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Analysis of the role of Bacillus subtilis ?(M) in ?-lactam resistance reveals an essential role for c-di-AMP in peptidoglycan homeostasis.

ABSTRACT: The Bacillus subtilis extracytoplasmic function (ECF) ? factor ?(M) is inducible by, and confers resistance to, several cell envelope-acting antibiotics. Here, we demonstrate that ?(M) is responsible for intrinsic ?-lactam resistance, with ?(X) playing a secondary role. Activation of ?(M) upregulates several cell wall biosynthetic enzymes including one, PBP1, shown here to be a target for the beta-lactam cefuroxime. However, ?(M) still plays a major role in cefuroxime resistance even in cells lacking PBP1. To better define the role of ?(M) in ?-lactam resistance, we characterized suppressor mutations that restore cefuroxime resistance to a sigM null mutant. The most frequent suppressors inactivated gdpP (yybT) which encodes a cyclic-di-AMP phosphodiesterase (PDE). Intriguingly, ?(M) is a known activator of disA encoding one of three paralogous diadenylate cyclases (DAC). Overproduction of the GdpP PDE greatly sensitized cells to ?-lactam antibiotics. Conversely, genetic studies indicate that at least one DAC is required for growth with depletion leading to cell lysis. These findings support a model in which c-di-AMP is an essential signal molecule required for cell wall homeostasis. Other suppressors highlight the roles of ECF ? factors in counteracting the deleterious effects of autolysins and reactive oxygen species in ?-lactam-treated cells.

SUBMITTER: Luo Y

PROVIDER: S-EPMC3306796 | biostudies-literature | 2012 Feb

REPOSITORIES: biostudies-literature

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Analysis of the role of Bacillus subtilis σ(M) in β-lactam resistance reveals an essential role for c-di-AMP in peptidoglycan homeostasis.

Luo Yun Y Helmann John D JD

Molecular microbiology 20120104 3

The Bacillus subtilis extracytoplasmic function (ECF) σ factor σ(M) is inducible by, and confers resistance to, several cell envelope-acting antibiotics. Here, we demonstrate that σ(M) is responsible for intrinsic β-lactam resistance, with σ(X) playing a secondary role. Activation of σ(M) upregulates several cell wall biosynthetic enzymes including one, PBP1, shown here to be a target for the beta-lactam cefuroxime. However, σ(M) still plays a major role in cefuroxime resistance even in cel ...[more]

PMID: 22211522

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Project description:In order to adjust to changing environmental conditions, bacteria use nucleotide second messengers to transduce external signals and translate them into a specific cellular response. Cyclic di-adenosine monophosphate (c-di-AMP) is the only known essential nucleotide second messenger. In addition to the well-established role of this second messenger in the control of potassium homeostasis, we observed that glutamate is as toxic as potassium for a c-di-AMP-free strain of the Gram-positive model bacterium Bacillus subtilis. In this work, we isolated suppressor mutants that allow growth of a c-di-AMP-free strain under these toxic conditions. Characterization of glutamate resistant suppressors revealed that they contain pairs of mutations, in most cases affecting glutamate and potassium homeostasis. Among these mutations, several independent mutations affected a novel glutamate transporter, AimA (Amino acid importer A, formerly YbeC). This protein is the major transporter for glutamate and serine in B. subtilis. Unexpectedly, some of the isolated suppressor mutants could suppress glutamate toxicity by a combination of mutations that affect phospholipid biosynthesis and a specific gain-of-function mutation of a mechanosensitive channel of small conductance (YfkC) resulting in the acquisition of a device for glutamate export. Cultivation of the c-di-AMP-free strain on complex medium was an even greater challenge because the amounts of potassium, glutamate, and other osmolytes are substantially higher than in minimal medium. Suppressor mutants viable on complex medium could only be isolated under anaerobic conditions if one of the two c-di-AMP receptor proteins, DarA or DarB, was absent. Also on complex medium, potassium and osmolyte toxicity are the major bottlenecks for the growth of B. subtilis in the absence of c-di-AMP. Our results indicate that the essentiality of c-di-AMP in B. subtilis is caused by the global impact of the second messenger nucleotide on different aspects of cellular physiology.

Dataset Information

Analysis of the role of Bacillus subtilis ?(M) in ?-lactam resistance reveals an essential role for c-di-AMP in peptidoglycan homeostasis.

Publications

Analysis of the role of Bacillus subtilis σ(M) in β-lactam resistance reveals an essential role for c-di-AMP in peptidoglycan homeostasis.

Similar Datasets

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