DNA sequences of promoter regions for the str and spc ribosomal protein operons in E. coli

Post, Leonard; Arfsten, Ann; Reusser, Fritz; Nomura, Masayasu

doi:10.1016/0092-8674(78)90096-x

Cited by 107 publications

(53 citation statements)

References 41 publications

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“…This is a greater decrement than that observed for the most polar trpB mutation known (29). Such [5][6]. The observed decrease in 3-galactosidase production associated with this DNA segment is consistent with the observation that the 3 and f' subunits of RNA polymerase are normally present in only one-fifth the molar yield of the ribosomal proteins (15).…”

Section: Resultssupporting

confidence: 84%

“…Recent reports that the genes for the a subunit (rpoA) and those for the 1 and 13' subunits (rpoBC) are in two separate transcription units, each of which also includes a number of ribosomal protein genes, suggest how this type of coordinate regulation could occur (4)(5)(6)(7). Yet under certain conditions, coordinate expression of the RNA polymerase subunits and ribosomal proteins is not observed.…”

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confidence: 99%

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Control features within the rplJL-rpoBC transcription unit of Escherichia coli.

Barry¹,

Squires²

1979

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

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Gene fusions constructed in vitro have been used to examine transcription regulatory signals from the operon which encodes ribosomal proteins L10 and L7/12 and the RNA polymerase P and #I subunits (the rplJL-rpoBC operon). Portions of this operon, which were obtained by in vitro deletions, have been placed between the ara promoter and the lacZ gene in the gene-fusion plasmid pMC81 developed by M. Casadaban and S. Cohen. The effect of the inserted DNA segment on the expression of the IacZ gene (in the presence and absence of arabinose) permits the localization of regulatory signals to discrete regions of the rpIJL-rpoBC operon. An element that reduces the level of distal gene expression to one-sixth is located on a fragment which spans the rplL-rpoB intercistronic region. This strongly supports the idea that there is an attenuator in this region. The terminator for the operon is located on a fragment which spans the 3' end of the rpoC gene. (13) or by the elicitation of the stringent response (9). The observed differential regulation of these cotranscribed genes has led to suggestions of attenuation or of mRNA processing as extra control devices (6,7,14,15). It has also been proposed that rpoBC may in fact have a second promoter that functions independently of the primary promoter for the rplJL-rpoBC transcript (7, 13).We have constructed hybrid plasmids between DNA fragments derived from selected segments of the rplJL-rpoBC region and the plasmid pMC81. These hybrid plasmids have been used to study the regulatory features contained in this region. MATERIALS AND METHODSBacterial Strains, Plasmids, and Enzymes. Strain HB133 (endoI-, rBMB, ara-, strA, lac-) and plasmid pBR322 (ampr, tetr) (16) were obtained from H. Boyer; strain MO1000 (r + m , recB, recC, sbcA, gal-, thi-, Atrp-tonB) was obtained from M. Oishi. Strain MC1000 (rjmj, AlacIPOZY, galK, galU, Aara-leu, strA) and plasmid pMC81 (colEl:tn3:araIO-trpBlacZ) (1)

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Section: Resultssupporting

confidence: 84%

mentioning

confidence: 99%

Control features within the rplJL-rpoBC transcription unit of Escherichia coli.

Barry¹,

Squires²

1979

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

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“…An efficiency of less than 100% could be due to (Figure 3, lane lb). When the complexes are made at a ten-fold higher ratio of enzyme to DNA (E/D = 3q), greater but still minor amounts of radioactivity are seen to migrate with Mbol-C, D, and E fragments in agreement with the Southern hybridization localization of transcription (Figure 3, lanes 2a and 2b) restriction fragments (e.g., 19,20). Conceivably such end-to-end transcription is also due to RNA polymerase core in the enzyme preparations used.…”

Section: Electrodhoresis Of Ternary Complexessupporting

confidence: 71%

Gel electrophoretic separation of transcription complexes: an assay for RNA polymerase selectivity and a method for promoter mapping

