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)

The single-copy RP2 gene in mice produces three major mRNAs, the abundances of which are significantly increased in the kidneys by the administration of testosterone. S1 nuclease analysis of the kidney mRNAs indicated that they differ in the lengths of their 3' untranslated regions as a result of the use of different polyadenylation sites. When the mRNAs from different inbred mouse strains were examined by Northern blot analysis, it was observed that the largest mRNA varies in size, whereas the sizes of the other mRNAs remain the same. In DBA/LiHa and DBA/2J mice, the largest mRNA is approximately 2,150 nucleotides long, whereas the corresponding mRNA in C57BL/6J and BALB/cJ mice is only 1,950 nucleotides in length. All of these strains also have RP2 mRNAs that are 1,450 and 1,350 nucleotides long. By S1 nuclease mapping and comparison of the sequence of cDNA clones representing these mRNAs in DBA/LiHa and C57BL/6J mice, we determined that this size difference or polymorphism observed in the largest mRNA is the result of the insertion of a member of the B1 family of repeats into the 3' untranslated region of the RP2 gene in DBA mice. This particular B1 repeat is transcribed by RNA polymerase III in vitro, and its transcriptional orientation is opposite to that of the RP2 transcript. The polymorphism described here is evidence for the mobility of B1 repetitive elements within the genome.
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PMID:Polymorphism in an androgen-regulated mouse gene is the result of the insertion of a B1 repetitive element into the transcription unit. 302 23

As shown by Southern blot analysis, the metallothionein-1 (MT-1) genes in rats comprise a multigene family. We present the sequence of the MT-1 structural gene and compare its features with other metallothionein genes. Three MT-1 pseudogenes which we sequenced apparently arose by reverse transcription of processed mRNA transcripts. Two of these, MT-1 psi a and MT-1 psi c, are retrogenes which derive from the MT-1 mRNA, having diverged from the MT-1 gene 6.9 and 2.6 million years ago, respectively. The third, MT-1 psi b, differs from the MT-1 cDNA by only three nucleotide alterations. Surprisingly, MT-1 psi b also preserves sequence homology for 142 base pairs 5' to the transcription initiation site of the parent gene; it contains a promoter sequence sufficient for specifying metal ion induction. We identified, by S1 nuclease mapping, an RNA polymerase II initiation site 432 base pairs 5' of the MT-1 transcription initiation site of the MT-1 structural gene which could explain the formation of the mRNA precursor to this pseudogene. We were unable to detect MT-1 psi b transcripts, either in liver tissue or after transfection. We conclude that the absence of detectable transcripts from this pseudogene is due to either a reduced level of transcription or the formation of unstable transcripts as a consequence of the lack of a consensus sequence normally found 3' of transcription termination in the MT-1 structural gene.
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PMID:Rat metallothionein-1 structural gene and three pseudogenes, one of which contains 5'-regulatory sequences. 302 30

We devised a strategy to measure the efficiency of transcription termination in vivo by RNA polymerase on polyomavirus DNA. Pulse-labeled nuclear RNA was hybridized with a single-stranded polyomavirus DNA fragment which spans the transcription initiation region. Hybrids were treated with RNase, bound to nitrocellulose filters, eluted with S1 nuclease, and analyzed by gel electrophoresis. The ratio of full-length to less-than-full-length DNA-RNA hybrids was used to calculate transcription termination frequency. We found that 50% of the polymerases terminated per traverse of the L DNA strand during the late phase of infection. The method for mapping in vivo pulse-labeled RNAs which we developed is potentially useful for studying unstable cellular or viral RNAs.
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PMID:Use of a novel S1 nuclease RNA-mapping technique to measure efficiency of transcription termination on polyomavirus DNA. 302 98

