ppApp alters transcriptional selectivity of <i>Escherichia coli</i> RNA polymerase

Travers, Andrew

doi:10.1016/0014-5793(78)80973-9

Cited by 21 publications

(13 citation statements)

References 16 publications

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“…To date, there has been only a single report of ppApp effects on transcription [30]. This report had suggested ppApp activates transcription of rRNA in a salt dependent manner, however this was done with phage lambda genomic DNA templates containing an rrn o peron rather than an isolated promoter and cellular extracts containing RNAP and other cellular factors; in addition, pppApp has not been studied.…”

Section: Resultsmentioning

confidence: 99%

Structure-function comparisons of (p)ppApp vs (p)ppGpp for Escherichia coli RNA polymerase binding sites and for rrnB P1 promoter regulatory responses in vitro

Bruhn-Olszewska

Molodtsov

Sobala

et al. 2018

Biochimica et Biophysica Acta (BBA) - Gene Regulatory Mechanism

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Precise regulation of gene expression is crucial for bacteria to respond to changing environmental conditions. In addition to protein factors affecting RNA polymerase (RNAP) activity, second messengers play an important role in transcription regulation, such as well-known effectors of the stringent response: guanosine 5'triphosphate-3'diphosphate and guanosine 3', 5'-bis(diphosphate) [(p)ppGpp]. Although much is known about importance of the 5' and 3' moieties of (p)ppGpp, the role of the guanine base remains somewhat cryptic. Here, we use (p)ppGpp's adenine analogs [(p)ppApp] to investigate how the nucleobase contributes to determine its binding site and transcriptional regulation. We determined X-ray crystal structure of Escherichia coli RNAP-(p)ppApp complex, which shows the analogs bind near the active site and switch regions of RNAP. We have also explored the regulatory effects of (p)ppApp on transcription initiating from the well-studied E. coli rrnB P1 promoter to assess and compare properties of (p)ppApp with (p)ppGpp. We demonstrate that contrary to (p)ppGpp, (p)ppApp activates transcription at this promoter and DksA hinders this effect. Moreover, pppApp exerts a stronger effect than ppApp. We also show that when ppGpp and pppApp are present together, the outcome depends on which one of them was pre-incubated with RNAP first. This behavior suggests a surprising Yin-Yang like reciprocal plasticity of RNAP responses at a single promoter, occasioned simply by pre-exposure to one or the other nucleotide. Our observations underscore the importance of the (p)ppNpp's purine nucleobase for interactions with RNAP, which may lead to a better fundamental understanding of (p)ppGpp regulation of RNAP activity.

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Section: Resultsmentioning

confidence: 99%

Structure-function comparisons of (p)ppApp vs (p)ppGpp for Escherichia coli RNA polymerase binding sites and for rrnB P1 promoter regulatory responses in vitro

Bruhn-Olszewska

Molodtsov

Sobala

et al. 2018

Biochimica et Biophysica Acta (BBA) - Gene Regulatory Mechanism

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“…Enzymes capable of synthesising (p)ppApp have been identified in the actinomycetes 12 and high levels of (p)ppApp accumulate in sporulating B. subtilis cells in a ribosome-dependent fashion 13 . A putative function for this nucleotide was identified in the 1970s, where ppApp was shown to positively affect transcription of rRNA in vitro, in contrast to the actions of (pp)pGpp 58 . In the last few years there has been a resurgence in interest in (p)ppApp.…”

Section: (Pp)pappmentioning

confidence: 99%

The stringent response and physiological roles of (pp)pGpp in bacteria

2020

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Response of guanosine 5'-triphosphate concentration to nutritional changes and its significance for Bacillus subtilis sporulation. J Bacteriol 146, 605-613, (1981). 88 Kriel, A. et al. Direct regulation of GTP homeostasis by (p)ppGpp: a critical component of viability and stress resistance. Mol Cell 48, 231-241, (2012). 89 Osaka, N. et al. Novel (p)ppGpp(0) suppressor mutations reveal an unexpected link between methionine catabolism and GTP synthesis in Bacillus subtilis. Mol

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“…In the ribosome-dependent stringent factor system from E. coli or B. subtilis the inhibitory effect of ppApp was less pronounced; 50 % reduction of ppGpp synthesis was observed at a concentration of 2 m M ppApp. Recently Travers [18] suggested that ppApp may act as an antagonist to ppGpp at the transcriptional level in E. coli; these results are not readily compatible with earlier reports that ppApp, like ppGpp, stimulates the induction of tryptophanase in E. coli [20]. An ATP-nucleotide pyrophosphate transferase activity has been isolated from various microorganisms including a number of Streptomyces strains [15,.…”

Section: The P P Gpp-syn Thesizing Systemmentioning

confidence: 96%

The Guanosine 3′,5′‐Bis(diphosphate) (ppGpp) Cycle

Richter

Fehr

Harder

1979

European Journal of Biochemistry

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The metabolism of guanosine 3',5'-bis(diphosphate) (ppcpp) has been studied in a number of different microbial species including Escherichia coli, Bacillus stearothermophilus, Bacillus subtilis and Bacillus brevis. The results can be summarized as follows.1. Generally, in the E. coli and B. subtilis systems, a stringent factor catalyses the ppGpp-synthesizing reaction only when activated by uncharged tRNA codon-specifically bound to ribosomes. in the B. stearothermophilus and B. brevis systems the stringent factor does not require activation by ribosomes. The physiological significance of this is reflected by the subcellular distribution : stringent factors requiring ribosomes for activity are exclusively found in ribosomal fractions, while those functioning independently of ribosomes are preferentially present in the high-speed supernatant fraction. Formation of ppGpp is quite specific in that neither ATP as pyrophosphate donor nor GDP (or GTP) as acceptor can be replaced by other nucleotides. However some nucleotides, particularly ppApp, effectively inhibit ppGpp production, while others, e. g. ppGpp or A(5')p4(5')A, are not inhibitory.2. Degradation of ppGpp is catalyzed by a guanosine 3',5'-bis(diphosphate) 3'-pyrophosphohydrolase present in the ribosomal fraction which specifically releases pyrophosphate from the 3' position of ppGpp. The activity is apparently associated with ribosomes and, unlike stringent factor, is not completely released under low-ionic-strength conditions. The enzyme has a molecular weight of 65 000-70000 and forms dimers and tetramers having molecular weights of 130 000 and 260000, respectively. It can also be released from ribosomes by 1 M potassium acetate extraction but then is insoluble in buffers of low ionic strength except in the presence of urea. It is inhibited by tRNA regardless of whether or not ribosomes are present.

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ppApp alters transcriptional selectivity of Escherichia coli RNA polymerase

Cited by 21 publications

References 16 publications

Structure-function comparisons of (p)ppApp vs (p)ppGpp for Escherichia coli RNA polymerase binding sites and for rrnB P1 promoter regulatory responses in vitro

Structure-function comparisons of (p)ppApp vs (p)ppGpp for Escherichia coli RNA polymerase binding sites and for rrnB P1 promoter regulatory responses in vitro

The stringent response and physiological roles of (pp)pGpp in bacteria

The Guanosine 3′,5′‐Bis(diphosphate) (ppGpp) Cycle

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