Chelm

Geiduschek

1979

Nucl Acids Res

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ABS ¶[RACI lWe describe a method for analyzing ternary transcription complexes, of RNA polymerase, DNA and nascent RN4-chains, by agarose gel electrophoresis.When the RNA of such complexes is -P-labelled, a simple comparison of the DNA fluorogram with an autoradiogram identifies transcriptionally active DNA molecules and restriction fragments in any mixture. Iwo limitations on the method are described: 1) retardation during electrophoresis of polymerase-DNA complexes relative to their conjugate bare DNA fragments; 2) failure of very large ternary complexes to enter gels.The following potential applications of the method are surveyed: transcription unit (elongation) mapping, separation of RNA molecules in a mixture of transcripts, dinucleotide primer mapping and identification of preferred template conformations. 1IiRODUCI.IONThe transcription reaction catalyzed by DNA-dependent RNA polymerase can be broken down into three steps: initiation, elongation, and termination. In vitro studies with purified hNA polymerases and auxiliary components have been crucial to furthering our understanding of the molecular mechanisms which regulate the selectivity and specificity of this reaction, at least for procaryotic systems (for review see, e.g., 1-3). In this paper we exploit the observation that transcription elongation complexes, that is, ternary complexes, of DNA, RNA polymerase and nascent RNA are stable during a variety of manipulations including electrophoresis through agarose gels. We show how this property can be used to assay transcriptional selectivity and to map transcription units with some advantages in speed and flexibility over already

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“…1 and 16 for reviews). DNA sequences for promoters for some of these operons have been determined (17,18), but no obvious common structure has been discovered that could be used to explain coordinated regulation.…”

Section: Resultsmentioning

confidence: 99%

Regulation of ribosomal protein synthesis in Escherichia coli by selective mRNA inactivation.

Fallon¹,

Jinks²,

Strycharz³

et al. 1979

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

116

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In an Escherichia coli strain lysogenic for Xspc2 transducing phage, an extra copy of ribosomal protein (r-protein) genes in the spc and a operons are carried on the phage chromosome. Expression of genes in the spc operon in this merodiploid strain was compared with that in a control "haploid" strain carrying XtrkA phage. It was found that the synthesis rate of spc mRNA, relative to other reference mRNA in the merodiploid strain, is about 2-fold higher than that in the control strain; yet, no dosage effect was observed in the synthesis rate of r-proteins in the spc or a operon. The spc mRNA was found to be more rapidly degraded in the merodiploid strain than in the control strain, and its steady-state amount, relative to reference mRNA, was only slightly higher in the merodiploid strain than in the control strain. Thus, E. coli cells have the ability to regulate the rate of r-protein synthesis regardless of the rate of transcription of r-protein genes, presumably by inactivation of the mRNA followed by its degradation. A model is proposed which involves selective inactivation of r-protein mRNA by a feedback mechanism. The model can explain coordinated synthesis of r-proteins and other observations related to selective expression of certain alleles in diploid strains.Ribosomes from Escherichia coli contain approximately 50 proteins. All of these proteins, with the exception of L7/L12, exist in a single copy per ribosome. In exponentially growing cells, there appears to be neither a significant pool of free ribosomal proteins (r-proteins) nor significant degradation of newly synthesized r-proteins. Therefore, all the r-proteins are apparently synthesized coordinately and stoichiometrically (for a review, see ref. 1). How this remarkable regulation is achieved is not known.In this paper, we report the results of experiments that show that E. coli cells have the ability to regulate the rate of r-protein synthesis regardless of the rate of transcription of r-protein genes. We then describe a new model which involves selective inactivation of r-protein mRNA by a feedback mechanism and discuss various available experimental observations. EXPERIMENTAL PROCEDURES Two E. coli K-12 strains were used. Both are lysogenic derivatives of strain N01230 (trkA401, kdpABC5, spCr, strr, fusr). One (NO1275) carries XtrkA transducing phage and XcI857S7 helper phage. The other (NO1328) carries Xspc2 transducing phage and the same helper phage. These strains and phages have been described (2, 3). All other experimental procedures are described in the figure and table legends. RESULTSWe initially asked whether an increase in the number of gene copies leads to an increase in the synthesis rate of corresponding r-proteins or if E. coli cells have the ability to regulate the synthesis rate regardless of gene copy numbers. The Xspc2 transducing phage carries 14 r-protein genes together with their promoters and 9 additional r-protein genes without their bacterial promoters (3, 4). The former 14 genes are organized into two operons ("...

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DNA sequences of promoter regions for the str and spc ribosomal protein operons in E. coli

Cited by 107 publications

References 41 publications

Control features within the rplJL-rpoBC transcription unit of Escherichia coli.

Control features within the rplJL-rpoBC transcription unit of Escherichia coli.

Gel electrophoretic separation of transcription complexes: an assay for RNA polymerase selectivity and a method for promoter mapping

Regulation of ribosomal protein synthesis in Escherichia coli by selective mRNA inactivation.

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