H4)2 tetramers lacking their tail domains, like the control templates with intact core histone octamers, protect approximately 146 base pairs of DNA against micrococcal nuclease digestion. The transcriptional inhibition caused by the association of DNA with core histone octamers is significantly reduced upon elimination of the tail domains of the (H3 x H4)2 tetramers. Apparently, the terminal domains of (H3 x H4)2 must be present to block transcription efficiently. These results show the important inhibitory role played by the tail domains of the histone (H3 x H4)2 tetramers, suggesting the involvement of these regions in transcriptional regulation.
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PMID:Transcriptional inhibitory role of the tail domains of histone (H3 x H4)2 tetramers. 975 Jan 70

Numerous studies suggest that C-ANCA are directly pathogenic in vasculitis by activating leucocytes (oxidative burst, enzyme release, endothelial cytotoxicity, etc.). We and others have shown that C-ANCA can also directly activate HUVEC, but the precise target on HUVEC is unknown. We show in this study that C-ANCA recognize various targets on the HUVEC membrane (different from PR3 in our model), leading to secondary cell activation. Polyclonal affinity-purified C-ANCA recognized targets on the unfixed endothelial membrane in fluorescent ELISA, flow cytometry, and immunoprecipitation studies. C-ANCA did not react with Fcgamma receptors. Reverse transcriptase-polymerase chain reaction (RT-PCR) experiments showed that HUVEC did not express PR3. The targets of polyclonal and monoclonal anti-PR3 antibodies on the endothelial membrane were not the same. Some epitopes were lost after trypsin-EDTA digestion and formaldehyde fixation of cells, whereas anti-PR3 targeted unfixed HUVEC. This suggests that anti-PR3 react with the endothelial membrane and recognize conformational epitopes shared with PR3. Endothelial cells may thus participate in the inflammation associated with Wegener's granulomatosis and contribute to the emergence of clinical manifestations.
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PMID:Anti-proteinase-3 (PR3) antibodies (C-ANCA) recognize various targets on the human umbilical vein endothelial cell (HUVEC) membrane. 993 66

Osteoblasts express protease-activated receptor-1 (PAR-1), which is activated by thrombin or by synthetic peptides corresponding to the new "tethered ligand" N-terminus of PAR-1 created by receptor cleavage. Both thrombin and human PAR-1-activating peptide stimulate an elevation of [Ca2+]i in the human SaOS-2 osteoblast-like cell line, but the peptide stimulates receptor-mediated Ca+ entry, whereas thrombin does not. Stimulation of proliferation in rat primary osteoblast-like cells is greater in response to rat PAR-1-activating peptide than to thrombin. Because the PAR-1-activating peptides are now known to activate PAR-2, the current study was undertaken to investigate whether osteoblasts express this receptor and, if so, whether this could account for the observed discrepancies between responses of osteoblasts to thrombin and to PAR-1-activating peptides. Reverse transcriptase-polymerase chain reaction (RT-PCR) and immunocytochemical studies demonstrated expression of PAR-2 by primary cultures of rat calvarial osteoblast-like cells. In immunohistochemical studies of embryonic mouse bones, osteoblasts showed positive staining for the presence of PAR-2. Activators of PAR-2 include trypsin, mast cell tryptase, gingipain-R, and synthetic peptides corresponding to the PAR-2 tethered ligand sequence. Treatment of primary rat osteoblast-like cells with rat PAR-2-activating peptide (SLIGRL), or SaOS-2 cells with human PAR-2-activating peptide (SLIGKV), caused a dose-dependent increase in [Ca2+]i. Trypsin or gingipain-R also induced an increase in intracellular calcium concentration, and caused reciprocal cross desensitization. Activators of PAR-2 caused a sharp peak in [Ca2+]i followed by a sustained plateau; [Ca2+]i returned to baseline levels upon treatment with ethylene-glycol tetraacetic acid (EGTA). Treatment of rat osteoblast-like cells in vitro with SLIGRL did not affect thymidine incorporation or endogenous alkaline phosphatase activity. The results presented here demonstrate that osteoblasts express PAR-2, and that such expression is able to account for the observed discrepancies between thrombin and PAR-1-activating peptides in their ability to evoke calcium entry, but not proliferative responses.
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PMID:Expression of protease-activated receptor-2 by osteoblasts. 1061 51

