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)

Torsional tension in DNA may be both a prerequisite for the efficient initiation of transcription and a consequence of the transcription process itself with the generation of positive torsional tension in front of the RNA polymerase and negative torsional tension behind it. To examine torsional tension in specific regions of genomic DNA in vivo, we developed an assay using photoactivated psoralen as a probe for unconstrained DNA superhelicity and x-rays as a means to relax DNA. Psoralen intercalates more readily into DNA underwound by negative torsional tension than into relaxed. DNA, and it can form interstrand DNA cross-links upon UVA irradiation. By comparing the amount of psoralen-induced DNA cross-links in cells irradiated with x-rays either before or after the psoralen treatment, we examined the topological state of the DNA in specific regions of the genome in cultured human 6A3 cells. We found that although no net torsional tension was detected in the bulk of the genome, localized tension was prominent in the DNA of two active genes. Negative torsional tension was found in the 5' end of the amplified dihydrofolate reductase gene and in a region near the 5' end of the 45S rRNA transcription unit, whereas a low level of positive torsional tension was found in a region near the 3' end of the dihydrofolate reductase gene. These results document an intragenomic heterogeneity of DNA torsional tension and lend support to the twin supercoiled domain model for transcription in the genome of intact human cells.
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PMID:Localized torsional tension in the DNA of human cells. 163 Oct 91

We have studied the chromatin structure of the Saccharomyces cerevisiae FBP1 gene, which codes for fructose-1,6-bisphosphatase. A strong, constitutive, DNase I, micrococcal nuclease and S1 nuclease hypersensitive site is present close to the 3' end of the coding region. In the repressed state, positioned nucleosomes exist around this site, and subtle changes occur in this nucleosomal organization upon derepression. A DNase I hypersensitive region is located within the promoter between positions -540 and -400 and its extends towards the gene in the derepressed state, leading to an alteration of nucleosomal positioning. Psoralen crosslinking of chromatin, which is used for the first time to study the mobility of restriction fragments from an RNA polymerase II gene, revealed that part of the promoter is nucleosome-free, in accordance with the results of DNase I digestion. A model is presented that, based on the chromatin structure, puts forward the hypothesis that the promoter UAS is located between -540 and -340. Finally, psoralen crosslinking, as well as digestions with micrococcal nuclease or restriction endonucleases suggests that most if not all of the copies of the active FBP1 gene are covered by nucleosomes.
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PMID:Chromatin structure of the yeast FBP1 gene: transcription-dependent changes in the regulatory and coding regions. 810 72

To explore the ability of triplex-forming oligodeoxyribonucleotides (TFOs) to inhibit genes responsible for dominant genetic disorders, we used two TFOs to block expression of the human rhodopsin gene, which encodes a G protein-coupled receptor involved in the blinding disorder autosomal dominant retinitis pigmentosa. Psoralen-modified TFOs and UVA irradiation were used to form photoadducts at two target sites in a plasmid expressing a rhodopsin-EGFP fusion, which was then transfected into HT1080 cells. Each TFO reduced rhodopsin-GFP expression by 70-80%, whereas treatment with both reduced expression by 90%. Expression levels of control genes on either the same plasmid or one co-transfected were not affected by the treatment. Mutations at one TFO target eliminated its effect on transcription, without diminishing inhibition by the other TFO. Northern blots indicated that TFO-directed psoralen photoadducts blocked progression of RNA polymerase, resulting in truncated transcripts. Inhibition of gene expression was not relieved over a 72 h period, suggesting that TFO-induced psoralen lesions are not repaired on this time scale. Irradiation of cells after transfection with plasmid and psoralen-TFOs produced photoadducts inside the cells and also inhibited expression of rhodopsin-EGFP. We conclude that directing DNA damage with psoralen-TFOs is an efficient and specific means for blocking transcription from the human rhodopsin gene.
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PMID:Blocking transcription of the human rhodopsin gene by triplex-mediated DNA photocrosslinking. 1105 28