A region that forms the S1 nuclease site in Co1E1 DNA is shown to code for an in vitro transcript, called S1 RNA-B, which contains a 5'-terminal GTP residue that exhibits noncomplementarity with the template's DNA sequence. The synthesis of S1 RNA-B initiates four bases upstream from the start point for S1 RNA-C. The initial four bases in S1 RNA-B and S1 RNA-C are identical. The relative synthesis of S1 RNA-B to S1 RNA-C is sensitive to the concentration of GTP, a substrate that is required for elongation past the +4 position in S1 RNA-C. Dinucleotides that are expected to only initiate synthesis of S1 RNA-C yield two transcripts that appear to initiate from the S1 RNA-C and S1 RNA-B start sites. In vitro studies involving other Co1E1 transcripts, RNA-B and RNA-C, provide similar observations concerning the noncomplementary initiation phenomenon. A model involving transcriptional slippage is suggested to explain the noncomplementary initiation phenomenon. The model proposes that the cycling reaction of Escherichia coli RNA polymerase produces tetranucleotides that are transposed to nearby upstream sequences for priming transcription.
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PMID:Synthesis of in vitro Co1E1 transcripts with 5'-terminal ribonucleotides that exhibit noncomplementarity with the DNA template. 302 12

Transcription of the sporulation gene spoVG of Bacillus subtilis is induced at the onset of spore formation and depends on the products of the regulatory genes spoOA, spoOB, and spoOH. We describe two effects of propagating the promoter region of spoVG on a multicopy plasmid replicon in B. subtilis cells. One effect is that transcription from the plasmid-borne spoVG promoter is altered with respect to the time of its induction and the dependence on spoO gene products. An example of this effect is that plasmid propagation was observed to relieve substantially the inhibitory effect of a mutation in spoOH, the spoO gene upon which spoVG promoter activity is most strongly dependent. We present results which suggest that propagation on a plasmid replicon causes an alteration in the conformation of spoVG promoter DNA which somehow compensates for the defective spoOH gene product. Plasmid propagation did not, however, entirely eliminate the requirement for the spoOH gene product; little or no spoVG-directed RNA synthesis was observed in cells bearing a putative spoOH deletion mutation, a finding which indicates that SpoOH protein plays an indispensable role in spoVG promoter utilization. Another effect of propagating the promoter region of spoVG on a multicopy plasmid is to inhibit sporulation. S1 nuclease mapping experiments suggest that amplification of spoVG on a multicopy plasmid causes the titration of a transcription factor or minor form of RNA polymerase holoenzyme required for utilization of one of the two overlapping promoters which comprise the spoVG transcription initiation region.
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PMID:Effects of plasmid propagation of a sporulation promoter on promoter utilization and sporulation in Bacillus subtilis. 302 29

A 205bp DNA fragment from the Streptomyces multi-copy plasmid pIJ101 has in vivo terminator activity both in Streptomyces lividans and in Escherichia coli. Termination of RNA synthesis, detected by high-resolution S1 nuclease mapping, occurs at precisely the same nucleotides in both organisms. This suggests that the E. coli RNA polymerase recognizes the same sequence elements and chooses the point(s) of termination in the same way as the corresponding S. lividans enzyme.
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PMID:Activity of a Streptomyces transcriptional terminator in Escherichia coli. 303 7

A cDNA, p1-88, was cloned from a library constructed using rabbit liver mRNA. Sequence analysis indicates that p1-88 is highly similar (congruent to 95%) to the cDNA, p1-8, that encodes rabbit liver cytochrome P-450 1 and that had been isolated from the same library. The predicted amino acid sequence of the protein encoded by p1-88, P-450 IIC4, differs at 25 of 487 amino acids from that encoded by p1-8. P-450 IIC4 was synthesized in vitro using rabbit reticulocyte lysate primed with RNA transcribed from the coding sequence of p1-88 using a bacteriophage T7 RNA polymerase/promoter system. P-450 IIC4 reacts with two monoclonal antibodies that recognize P-450 1 and exhibits the same relative electrophoretic mobility as P-450 1. In contrast, the reactivity of a third monoclonal antibody recognizing P-450 1, 1F11, toward P-450 IIC4 synthesized in vitro is greatly diminished. The latter antibody extensively inhibits hepatic progesterone 21-hydroxylase activity and recognizes phenotypic differences among rabbits in the microsomal concentration of P-450 1. This difference in the immunoreactivity of P-450 IIC4 and P-450 1 with the 1F11 antibody suggests that P-450 IIC4 does not contribute significantly to hepatic progesterone 21-hydroxylase activity. S1 nuclease mapping demonstrates that the expression of mRNAs corresponding to p1-88 are expressed to equivalent extents in rabbits exhibiting high and low expression of mRNAs corresponding to p1-8. Thus, P-450 1 differs from the protein encoded by p1-88, in its regulation, immunoreactivity, and by inference its catalytic properties although the amino acid sequences of P-450 1 and P-450 IIC4 are highly similar (congruent to 95%).
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PMID:Characterization of a second gene product related to rabbit cytochrome P-450 1. 303 48

