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)

mRNA's homologous to the herpes simplex virus type 1 DNA restriction endonuclease fragment BamHI p, which contains the thymidine kinase gene, have been identified and mapped by hybrid-arrested translation and mRNA selection. Such mRNA's, when translated in vitro, directed the synthesis of polypeptides of apparent molecular weights 43,000 (VI43) and 39,000 (VI39). mRNA for enzymatically active thymidine kinase was enriched by more than 20-fold after selection. Mapping was carried out with restriction endonuclease fragments of BamHI p, and locations of the 5' and 3' termini of VI43 mRNA were deduced. Analysis of nucleotide sequences around the 5' terminus revealed several consensus sequences commonly found at the start of eucaryotic mRNA's and which are presumably involved in initiation of transcription by RNA polymerase II. Translation of mRNA's for VI43, VI39, and the thymidine kinase enzyme was arrested only by a 1,170-base-pair region of BamHI p. Since this region is insufficient for adjacent genes, coding sequences for VI43 and VI39 must overlap; the possible relationship of these two polypeptides is discussed. A virus-induced product equivalent to VI39 was detected in infected cells.
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PMID:Identification and mapping of two polypeptides encoded within the herpes simplex virus type 1 thymidine kinase gene sequences. 626 30

We have identified a transcriptional control region of the herpes simplex virus tk gene. This finding results from in vivo transcription assays of specific deletion mutants constructed in vitro. A region located between 40 and 100 nucleotides upstream from the putative transcription start site of the tk gene can promote transcription by RNA polymerase form II in the complete absence of the mRNA-coding component of the gene. When the region is deleted enzymatically from a 5' direction, accurate transcriptional expression is reduced by a factor of 50. The small remaining level of accurate transcription is eliminated by deletion of sequences from 32 to 16 nucleotides upstream from the the structural gene. It appears that the sequences between 16 and 32 nucleotides upstream from the 5' terminus of the tk gene are required to specify the exact start site of transcription. Control of both the efficiency of transcription and rough specification of the position of initiation, however, depends on sequences 40--100 nucleotides upstream from the tk structural gene.
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PMID:Analysis of transcriptional regulatory signals of the HSV thymidine kinase gene: identification of an upstream control region. 626 44

The hybrid plasmid pTK1 consists of the herpes simplex virus type 1 (HSV-1) BamHI p fragment, which contains the thymidine kinase (TK) gene, inserted into the vector pAT 153. When pTK1 DNA was microinjected into nuclei of Xenopus laevis oocytes, functional HSV-1-specific TK was produced, showing that transcription and translation of the gene occurred. Investigation of pTK1-specific RNA by "Southern' blot hybridization revealed that all regions of the hybrid plasmid were transcribed by RNA polymerase II, but sequences present in TK mRNA were most highly represented in stable transcripts.
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PMID:Transcription and translation of the herpes simplex virus type 1 thymidine kinase gene after microinjection into Xenopus laevis oocytes. 627 Feb 58

We describe a 2560 base pair herpes simplex virus type 1 (HSV-1) DNA sequence containing the entire immediate-early mRNA-5 (IEmRNA-5) gene. The 3' and 5' termini of IEmRNA-5 were mapped within this DNA sequence by single-strand specific endonuclease protection experiments. The IEmRNA-5 gene contains DNA sequences from both the unique (Us) and reiterated (TRs/IRs) regions of the HSV-1 DNA short component and is interrupted by a single intron mapping in TRs/IRs. A search of the transcribed DNA sequence revealed no initiator codon within TRs/IRs. The first ATG was located 6 bases into Us sequences and this reading frame (316 codons) was also observed in the 3' transcribed region. The oligonucleotide sequences adjacent to the IEmRNA-5 termini are discussed in relation to those of the HSV-1 thymidine kinase gene and other genes transcribed by RNA polymerase II.
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PMID:DNA sequence of an immediate-early gene (IEmRNA-5) of herpes simplex virus type I. 627 43

