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)

Transcriptional stimulation by the model activator GAL4-VP16 (a chimeric protein consisting of the DNA-binding domain of the yeast activator GAL4 and the acidic activation domain of the herpes simplex virus protein VP16) involves a series of poorly understood protein-protein interactions between the VP16 activation domain and components of the RNA polymerase II general transcription machinery. One of these interactions is the VP16-mediated binding and recruitment of transcription factor TFIIB. However, TATA box-binding protein (TBP)-associated factors (TAFs), or coactivators, are required for this interaction to culminate in productive transcription complex assembly, and one such TAF, Drosophila TAF40, reportedly forms a ternary complex with VP16 and TFIIB. Due to TFIIB's central role in gene activation, we sought to directly visualize the surfaces of this protein that mediate formation of the ternary complex. We developed an approach called protease footprinting in which the broad-specificity proteases chymotrypsin and alkaline protease were used to probe binding of 32P-end-labeled TFIIB to GAL4-VP16 or TAF40. Analysis of the cleavage products revealed two regions of TFIIB protected by VP16 from protease attack, one of which overlapped with a region protected by TAF40. The close proximity of the VP16 and TAF40 binding sites on the surface of TFIIB suggests that this region could act as a regulatory interface mediating the effects of activators and coactivators on transcription complex assembly.
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PMID:Protease footprinting reveals a surface on transcription factor TFIIB that serves as an interface for activators and coactivators. 759 78

Introduction of genes for cytokine receptors into hematopoietic stem/progenitor cells (HSC/HPC) may be of clinical use in the future. We recently reported that retroviral-mediated transduction of either the human erythropoietin receptor (hEpoR) or interleukin-9 receptor (hIL-9R) genes into highly purified HSC/HPC from cord blood (CB) resulted in increased numbers of detectable cytokine-responsive erythroid progenitors (burst-forming units-erythroid [BFU-E]). In the present study, we evaluated if this increase could be further enhanced by cotransducing both these genes into single isolated HSC/HPC. Single CD34++CD33-or low-expressing cells from CB were transduced with viral supernatant containing the hEpoR or hIL-9R genes or cotransduced with both genes. In the presence of Steel factor (SLF), interleukin-3 (IL-3), granulocyte-macrophage colony-stimulating factor (GM-CSF), erythropoietin (Epo), and IL-9, the numbers of erythroid colonies formed were significantly increased after transduction of cells with either the hIL-9R or hEpoR gene compared to mock-transduced cells. This increase was significantly enhanced in cells cotransduced with both genes compared with either gene alone. Integration and expression of both genes was confirmed by polymerase chain reaction (PCR) and reverse-transcriptase (RT)-PCR analysis, respectively. The data demonstrate that myeloid progenitors can be transduced at the single-cell level with both hEpoR and hIL-9R genes with resultant enhanced proliferation of these progenitors in the erythroid lineage by combinations of cytokines including Epo and IL-9.
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PMID:Influence of retroviral-mediated gene transduction of both the recombinant human erythropoietin receptor and interleukin-9 receptor genes into single CD34++CD33-or low cord blood cells on cytokine-stimulated erythroid colony formation. 864 64

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

Mammalian reovirus virions undergo partial disassembly of the outer capsid upon exposure to proteases in vitro, producing infectious subvirion particles (ISVPs) that lack protein sigma3 and contain protein mu1/mu1C as endoprotease-generated fragments mu1delta/delta and phi. ISVPs are thought to be required for two early steps in reovirus infection: membrane penetration and activation of the particle-bound viral transcriptase complexes. Genetic and biochemical evidence implicates outer-capsid protein mu1 in both these steps. To determine whether the cleavage of mu1/mu1C is relevant to the unique properties of ISVPs, we analyzed the properties of novel subvirion particles that lacked sigma3 yet retained mu1/mu1C in an uncleaved but cleavable form. These detergent-plus-protease subvirion particles (dpSVPs) were produced by treating virions with chymotrypsin in the presence of micelle-forming concentrations of alkyl sulfate detergents. Infections with dpSVPs in murine L or canine MDCK cells provided evidence that the cleavage of mu1/mu1C during viral entry into these cells is dispensable for reovirus infection. Additionally, dpSVPs behaved like ISVPs in their capacity to permeabilize lipid bilayers and to undergo transcriptase activation in vitro, supporting the conclusion that cleavage of mu1/mu1C to mu1delta/delta and phi during viral entry is not required for either membrane penetration or transcriptase activation in cells. The capacity of alkyl sulfate detergents to inhibit the cleavage of mu1/mu1C in a reversible fashion suggests a specific association between virus particle and detergent micelles that may mimic virus particle-phospholipid membrane interactions during reovirus entry into cells.
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PMID:Protease cleavage of reovirus capsid protein mu1/mu1C is blocked by alkyl sulfate detergents, yielding a new type of infectious subvirion particle. 942 Feb 47

