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)

Sequence-specific methylation of the promoter and adjacent regions in mammalian genes transcribed by RNA polymerase II leads to the inhibition of these genes. So far, RNA polymerase III-transcribed genes have not been investigated in depth. We therefore studied methylation effects on the RNA polymerase III-transcribed VAI gene of adenovirus type 2 DNA. The VAI gene contains 20 5'-CG-3' dinucleotides, of which 4 (20%) can be methylated by HpaII (5'-CCGG-3') and HhaI (5'-GCGC-3'). Three of these 5'-CG-3' sequences are located close to the internal regulatory region of the VAI segment. An unmethylated, a 5'-CCGG-3'- and 5'-GCGC-3'-methylated, and a 5'-CG-3'-methylated pUC18 construct containing the VAI and VAII regions were transfected into mammalian cells. In many experiments, an inactivating effect of 5'-CCGG-3' and 5'-GCGC-3' DNA methylation on the VAI region was not observed. In contrast, methylation of all 20 5'-CG-3' sequences in the VAI region by a CpG-specific DNA methyltransferase from Spiroplasma species did interfere with VAI transcription. Transcription of the VAI- and VAII- and of the VAI-containing constructs was also shown to be inhibited in an in vitro cell-free transcription system after the constructs had been methylated at the 5'-CCGG-3' and 5'-GCGC-3' sequences or at all 5'-CG-3' sequences. When an oligodeoxyribonucleotide which carried the internal control block A of the VAI region was methylated at three 5'-CG-3' sequences, the formation of a complex with HeLa nuclear proteins was abrogated. The results presented support the notion that the VAI gene transcribed by the DNA-dependent RNA polymerase III is also inactivated by methylation of the decisive 5'-CG-3' sequences.
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PMID:Adenovirus type 2 VAI RNA transcription by polymerase III is blocked by sequence-specific methylation. 200 41

Methylation of cytosine in the DNA inhibits the transcription by RNA polymerase II in higher eukaryotes, but has no influence on RNA polymerase I transcription. The effect on RNA polymerase III was unknown, so far. Two polymerase III genes: a type 1 5S rRNA gene and a type 2 tRNA gene were methylated in vitro with a purified eukaryotic DNA methyltransferase (EC2.1.1.37) and their transcription was analyzed in Xenopus oocytes. The 5S rRNA gene, an oocyte 5S rRNA gene from X. laevis which is subject to developmental inactivation, was not affected by methylation. Conversely, transcription of the tRNA gene was 80% inhibited by methylation with the eukaryotic methyltransferase. HhaI and HpaII methylation left its transcription unaffected.
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PMID:DNA methylation inhibits transcription by RNA polymerase III of a tRNA gene, but not of a 5S rRNA gene. 240 61

In vitro transcription in a HeLa cell lysate by RNA polymerase II directed by a chicken feather keratin gene promotor has been studied using unmethylated template DNA and DNA methylated in vitro by HpaII methylase. The efficiency of specific gene transcription from methylated DNA was dependent on topology of the input DNA, the most significant effect being complete inhibition of transcription from one template which contained three methylation sites, one just 5' and two greater than 500 bases 3' to the site of transcription initiation. The inhibition of transcription depends on a factor(s) which is variably present in lysate preparations and is labile on storage at -70 degrees.
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PMID:Effects of DNA methylation on specific transcription by RNA polymerase II in vitro. 241 49

The genome of the large icosahedral DNA virus, frog virus 3 (FV3), is heavily methylated at the cytosine residues of dCdG dinucleotide pairs, with more than 22% of the total cytosine residues in the form of 5-methylcytosine (5mC). This methylation is carried out postreplicatively in the cytoplasm of infected cells by a virus-encoded DNA methyltransferase. DNA methyltransferase activity was shown to copurify with a 26 kD virus-induced, DNA-binding protein that had an altered mobility in extracts from cells infected with a DNA-methyl-transferase deficient mutant of FV3. Immediately after infection, the highly methylated parental DNA is transcribed in the nucleus by the host cell RNA polymerase II. As FV3 induces the synthesis of a protein that can override the inhibitory effect of methylation on the transcription of exogenous promoters methylation in vitro, we suggest that this protein is a factor evolved by this virus to allow transcription from methylated promoters by eukaryotic RNA polymerase II.
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PMID:Transcription of methylated viral DNA by eukaryotic RNA polymerase II. 247 31

The genome of the iridovirus, frog virus 3, is highly methylated at cytosine residues by a virus-encoded DNA methyltransferase. We have shown previously that an FV3-induced trans-acting protein alters either host RNA polymerase II or methylated template to allow transcription from promoters inactivated by methylation. We now present evidence that the ability of FV3-infected cells to transcribe methylated DNA is specific for DNA methylated at cytosine residues. Eukaryotic promoters were inactivated by methylation of either adenine or cytosine residues, and tested for transcriptional activity. Only promoters inactivated by cytosine methylation were transcribed in FV3-infected cells. We also show that the dinucleotide sequence in which the methylcytosine is found appears to have no effect on the ability of FV3 to trans-activate the methylated promoters.
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PMID:Infection with frog virus 3 allows transcription of DNA methylated at cytosine but not adenine residues. 362 76

