Topoisomerization of plasmid DNA in Escherichia coli infected with bacteriophage T4

Albright, Lisa M.; Geiduschek, E. Peter

doi:10.1016/0022-2836(86)90005-7

Cited by 7 publications

(7 citation statements)

References 44 publications

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“…la and b, compare lane 12 with lane 11). Thus, the T4 topoisomerase II relaxed negative supercoils and the E. coli DNA gyrase maintained negative supercoils in pTE114 DNA after T4 infection, as was previously shown for pLA4 (3).…”

Section: Resultssupporting

confidence: 80%

“…During infection by T4 reference-type phage, the average linking number of the topoisomer distribution of pTE114 increased; we refer to this process as relaxation. We have argued that this relaxation, seen in protein-free DNA, indicates a decrease in both restrained and unrestrained supercoils in vivo (3). The increase in average linking number was accompanied by a dramatic broadening of the narrow distribution that was characteristic of uninfected cells ( Fig.…”

Section: Resultsmentioning

confidence: 91%

“…In our initial experiments, we used a "minipromoter" plasmid, pTE114, containing only 35 bp of T4 DNA spanning the gene 23 promoter (14). pTE114 sustained changes in topoisomer distribution during T4 infection that were comparable to those previously analyzed in pLA4, which contains the entire gene 23 transcription unit (3). During infection by T4 reference-type phage, the average linking number of the topoisomer distribution of pTE114 increased; we refer to this process as relaxation.…”

Section: Resultsmentioning

confidence: 94%

“…We have argued that there are two reasons for this topoisomerization (3). The first is competition between the T4-encoded topoisomerase II supercoils, with concomitant action by topoisomerases to equilibrate torsional stress.…”

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confidence: 99%

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Bacteriophage T4 late transcription from plasmid templates is enhanced by negative supercoiling

1988

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Concurrent viral replication is normally required to activate bacteriophage T4 late promoters; replication is thought to provide a template structure which is competent for late transcription. Transcription from plasmid-borne T4 late promoters, however, is independent of replication in vivo and in vitro. In this work, we have shown that, when the late gene 23 promoter is located on a plasmid, its utilization in vivo depends upon the ability of host DNA gyrase to maintain some degree of negative superhelicity. This suggests that an alternative pathway exists for activation of late promoters: DNA which is under sufficient negative torsional stress is already competent for late transcription. We also describe a method for isolating ternary complexes of plasmid DNA, RNA polymerase, and nascent RNA which have initiated transcription in vivo. The topoisomer distribution of such ternary complexes prepared from T4-infected cells showed that, late in infection, transcriptional activity resides primarily in the subset of the plasmid population with the most negatively supercoiled topoisomers. However, the overall transcriptional pattern in these ternary complexes indicated that both vector and T4 sequences are actively transcribed. Much of this transcriptional activity could be independent of gp55, the T4-specific RNA polymerase-binding protein that confers late promoter recognition.During infection of Escherichia coli by bacteriophage T4, a program of modifications to the host RNA polymerase generates three major temporal classes of phage transcripts (reviewed in references 7, 15, and 42). Late transcription requires the products of T4 genes 33, 45, and 55. Gene product 55 (gp55) confers specificity for late T4 promoters on RNA polymerase core (26). The roles of gp33 and gp45 in transcription remain unclear, but gp45 is known also to play a role in DNA replication as an accessory protein to the replication complex (1). Late promoters share the sequence TATAAATA centered at about position -10 from the start of transcription, but require no position -35 sequence (12,14).Late transcription is also distinguished by a requirement for concurrent viral DNA replication (45). Late transcription can be uncoupled from replication in genetic backgrounds which are thought to stabilize nicks in the DNA. These observations led to the hypothesis that late transcription requires a competent template which has been activated by DNA replication (46). Uncoupling can also be achieved by placing the promoter on a plasmid template (21, 23). Late transcription from plasmid templates in vivo is otherwise faithful in its initiation sites and in regulation by genes 33, 45, and 55 (14, 23). Additionally, supercoiled plasmids are efficient templates for in vitro transcription with purified T4 late RNA polymerase (24). Therefore it appears that plasmid templates are intrinsically competent for late transcription.The topoisomer profile of plasmids changes drastically after T4 infection. We have argued that there are two reasons for this topoisomerizat...