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

The human ov-serpin monocyte neutrophil elastase inhibitor (MNEI) is encoded by a single gene SERPINB1. It is a highly efficient inhibitor of neutrophil granule proteases. Four murine genes with high sequence identity with MNEI were identified and fully sequenced, and these were named EIA, EIB, EIC, and EID. EIA, EIB and EIC showed the same seven-exon gene structure as SERPINB1. However, EIC included an additional, alternatively spliced, exon due to the insertion of an endogenous retrovirus-like sequence. EID lacked several exons and is a pseudogene. Reverse transcriptase-PCR showed that EIA, like MNEI, is expressed at high levels in many tissues. EIB is mainly expressed in brain, and EIC was only expressed as splicing variants unlikely to encode a functional serpin. Upon incubation with serine proteases, EIA formed inhibitory covalent complexes with pancreatic and neutrophil elastases, cathepsin G, proteinase-3, and chymotrypsin, as previously shown for MNEI, whereas EIB was only able to do so with cathepsin G. According to the new serpin nomenclature, the genes encoding EIA, EIB, EIC, and EID will be called Serpinb1, Serpinb1b, Serpinb1c, and Serpinb1-ps1. These data demonstrate that the four murine homologs of MNEI have met different evolutionary fates, and that EIA is the mouse ortholog of MNEI.
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PMID:Characterization of four murine homologs of the human ov-serpin monocyte neutrophil elastase inhibitor MNEI (SERPINB1). 1218 54

Histone modifications have been implicated in stem cell maintenance and differentiation. We have analyzed genome-wide changes in gene expression and histone modifications during differentiation of multipotent human primary hematopoietic stem cells/progenitor cells (HSCs/HPCs) into erythrocyte precursors. Our data indicate that H3K4me1, H3K9me1, and H3K27me1 associate with enhancers of differentiation genes prior to their activation and correlate with basal expression, suggesting that these monomethylations are involved in the maintenance of activation potential required for differentiation. In addition, although the majority of genes associated with both H3K4me3 and H3K27me3 in HSCs/HPCs become silent and lose H3K4me3 after differentiation, those that lose H3K27me3 and become activated after differentiation are associated with increased levels of H2A.Z, H3K4me1, H3K9me1, H4K20me1, and RNA polymerase II in HSCs/HPCs. Thus, our data suggest that gene expression changes during differentiation are programmed by chromatin modifications present at the HSC/HPC stage and provide a resource for enhancer and promoter identification.
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PMID:Chromatin signatures in multipotent human hematopoietic stem cells indicate the fate of bivalent genes during differentiation. 1912 86

Previous studies suggest that furanyl-rhodanines might specifically inhibit bacterial RNA polymerase (RNAP). We further explored three compounds from this class. Although they inhibited RNAP, each compound also inhibited malate dehydrogenase and chymotrypsin. Using biosensors responsive to inhibition of macromolecular synthesis and membrane damaging assays, we concluded that in bacteria, one compound inhibited DNA synthesis and another caused membrane damage. The third rhodanine lacked antibacterial activity. We consider furanyl-rhodanines to be unattractive RNAP inhibitor drug candidates.
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PMID:Furanyl-rhodanines are unattractive drug candidates for development as inhibitors of bacterial RNA polymerase. 2066 Jun 93

Somatic or de novo mutations of Additional sex combs-like 1 (ASXL1) frequently occur in patients with myeloid malignancies or Bohring-Opitz syndrome, respectively. We have reported that global loss of Asxl1 leads to the development of myeloid malignancies and impairs bone marrow stromal cell (BMSC) fates in mice. However, the impact of Asxl1 deletion in the BM niche on hematopoiesis remains unclear. Here, we showed that BMSCs derived from chronic myelomonocytic leukemia patients had reduced expression of ASXL1, which impaired the maintaining cord blood CD34+ cell colony-forming capacity with a myeloid differentiation bias. Furthermore, Asxl1 deletion in the mouse BMSCs altered hematopoietic stem and progenitor cell (HSC/HPC) pool and a preferential myeloid lineage increment. Immunoprecipitation and ChIP-seq analyses demonstrated a novel interaction of ASXL1 with the core subunits of RNA polymerase II (RNAPII) complex. Convergent analyses of RNA-seq and ChIP-seq data revealed that loss of Asxl1 deregulated RNAPII transcriptional function and altered the expression of genes critical for HSC/HPC maintenance, such as Vcam1. Altogether, our study provides a mechanistic insight into the function of ASXL1 in the niche to maintain normal hematopoiesis; and ASXL1 alteration in, at least, a subset of the niche cells induces myeloid differentiation bias, thus, contributes the progression of myeloid malignancies.
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PMID:Loss of ASXL1 in the bone marrow niche dysregulates hematopoietic stem and progenitor cell fates. 2942 72

The mainstream approach to antiviral drugs against COVID-19 is to focus on key stages of the SARS-CoV-2 life cycle. The vast majority of candidates under investigation are repurposed from agents of other indications. Understanding of protein-inhibitor interactions at molecular scale will provide crucial insights for drug discovery to stop this pandemic. In this article, we summarize and analyze the most recent structural data on several viral targets with the presence of promising inhibitors for COVID-19 in the perspective of modes of action (MOA) to unravel insightful mechanistic features with atomistic resolution. The targets include spike glycoprotein and various host proteases mediating the entry of the virus into the cells, viral chymotrypsin- and papain-like proteases, and RNA dependent RNA polymerase. The main purpose of this review is to present detailed MOA analysis to inspire fresh ideas for both de novo drug design and optimization of known scaffolds to combat COVID-19.
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PMID:Molecular Insights into Small Molecule Drug Discovery for SARS-CoV-2. 3272 80


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