Expression of the genes in the phosphate regulon, including the pstS (phoS) and phoB genes, is positively regulated by PhoB protein when phosphate is limited. We purified PhoB protein from overproducing cells and studied its interaction with the pstS gene. It binds specifically to the DNA fragment containing the promoter region of pstS. The transcription initiation site of the gene in vivo was identified by S1 nuclease mapping and primer-extension experiments. In-vitro transcription of pstS was activated by the PhoB protein, and the initiation site of transcription agreed with the in-vivo initiation site. Activation of in-vitro transcription by PhoB protein required both the normal sigma factor (sigma 70) and core RNA polymerase. PhoB protein binding sites on the promoter regions of pstS and phoB were determined by footprinting experiments with DNase I and a methylating agent. In both cases the protein binds to the pho box, the concensus sequence shared by regulatory regions of genes in the phosphate regulon. Our findings indicate that PhoB protein recognizes and binds to the pho box and activates transcription of the genes in the phosphate regulon.
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PMID:Regulation of the phosphate regulon of Escherichia coli. Activation of pstS transcription by PhoB protein in vitro. 305 25

The nucleotide sequence of an 878-bp BamHI-BglII restriction endonuclease fragment from citrate utilization transposon Tn3411 was determined, and was compared with that from plasmid pMS185 [Sasatsu et al., J. Bacteriol. 164 (1985) 983-993]. A long open reading frame for a 379-amino acid (aa) polypeptide (citB) was found 5' to the citA gene (431-aa membrane protein) in Tn3411 as well as in pMS185. Promoter regions were identified by RNA polymerase filter-binding assays, S1 nuclease mapping and cit-lac fusion experiments. The results indicated that two genes (citA and citB) have separate promoters, and the location of the promoter for the citB gene in the Tn3411 nucleotide sequence was different from that in pMS185. The regulation of transcription of the two genes (citA and citB) was characterized by the use of cit-lacZ fusions. The level of the citB promoter activity was about five-fold higher than that of the citA gene promoter, and transcription from both was not induced by citrate. Synthesis of the mRNA for the citB gene (especially with the wild-type Cit+ determinant) was suppressed by citrate, accompanying growth suppression of Escherichia coli. The citB gene expressed in E. coli minicells produced a membrane-associated 37.5-kDa polypeptide.
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PMID:Promoters and transcription of the plasmid-mediated citrate-utilization system in Escherichia coli. 306 41

The control region of the carAB operon, encoding carbamoylphosphate synthetase, comprises two tandem promoters (P1, upstream and P2, downstream) located 67 base-pairs apart and repressed respectively by pyrimidines and arginine. RNA polymerase and pure arginine repressor bind to the P2 region in mutually exclusive ways. Repressor protects the two adjacent palindromic ARG boxes overlapping P2 against DNase I. Binding of RNA polymerase to P1 is abnormal; the region protected against DNase I is shifted upstream by about 20 nucleotides with respect to the position expected from the transcription startpoint. This pattern is not due to interference with polymerase binding at P2, since it is observed also in the presence of repressor and on an isolated P1 region. Binding of RNA polymerase is relatively weak and heparin-sensitive suggesting that, in vivo, an ancillary factor is required to promote the formation of an open complex. S1 nuclease mapping experiments show that the simultaneous presence of pyrimidines and arginine represses the downstream arginine-specific promoter (P2) more efficiently than arginine alone. This effect is not due to a direct regulatory interaction between pyrimidines and P2, since it is not observed when P1 is inactivated by insertion mutations or partial deletion. It has been shown that transcription initiated at P1 can proceed even when arginine represses P2. We therefore suggest that P2 operator-arginine repressor complex is destabilized by RNA polymerase binding at P1 or transcription from P1. We describe a novel technique to select for expression-down mutants in a lac fusion context.
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PMID:Molecular interactions in the control region of the carAB operon encoding Escherichia coli carbamoylphosphate synthetase. 306 19


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