Protease-activated receptor-2 (PAR-2) is distributed throughout the gastrointestinal systems. The present study investigated the role for PAR-2 in the rat salivary glands. PAR-2 mRNA was detected in the sublingual, submaxillary, and parotid glands by a reverse-transcriptase polymerase chain reaction. In the isolated sublingual gland that exhibited the strongest signal for PAR-2, Ser-Leu-Ile-Gly-Arg-Leu-NH(2), a PAR-2-activating peptide, and trypsin, a PAR-2-activating enzyme, but not thrombin that can activate PARs 1, 3, and 4, triggered secretion of N-acetylneuraminic acid, an indicator of mucin, that was a unique major sialic acid detectable after hydrolysis of the sublingual mucin with 0.1 N HCl. The PAR-2-mediated secretion of mucin was attenuated by genistein, a tyrosine kinase inhibitor, but not by inhibitors of protein kinase C and phosphatidyl inositol 3'-kinase. Thus, PAR-2 is expressed by the three distinct salivary glands in the rat, and sublingual PAR-2 appears to play a role in triggering mucin secretion, at least in part, via activation of tyrosine kinase.
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PMID:Activation of protease-activated receptor-2 (PAR-2) triggers mucin secretion in the rat sublingual gland. 1073 43

The developmental regulatory protein sigma(F) of Bacillus subtilis, a member of the sigma(70)-family of bacterial RNA polymerase sigma factors, is negatively regulated by the anti-sigma factor SpoIIAB, which binds to sigma(F), sequestering it in an inactive complex. SpoIIAB binding to sigma(F) is strongly stimulated by ATP. Here, we use a combination of gel filtration chromatography, dynamic light-scattering, analytical ultracentrifugation, limited proteolysis with N-terminal sequencing and electrospray mass spectrometry, and deletion analysis to probe the SpoIIAB-sigma(F) complex. The studies were facilitated by investigating the homologs from Bacillus stearothermophilus as well as co-expression of the proteins in Escherichia coli, allowing purification of large quantities of the in vivo assembled complex. We determined the stoichiometry of the complex to be SpoIIAB(2):sigma(F)(1). Alone, sigma(F) is rapidly degraded by the protease trypsin. In the complex with SpoIIAB, however, sigma(F) is remarkably resistant to proteolysis. Analysis of the protease cleavage data indicates the anti-sigma binds sigma(F) through contacts with mutliple conserved regions of the sigma factor, supporting previous findings based on genetic data.
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PMID:The anti-sigma factor SpoIIAB forms a 2:1 complex with sigma(F), contacting multiple conserved regions of the sigma factor. 1086 95

We have recovered a virion from defective cDNA of Sendai virus (SeV) that is capable of self-replication but incapable of transmissible-virion production. This virion delivers and expresses foreign genes in infected cells, and this is the first report of a gene expression vector derived from a defective viral genome of the Paramyxoviridae. First, functional ribonucleoprotein complexes (RNPs) were recovered from SeV cloned cDNA defective in the F (envelope fusion protein) gene, in the presence of plasmids expressing nucleocapsid protein and viral RNA polymerase. Then the RNPs were transfected to the cells inducibly expressing F protein. Virion-like particles thus obtained had a titer of 0.5 x 10(8) to 1. 0 x 10(8) cell infectious units/ml and contained F-defective RNA genome. This defective vector amplified specifically in an F-expressing packaging cell line in a trypsin-dependent manner but did not spread to F-nonexpressing cells. This vector infected and expressed an enhanced green fluorescent protein reporter gene in various types of animal and human cells, including nondividing cells, with high efficiency. These results suggest that this vector has great potential for use in human gene therapy and vaccine delivery systems.
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PMID:A cytoplasmic RNA vector derived from nontransmissible Sendai virus with efficient gene transfer and expression. 1086 70


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