Several animal viruses are known to contain significant amounts of polyamines but so far the function of these viral components is poorly understood. In this study the role of polyamines in the replication of two different types of viruses, herpes simplex virus type 2 and Semliki Forest virus (SFV) has been investigated. Purified SFV was found to contain fairly small amounts of polyamines, sufficient to neutralize only about 3% of viral nucleic acid phosphate, i.e., 1/20 of that found in herpes simplex virus. The production of both viruses was, however, markedly inhibited in BHK21 cells depleted of polyamines by treatment with alpha-difluoromethylornithine, a specific inhibitor of polyamine synthesis. This inhibition was reversed by putrescine, spermidine and spermine, and at least partly reversed by several other diamines and polyamine homologs. The activity of viral RNA polymerase induced by SFV infection was markedly reduced in polyamine-depleted cells but increased rapidly after addition of spermidine to the culture medium. It appears that the inhibition of virus production in polyamine-depleted cells is due in part to malfunction of protein synthetic machinery of the host cell. The possibility that other steps in virus synthesis and assembly are affected by polyamine deficiency is currently being investigated.
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PMID:Roles of polyamines in the replication of animal viruses. 627 79

We used partially purified RNA polymerase II from uninfected (Pol II) and from herpes simplex virus type 1 (HSV-1) infected HEp-2 cells (Pol II-H) to transcribe HSV-1 DNA in vitro. Gel electrophoretic analysis of the products produced from native HSV-1 DNA yielded weight average chain lengths of 4.0 and 3.5 kb for the Pol II and Pol II-H products, respectively. Blot hybridization analyses of the HSV DNA transcripts showed that both enzymes transcribed RNA from essentially all regions of the genome. However, Pol II preferentially transcribed regions coding for the immediate-early or alpha mRNAs, whereas Pol II-H preferentially copied regions coding for the early (beta) and late (gamma) gene products. Transcriptional analyses of the cloned HSV-1 Bam HI-Q fragment (containing the thymidine kinase (TK) gene) and its subfragments showed that (1) the major transcripts produced by Pol II-H were distinctly different from those produced by Pol II; (2) Pol II and Pol II-H utilized different promoters for the synthesis of major transcripts; (3) both enzymes produced three minor transcripts that were partially overlapping and in opposite direction to the TK gene; and (4) only Pol II-H initiated transcription from the TK promoter. In contrast, both Pol II and Pol II-H generated an identical set of transcripts from an adenovirus 2 early region DNA fragment. The sizes of the products suggest that RNA processing may be occurring in vitro. These results show that HSV-1 infection alters the in vitro transcriptional specificity of RNA polymerase II and demonstrate that this system should be useful for studying in vitro the regulation of gene transcription.
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PMID:Regulation of herpes simplex virus gene transcription in vitro. 629 54

A transcription unit at the herpes simplex virus (HSV) type 2 transforming region, mtr-2 (map coordinates 0.580-0.625), comprises two early, unspliced mRNAs of 4.5 kb and 1.2 kb which are 3' co-terminal; a region including that specifying the 1.2-kb mRNA has been sequenced. The putative translated portions of these two mRNAs do not overlap and this feature, together with the arrangement of the mRNAs, is similar to the apparently equivalent co-linear HSV-1 locus which however does not transform. A putative stem and loops structure containing a TATA box is located upstream from the 5' terminus of the 1.2-kb mRNA within the translated portion of the 4.5-kb mRNA. Evidence for the generation of this structure by intra-strand reassociation under our hybridisation conditions has been obtained and possibilities are that it may function as a transcription-activated promoter or as an RNA polymerase pause site. A comparison of the equivalent HSV-2 and HSV-1 regions reveals a conserved sequence downstream from the 3' co-terminus which is present at a similar location in many eukaryotic genes (consensus sequence YGTGTTYY). The overall sequence conservation at this transcription unit is high except for regions located at: (1) the untranslated leaders of the 1.2-kb mRNAs; (2) the N termini of the polypeptides specified by the HSV-2/HSV-1 1.2-kb mRNAs; (3) the intergenic region beyond the 3' co termini. Regions (2) and (3) are located within a transforming fragment of HSV-2. The possible significance of these data for HSV-mediated cell transformation is discussed.
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PMID:DNA sequence homology between two co-linear loci on the HSV genome which have different transforming abilities. 631 8

