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DksA potentiates direct activation of amino acid promoters by ppGpp.

ABSTRACT: Amino acid starvation in Escherichia coli results in a spectrum of changes in gene expression, including inhibition of rRNA and tRNA promoters and activation of certain promoters for amino acid biosynthesis and transport. The unusual nucleotide ppGpp plays an important role in both negative and positive regulation. Previously, we and others suggested that positive effects of ppGpp might be indirect, resulting from the inhibition of rRNA transcription and, thus, liberation of RNA polymerase for binding to other promoters. Recently, we showed that DksA binds to RNA polymerase and greatly enhances direct effects of ppGpp on the negative control of rRNA promoters. This conclusion prompted us to reevaluate whether ppGpp might also have a direct role in positive control. We show here that ppGpp greatly increases the rate of transcription initiation from amino acid promoters in a purified system but only when DksA is present. Activation occurs by stimulation of the rate of an isomerization step on the pathway to open complex formation. Consistent with the model that ppGpp/DksA stimulates amino acid promoters both directly and indirectly in vivo, cells lacking dksA fail to activate transcription from the hisG promoter after amino acid starvation. Our results illustrate how transcription factors can positively regulate transcription initiation without binding DNA, demonstrate that dksA directly affects promoters in addition to those for rRNA, and suggest that some of the pleiotropic effects previously associated with dksA might be ascribable to direct effects of dksA on promoters involved in a wide variety of cellular functions.

SUBMITTER: Paul BJ

PROVIDER: S-EPMC1142371 | biostudies-literature | 2005 May

REPOSITORIES: biostudies-literature

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DksA potentiates direct activation of amino acid promoters by ppGpp.

Paul Brian J BJ Berkmen Melanie B MB Gourse Richard L RL

Proceedings of the National Academy of Sciences of the United States of America 20050517 22

Amino acid starvation in Escherichia coli results in a spectrum of changes in gene expression, including inhibition of rRNA and tRNA promoters and activation of certain promoters for amino acid biosynthesis and transport. The unusual nucleotide ppGpp plays an important role in both negative and positive regulation. Previously, we and others suggested that positive effects of ppGpp might be indirect, resulting from the inhibition of rRNA transcription and, thus, liberation of RNA polymerase for b ...[more]

PMID: 15899978

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Project description:?S is an alternative sigma factor, encoded by the rpoS gene, that redirects cellular transcription to a large family of genes in response to stressful environmental signals. This so-called ?S general stress response is necessary for survival in many bacterial species and is controlled by a complex, multifactorial pathway that regulates ?S levels transcriptionally, translationally, and posttranslationally in Escherichia coli It was shown previously that the transcription factor DksA and its cofactor, ppGpp, are among the many factors governing ?S synthesis, thus playing an important role in activation of the ?S stress response. However, the mechanisms responsible for the effects of DksA and ppGpp have not been elucidated fully. We describe here how DksA and ppGpp directly activate the promoters for the anti-adaptor protein IraP and the small regulatory RNA DsrA, thereby indirectly influencing ?S levels. In addition, based on effects of DksAN88I, a previously identified DksA variant with increased affinity for RNA polymerase (RNAP), we show that DksA can increase ?S activity by another indirect mechanism. We propose that by reducing rRNA transcription, DksA and ppGpp increase the availability of core RNAP for binding to ?S and also increase transcription from other promoters, including PdsrA and PiraP By improving the translation and stabilization of ?S, as well as the ability of other promoters to compete for RNAP, DksA and ppGpp contribute to the switch in the transcription program needed for stress adaptation.IMPORTANCE Bacteria spend relatively little time in log phase outside the optimized environment found in a laboratory. They have evolved to make the most of alternating feast and famine conditions by seamlessly transitioning between rapid growth and stationary phase, a lower metabolic mode that is crucial for long-term survival. One of the key regulators of the switch in gene expression that characterizes stationary phase is the alternative sigma factor ?S Understanding the factors governing ?S activity is central to unraveling the complexities of growth, adaptation to stress, and pathogenesis. Here, we describe three mechanisms by which the RNA polymerase binding factor DksA and the second messenger ppGpp regulate ?S levels.

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DksA potentiates direct activation of amino acid promoters by ppGpp.

Publications

DksA potentiates direct activation of amino acid promoters by ppGpp.

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