The nucleotide (p)ppGpp is critical for stress resistance and antibiotic tolerance. In the Gram-negative bacterium Escherichia coli, (p)ppGpp interacts with RNA polymerase to reprogram transcription upon stress. However, in Firmicutes, a major class of Gram- positive bacteria comprised of pathogens (Staphylococcus, Enterococcus, Listeria, Bacillus anthracis) and beneficial microbes, how (p)ppGpp mediates stress resistance remains poorly understood. The central hypothesis of this proposal is that, in Firmicutes, (p)ppGpp regulates GTP levels to adjust pleiotropic aspects of cellular metabolism to the external environment, thus maintaining homeostasis and facilitating antibiotic tolerance. Our recent work with the bacterium Bacillus subtilis, a widely studied, genetically amenable model for Firmicutes, lends strong support for this hypothesis. Integrating genomic and metabolomic approaches with genetics and biochemistry, we identified enzymes in the GTP biosynthesis pathway as major targets for (p)ppGpp action, and unmasked a novel GTP-(p)ppGpp feedback loop that is critical for cellular viability. Our discoveries thus provide a previously unrecognized view of (p)ppGpp synthesis and function in B. subtilis.
We aim to build on this foundation to develop a comprehensive model of stress-management via (p)ppGpp-GTP in B. subtilis that our data predicts will be applicable to Firmicutes and beyond.
The specific aims are: 1) Characterize how (p)ppGpp regulates its targets Gmk and HprT;2) Elucidate how (p)ppGpp controls GTP feedback and glycolysis;and 3) Define how (p)ppGpp contributes to antibiotic tolerance.
We are investigating how Gram-positive bacteria, including many pathogens, use the signaling nucleotide (p)ppGpp for tolerating antibiotic treatment. The proposed study helps us to find strategies to prevent (p)ppGpp-mediated drug tolerance and potentiate the efficacy of antimicrobial treatment.
|Li, Wenting; Bouveret, Emmanuelle; Zhang, Yan et al. (2016) Effects of amino acid starvation on RelA diffusive behavior in live Escherichia coli. Mol Microbiol 99:571-85|
|Gaca, Anthony O; Kudrin, Pavel; Colomer-Winter, Cristina et al. (2015) From (p)ppGpp to (pp)pGpp: Characterization of Regulatory Effects of pGpp Synthesized by the Small Alarmone Synthetase of Enterococcus faecalis. J Bacteriol 197:2908-19|
|Liu, Kuanqing; Bittner, Alycia N; Wang, Jue D (2015) Diversity in (p)ppGpp metabolism and effectors. Curr Opin Microbiol 24:72-9|
|Liu, Kuanqing; Myers, Angela R; Pisithkul, Tippapha et al. (2015) Molecular mechanism and evolution of guanylate kinase regulation by (p)ppGpp. Mol Cell 57:735-49|
|Zhang, Yan; Mooney, Rachel A; Grass, Jeffrey A et al. (2014) DksA guards elongating RNA polymerase against ribosome-stalling-induced arrest. Mol Cell 53:766-78|
|Kriel, Allison; Brinsmade, Shaun R; Tse, Jessica L et al. (2014) GTP dysregulation in Bacillus subtilis cells lacking (p)ppGpp results in phenotypic amino acid auxotrophy and failure to adapt to nutrient downshift and regulate biosynthesis genes. J Bacteriol 196:189-201|
|Maduike, Nkabuije Z; Tehranchi, Ashley K; Wang, Jue D et al. (2014) Replication of the Escherichia coli chromosome in RNase HI-deficient cells: multiple initiation regions and fork dynamics. Mol Microbiol 91:39-56|
|Bittner, Alycia N; Kriel, Allison; Wang, Jue D (2014) Lowering GTP level increases survival of amino acid starvation but slows growth rate for Bacillus subtilis cells lacking (p)ppGpp. J Bacteriol 196:2067-76|
|Arjes, Heidi A; Kriel, Allison; Sorto, Nohemy A et al. (2014) Failsafe mechanisms couple division and DNA replication in bacteria. Curr Biol 24:2149-55|
|Denapoli, Jessica; Tehranchi, Ashley K; Wang, Jue D (2013) Dose-dependent reduction of replication elongation rate by (p)ppGpp in Escherichia coli and Bacillus subtilis. Mol Microbiol 88:93-104|
Showing the most recent 10 out of 17 publications