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Section: Resultssupporting

confidence: 80%

Section: Resultsmentioning

confidence: 91%

Section: Resultsmentioning

confidence: 94%

mentioning

confidence: 99%

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Bacteriophage T4 late transcription from plasmid templates is enhanced by negative supercoiling

1988

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“…The protocol used to investigate changes in DNA topology was adapted from previous procedures (Pettijohn and Pfenniger, 1980;Albright and Geiduschek. 1986;BaIke and Gralla.…”

Section: Dna Supercoiling Can Change During Starvation In Vivomentioning

confidence: 99%

Interrelated effects of DNA supercoiling, ppGpp, and low salt on melting within the Escherichia coli ribosomal RNA rrnB P₁ promoter

Ohlsen

Gralla

1992

Molecular Microbiology

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The formation of complexes containing high levels of DNA melting at the ribosomal RNA rrnB P1 promoter in vitro is shown to be facilitated by DNA supercoiling or low salt. The effector nucleotide ppGpp is ineffective under these conditions. The loss of supercoils or addition of salt increases the effectiveness of ppGpp in inhibiting formation of these complexes. In vivo plasmid DNA supercoiling is shown to decrease during starvation protocols that also increase levels of ppGpp. The results suggest that ppGpp regulation may be affected by the state of DNA supercoiling in vivo.

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Transcription and messenger RNA processing upstream of bacteriophage T4 gene 32

1989

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Bacteriophage T4 gene 32 lies at the 3' end of a complex transcription unit which includes genes 33, 59, and several open reading frames. In the course of an infection, four major transcripts are synthesized from this unit: two overlapping polycistronic transcripts about 3800 and 2800 nucleotides in length, and two monocistronic gene 32 transcripts about 1150 and 1100 nucleotides in length. These transcripts are made at different times in infection and the polycistronic transcripts have segmental differences in stability. Messenger RNA processing yields a 1025 nucleotide monocistronic gene 32 transcript, and a 135 nucleotide transcript containing part of the gene 59 coding sequence. Processing depends on Escherichia coli encoded ribonuclease E. This pattern of transcription and processing leads to the synthesis of gene 32 mRNA throughout infection, whereas transcripts encoding the upstream genes are present only early in infection. The 3800 nucleotide polycistronic transcript initiates at a promoter that does not require T4 encoded factors for activity. However, full-length synthesis of this transcript depends on the T4 mot gene product. The region upstream of gene 32 also contains four E. coli-like promoters that are active on chimeric plasmids in uninfected cells, but inactive in bacteriophage T4. The location of these cryptic T4 promoters is intriguing in that they lie near the 5' ends of open reading frame B, gene 59 and gene 32. They could play a role in phage development under particular conditions of growth or in bacterial hosts other than those examined here.

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Topoisomerization of plasmid DNA in Escherichia coli infected with bacteriophage T4

Cited by 7 publications

References 44 publications

Bacteriophage T4 late transcription from plasmid templates is enhanced by negative supercoiling

Bacteriophage T4 late transcription from plasmid templates is enhanced by negative supercoiling

Interrelated effects of DNA supercoiling, ppGpp, and low salt on melting within the Escherichia coli ribosomal RNA rrnB P₁ promoter

Transcription and messenger RNA processing upstream of bacteriophage T4 gene 32

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Topoisomerization of plasmid DNA in Escherichia coli infected with bacteriophage T4

Cited by 7 publications

References 44 publications

Bacteriophage T4 late transcription from plasmid templates is enhanced by negative supercoiling

Bacteriophage T4 late transcription from plasmid templates is enhanced by negative supercoiling

Interrelated effects of DNA supercoiling, ppGpp, and low salt on melting within the Escherichia coli ribosomal RNA rrnB P1 promoter

Transcription and messenger RNA processing upstream of bacteriophage T4 gene 32

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Interrelated effects of DNA supercoiling, ppGpp, and low salt on melting within the Escherichia coli ribosomal RNA rrnB P₁ promoter