Mapping of transcription units in the bacteriophage T4 tRNA gene cluster

Goldfarb, Alex; Daniel, Violet

doi:10.1016/0022-2836(81)90039-5

Cited by 24 publications

(10 citation statements)

References 33 publications

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“…They were separated by a modification of the acidic urea agarose gel electrophoresis procedure described by Rosen et al (1975) and visualized by autoradiography. The 16S rRNA bands were excised and eluted electrophoretically as described by Goldfarb and Daniel (1981). The 16S rRNAs were precipitated from the eluates and then washed and dried as described above for the total nucleic acids.…”

Section: Preparation Of 16s Rrnasmentioning

confidence: 99%

Taxonomic relations between archaebacteria including 6 novel genera examined by cross hybridization of DNAs and 16S rRNAs

Prangishvilli

Huber

et al. 1982

J Mol Evol

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Summary. DNAs from 16 species of archaebacteria including 6 novel isolates were hybridized with 16S rRNAs from 7 species representing different orders or groups of the urkingdom of archaebacteria.The yields, normalized for the number of genes per ¡ig of DNA, and the temperature stabilities of all hybrids were determined and related to each other.A taxonomic tree constructed from such fractional stability data reveals the same major divisions as that derived from comparative cataloging of 16S rRNA sequences. The extreme halophiles appear however as a distinct order besides the three known divisions of methanogens.The methanogens, the halophiles and Thermoplasma form one of two clearly recognizable branches of the archaebacterial urkingdom. The order represented by Sulfolobus and the related novel order Thermoproteales form the other branch.Three novel genera, Thermoproteus, Desulfurococcus and the "stiff filaments" represent three families of this order.The extremely thermophilic methanogen Methanothermusfervidus belongs to the Methanobacteriales. SN1, a methanogen from Italy, appears as another species of the genus Methanococcus. Another novel methanogen, M3, represents a genus or family of the order Methanomicrobidles.

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Section: Preparation Of 16s Rrnasmentioning

confidence: 99%

Taxonomic relations between archaebacteria including 6 novel genera examined by cross hybridization of DNAs and 16S rRNAs

Prangishvilli

Huber

et al. 1982

J Mol Evol

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“…1). They are initiated at different promoters, P1 and P2, and terminated at the same terminator site (33,34). The experiments reported here show that T4-modified RNA polymerase fails to recognize several ofthe T4 promoters including the P2 promoter of the tRNA genes.…”

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confidence: 70%

“…The samples were treated with DNase I (RNase-free, Boehringer Mannheim), deproteinized with phenol, and prepared for electrophoresis as described (33).…”

Section: Methodsmentioning

confidence: 99%

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Changes in the promoter range of RNA polymerase resulting from bacteriophage T4-induced modification of core enzyme.

Goldfarb¹

1981

Proc. Natl. Acad. Sci. U.S.A.

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Primary transcripts made in vitro on bacteriophage T4 DNA by RNA polymerase isolated from normal or T4-infected Escherichia coli were compared by gel electrophoresis. Bacteriophage-modified RNA polymerase fails to initiate transcription at certain promoters recognized by unmodified enzyme. In the T4 tRNA gene region, only one ofthe two promoters is active with the modified RNA polymerase. Reconstitution of separated RNA polymerase components demonstrates that this change in promoter site selection results from the modification of core enzyme and not a factor.During the development of bacteriophage T4 in its host Escherichia coli, complex changes occur in the transcription process. These changes include the shutoff of host transcription and sequential expression of three classes of bacteriophage genes, served by "early," "middle," and "late" promoters. The host RNA polymerase (nucleosidetriphosphate:RNA nucleotidyltransferase, EC 2.7.7.6) is apparently used for all transcription throughout bacteriophage infection, and it is generally believed that at least some ofthe changes in the transcription pattern are achieved through phage-induced changes in RNA polymerase specificity (for review, see ref. 1).Several modifications of RNA polymerase have been observed after T4 infection. They include chemical modification of the existing RNA polymerase subunits (2-13) and association with the enzyme of four small T4-coded polypeptides (14)(15)(16)(17)(18)(19)(20). Two of these new polypeptides (Mr 12,000 and 22,000) are the products ofT4 genes 33 and 55, which are required for the transcription ofthe late genes (15,16,18). No relationship has been established between other T4-induced RNA polymerase modifications and switches in the transcription pattern.Clearly, progress in understanding the molecular mechanisms of T4 transcription control depends on the development of in vitro systems that link particular RNA polymerase modifications with changes in transcription selectivity. Purified E. coli RNA polymerase in vitro recognizes only early promoters (21-23). In vitro transcription from middle (24) and late (25,26) promoters has been demonstrated only in crude lysates of T4-infected cells. The study of functional changes in T4-modified RNA polymerase by using purified transcription systems has thus far produced limited information. The modified enzyme competes poorly with the host RNA polymerase for template DNA (27), has a higher transition temperature for rapidly initiating transcription complexes (28), and is inhibited by 0.2 M KC1 (16). In contrast, the activity of the host RNA polymerase is markedly stimulated by KC1. The salt sensitivity of the modified enzyme is caused by a T4-coded polypeptide (Mr 10,000) that is associated with oa factor. RNA polymerase containing this protein fails to initiate transcription on T4 DNA at 0.2 M salt; this effect, however, can be overcome if 1% Triton X-405 is present at the moment of initiation (19,20). It has been shown that modified RNA polymerase transcribes certain bac...

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“…An easy way to identify these transcripts and to approximately map their endpoints on the T4 chromosome is to use as transcription templates DNA from T4 deletion mutants. This approach has already provided an approximate transcription map of the T4 tRNA gene cluster (4,6,7) and helped to elucidate some features of its control (8,9). In this work we employed the deletion analysis of in vitro transcripts to identify the products of another T4 genetic region -the D region, and to approximately map its transcription control signals.…”

Section: Introductionmentioning

confidence: 99%

Mapping of in vitro transcription units and identification of primary transcripts of the D region of bacteriophage T4

Goldfarb

Burger

1981

Nucl Acids Res

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The D region of bacteriophage T4 is comprised of six closely linked genes which are situated between 161 kb and 165 kb on the T4 chromosome. We studied the transcription of these genes in vitro by using DNA templates derived from a series of deletion mutants in this region. The mixture of primary products made by Escherichia coli RNA polymerase were fractionated by gel electrophoresis into discrete RNA species. The results obtained together with the known map positions of the deletions allowed to identify four wild-type and several deletion-specific transcripts of the D region. The end points of these transcripts were approximately mapped. The results demonstrate that the D region has two promoters and two terminators, an organisation which is similar to the previously established organisation of the T4 tRNA gene cluster.

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Mapping of transcription units in the bacteriophage T4 tRNA gene cluster

Cited by 24 publications

References 33 publications

Taxonomic relations between archaebacteria including 6 novel genera examined by cross hybridization of DNAs and 16S rRNAs

Taxonomic relations between archaebacteria including 6 novel genera examined by cross hybridization of DNAs and 16S rRNAs

Changes in the promoter range of RNA polymerase resulting from bacteriophage T4-induced modification of core enzyme.

Mapping of in vitro transcription units and identification of primary transcripts of the D region of bacteriophage T4

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