A 2.5 X 10(3) base-pair segment of Bacillus sphaericus R DNA cloned in Escherichia coli has previously been shown to carry the functional BspRI modification methylase gene. The approximate location of the gene on this DNA segment and its direction of transcription were established by subcloning experiments. The nucleotide sequence of the relevant region was determined by the Maxam-Gilbert procedure. An open reading frame that can code for a 424 amino acid protein was found. The calculated molecular weight (48,264) of this protein is in fair agreement with previous estimates (50,000 to 52,000). The synthesis of this protein was demonstrated in E. coli minicells. The initiation point of transcription by E. coli RNA polymerase was localized by in vitro transcription experiments. The open reading frame starts 29 base-pairs downstream from the transcription initiation site and it is preceded by a sequence showing extensive Shine-Dalgarno complementarity. Subcloning experiments and translation in minicells suggest that after removal of this translational initiation site, a secondary start site 29 amino acids downstream can also start translation in E. coli, and this shorter protein retains the methylase activity. The overall base composition of the gene and the codon usage indicate a strong preference for A.T base-pairs.
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PMID:Structure of the Bacillus sphaericus R modification methylase gene. 631 47

We have determined the nucleotide sequence of a 4.0-kilobase DNA fragment containing the genes of the PstI restriction-modification system. Two large open reading frames were identified within the sequence and were ascribed to the restriction enzyme and methylase by the analysis of a series of deletion mutants. The two genes are encoded on opposite DNA strands, and hence must be transcribed from separate promoters rather than as a polycistronic message. The sequence of the first 10 amino acids of the restriction endonuclease was determined by sequential Edman degradation of the purified protein, permitting the alignment of the polypeptide with the DNA sequence. The NH2 terminus of the modification enzyme was established by sequential Edman degradation of the protein synthesized in bacterial minicells with different radiolabeled amino acids. The initiation codons of the two genes are separated by 130 base pairs. The deduced amino acid sequences indicate that the restriction endonuclease contains 326 amino acids with a calculated Mr = 37,370; the modification enzyme is composed of 507 amino acids with a calculated Mr = 56,830. There is no significant homology between the two proteins at the level of the primary structure. Antibody raised against the purified restriction endonuclease did not immunoprecipitate the modification enzyme. The transcription initiation sites were mapped using mung bean nuclease. Both of the transcripts begin with adenosine. The initiation sites are separated by only 70 base pairs. This close proximity suggests that the promoters for the two divergent genes overlap. DNase I protection experiments show that Escherichia coli RNA polymerase has a higher affinity for the methylase promoter than for the restriction enzyme promoter.
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PMID:The organization and complete nucleotide sequence of the PstI restriction-modification system. 633 92

Further research on bacteriophage T7 DNA penetration mechanism into E. coli cells during the infection was carried out. The DNA-RNA-hybridization on nitrocellulose filters revealed that in the presence of chloramphenicol the T7 DNA penetration from the virion into a host cell was coupled with its transcription by the bacterial RNA polymerase. The data obtained indicate that in the absence of antibiotics the penetration of a part of T7 genome which correspondes to class II and III genes is coupled with its transcription by a phage-specific RNA polymerase. Along with this the host restriction-modification system when its activity is not inhibited by the phage-induced proteins will be able to cleave the penetrated T7 DNA just after its transcription was accomplished. Considering these data along with our conception on direct involvement of transcription in T7 DNA penetration process during the infection one can suggest that E. coli RNA polymerase molecules which provide the phage DNA transport, are localized at the inner surface of cytoplasmic membrane.
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PMID:[Coupling of bacteriophage T7 DNA penetration with its transcription, during infection]. 635 19

DNA methyltransferase was purified 310-fold from a green alga, Chlamydomonas reinhardi vegetative cells. The native enzyme of molecular weight 55 000--58 000 catalyzed the transfer of methyl groups from S-adenosylmethionine to the 5 position of cytosine in DNA. Native DNA accepted methyl groups 10-fold more than did denatured DNA. The sequence specificity analysis of methylated deoxycytidine in vitro revealed that the enzyme introduces methyl groups preferentially into sequences containing 5'd(T-mC-R)3'. Kinetic analysis of the reaction indicated that the enzyme obeys a random sequential mechanism. The extent of saturation with methyl groups depends upon the species from which the DNA was obtained. Kinetic analysis of the reaction catalyzed by RNA polymerase II has indicated that DNA methylation decreases the rate of initiation of RNA synthesis, but does not affect the rate of RNA chain elongation.
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PMID:Deoxyribonucleic acid methyltransferase from the eukaryote, Chlamydomonas reinhardi. 737 44

The effect of 5-aza-CR and 5-aza-2'-deoxycytidine (5-aza-CdR) on cell differentiation and DNA methylation of HL-60 cells was studied. The differentiation index of HL-60 cells was measured after being treated with drugs by using the NBT stain method. DNA methylase activities of HL-60 cells treated with the drugs were assayed by using 3H-methyl-S-adenosylmethionine (3H-SAM) as a methyl donor. The DNA methylation level of HL-60 cells treated with the drugs was measured by HPLC. The results showed that the HL-60 cell differentiation index was increased after being treated with 5-aza-CR or 5-aza-CdR at a certain concentration for 4 days. But, at the same time, DNA methylase activity and the DNA methylation level were decreased. And all these changes were related to the concentration of the drugs. 5-Aza-CdR was more efficient than 5-aza-CR. We also assayed the E. coli RNA polymerase activity in vitro by using different DNA templets different in DNA methylation level. We found that the transcriptional activity of RNA polymerase was increased with the decrease of the DNA methylation level of HL-60 cells.
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PMID:The effect of 5-azacytidine (5-aza-CR) and its analogue on cell differentiation and DNA methylation of HL-60 cells. 769 Oct 71


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