An in vitro approach has been used to study trout protamine gene expression using various recombinant plasmids containing trout protamine genes as templates in the HeLa cell lysate transcription system. The specific RNA transcript which is protected against S1 nuclease digestion by hybridization to the protamine gene sequence is alpha-amanitin sensitive (1 micrograms/mL), showing that RNA polymerase II is involved. The sizes of transcripts from templates linearized with Bam HI, Rsa I, and Hpa II (all downstream from the putative TATA box) are consistent with those predicted from the known sequence of the protamine gene. Digestion at an Alu I site only 14 base pairs (bp) upstream from TATA box has no effect on the accuracy of transcription in vitro; however, cutting at an Ava II site 9 bp downstream from the TATA box (reading from the first T) abolishes transcription. Chimeric plasmids, in which a herpes simplex virus (HSV-1) thymidine kinase (tk) promoter is tandemly inserted upstream from the trout protamine DNA sequences or as a replacement of the natural protamine promoter, were constructed. Use of these plasmids allowed an examination in a single assay of eight different putative promoter sequences (TATAAAA, TATAAA, TACAAA, TATATA, TATTTAA, CATATTA, TATATTAT, and TATTTAT) that are localized in either the protamine or the tk genes. The canonical TATAAAA promoter (the natural protamine promoter) was the strongest one and, in its presence, none of the others were used significantly for transcription. However, when this promoter was removed the weaker promoters were able to promote transcription.
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PMID:Transcription of a trout protamine gene in vitro: the effects of alteration of promoters. 632 91

Herpes simplex virus type 1 (HSV-1) establishes latency in human sensory ganglia, during which time the viral genome is transcriptionally silent with the exception of the latency-associated transcripts (LATs). The most abundant LAT is a 2-kb RNA whose biosynthesis is poorly characterized. The 2-kb LAT may be a primary transcript, or its synthesis may involve splicing and/or other forms of processing. Two potential RNA polymerase II promoters (LAP1 and LAP2) upstream of the 2-kb LAT 5' end have been identified. To investigate the role played by LAP1 and LAP2 in the synthesis of the 2-kb LAT under lytic and latent conditions, we analyzed HSV-1 mutants which contain deletions of one or both of these promoters. During lytic infection in cell culture, the cis elements critical for the normal accumulation of the 2-kb LAT were mapped to LAP2, while LAP1 sequences were largely dispensable. The 5' ends of the major 2-kb LATs produced by the wild-type and LAP deletion viruses were examined by primer extension analysis and were all found to be identical (+/- 2 bp). The accumulation of the 2-kb LAT during latent infections of murine trigeminal ganglia was examined by Northern (RNA) blot and by reverse transcription-PCR. In contrast to the results found in lytic infections, the critical cis elements needed for 2-kb LAT accumulation during latency were mapped to LAP1. Deletion of LAP1 resulted in a 500-fold reduction in 2-kb LAT accumulation, whereas deletion of LAP2 resulted in only a 2- to 3-fold reduction. Deletion of both LAP1 and LAP2 resulted in undetectable levels of the 2-kb LAT. Our results indicate that both LAP1 and LAP2 are critical for 2-kb LAT expression but under different conditions. LAP1 is essential for LAT expression during latency, while LAP2 is primarily responsible for LAT expression in lytic infections in cell culture. LAP1 and LAP2 may prove to be functionally independent promoter elements that control 2-kb LAT expression during different stages of HSV-1 infections.
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PMID:Two herpes simplex virus type 1 latency-active promoters differ in their contributions to latency-associated transcript expression during lytic and latent infections. 749 2

Regulation of chain elongation by RNA polymerase II can have an important effect on gene expression (Bentley, D. (1995) Curr. Opin. Genet. Dev. 5, 210-216; Yankulov, K., Blau, J., Purton, T., Roberts, S., and Bentley, D. (1994) Cell 77, 749-759); however the mechanisms that control this step in transcription are not well understood. The adenosine analogue 5,6-dichloro-1-beta-D-ribofuranosylbenzimidazole (DRB) has long been used as an inhibitor of RNA polymerase II elongation, but its target is not known. We show that DRB is a potent inhibitor of Cdk-activating kinase, associated with the general transcription factor TFIIH. Two other inhibitors of this kinase, H-7 and H-8, also inhibited transcriptional elongation. Furthermore, TFIIH kinase bound specifically to the herpes simplex virus VP16 activation domain which stimulates polymerase II elongation in addition to initiation (Yankulov, K., Blau, J., Purton, T., Roberts, S., and Bentley, D. (1994) Cell 77, 749-759). Our results suggest that DRB affects transcription by inhibiting the TFIIH-associated kinase and that this kinase functions in the control of elongation by RNA polymerase II.
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PMID:The transcriptional elongation inhibitor 5,6-dichloro-1-beta-D-ribofuranosylbenzimidazole inhibits transcription factor IIH-associated protein kinase. 759 83


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