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ppGpp: magic beyond RNA polymerase

Nature Reviews Microbiology volume 10, pages 203212 (2012) | Download Citation


During stress, bacteria undergo extensive physiological transformations, many of which are coordinated by ppGpp. Although ppGpp is best known for enhancing cellular resilience by redirecting the RNA polymerase (RNAP) to certain genes, it also acts as a signal in many other cellular processes in bacteria. After a brief overview of ppGpp biosynthesis and its impact on promoter selection by RNAP, we discuss how bacteria exploit ppGpp to modulate the synthesis, stability or activity of proteins or regulatory RNAs that are crucial in challenging environments, using mechanisms beyond the direct regulation of RNAP activity.

Key points

  • To tolerate stress, many bacteria rely on the signalling nucleotides guanosine tetraphosphate and guanosine pentaphosphate (collectively referred to as ppGpp) to coordinate physiological adaptations.

  • A wealth of genetic, molecular and biochemical studies have determined that ppGpp and DnaK suppressor (DksA) bind directly to RNA polymerase (RNAP) to regulate promoter selection.

  • Beyond altering promoter selection by RNAP, ppGpp also modulates the expression, stability, oligomerization and activity of a variety of proteins and regulatory RNAs by direct and indirect mechanisms.

  • In addition to increasing the pool of RNAP that is available to alternative σ-factors in Escherichia coli, ppGpp increases the stability of the stationary phase σ-factor, σS, and the activity of the envelope stress σ-factor, σE.

  • By inducing expression of the Legionella pneumophila regulatory RNAs RsmY and RsmZ, ppGpp relieves carbon storage regulator (CsrA)-mediated repression of the translation of mRNAs that are important for transmission.

  • By binding to Salmonella enterica subsp. enterica serovar Typhimurium and Francisella tularensis transcription factors, ppGpp enhances oligomerization and promotes the formation of particular protein complexes, resulting in activation of virulence factor expression.

  • ppGpp binds to and inhibits the activity of a variety of enzymes — namely, polynucleotide phosphorylase of actinomycetes, lysine decarboxylase of E. coli, inosine monophosphate dehydrogenase of gammaproteobacteria and firmicutes, and the Bacillus subtilis phosphodiesterase YybT, which degrades cyclic di-AMP.

  • The synthesis of ppGpp also alters the amount or the impact of other regulatory nucleotides, including GTP (an inhibitor of the CodY repressor of firmicutes) and cyclic di-GMP (an inducer of biofilm matrix production by E. coli).

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  1. 1.

    & (p)ppGpp: still magical? Annu. Rev. Microbiol. 62, 35–51 (2008).

  2. 2.

    , , & Regulation of σ factor competition by the alarmone ppGpp. Genes Dev. 16, 1260–1270 (2002). Genetic and biochemical studies that establish the concept of σ-factor competition.

  3. 3.

    , , & ppGpp conjures bacterial virulence. Microbiol. Mol. Biol. Rev. 74, 171–199 (2010).

  4. 4.

    , & Advances in bacterial promoter recognition and its control by factors that do not bind DNA. Nature Rev. Microbiol. 6, 507–519 (2008).

  5. 5.

    , & Regulation of alternative sigma factor use. Annu. Rev. Microbiol. 65, 37–55 (2011).

  6. 6.

    , & Control of rRNA expression by small molecules is dynamic and nonredundant. Mol. Cell 12, 125–134 (2003).

  7. 7.

    , , & ppGpp is the major source of growth rate control in E. coli. Environ. Microbiol. 13, 563–575 (2011).

  8. 8.

    , & Nutritional control of elongation of DNA replication by (p)ppGpp. Cell 128, 865–875 (2007). An analysis of DNA replication in E. coli cultures and with purified components identifies direct inhibition of DNA primase activity by ppGpp.

  9. 9.

    et al. Second messenger signalling governs Escherichia coli biofilm induction upon ribosomal stress. Mol. Microbiol. 72, 1500–1516 (2009). A genetic study demonstrating synergy between the ppGpp and c-di-GMP signalling pathways.

  10. 10.

    et al. YybT is a signaling protein that contains a cyclic dinucleotide phosphodiesterase domain and a GGDEF domain with ATPase activity. J. Biol. Chem. 285, 473–482 (2010). A biochemical analysis showing that ppGpp inhibits the degradation of c-di-AMP by YybT.

  11. 11.

    Signal transduction and regulatory mechanisms involved in control of the σS (RpoS) subunit of RNA polymerase. Microbiol. Mol. Biol. Rev. 66, 373–395 (2002).

  12. 12.

    & Role of RpoS in virulence of pathogens. Infect. Immun. 78, 887–897 (2010).

  13. 13.

    Signal sensory systems that impact σ54-dependent transcription. FEMS Microbiol. Rev. 35, 425–440 (2010).

  14. 14.

    et al. Increased RNA polymerase availability directs resources towards growth at the expense of maintenance. EMBO J. 28, 2209–2219 (2009).

  15. 15.

    , & The RpoS-mediated general stress response in Escherichia coli. Annu. Rev. Microbiol. 65, 189–213 (2011).

  16. 16.

    , & The σS subunit of RNA polymerase as a signal integrator and network master regulator in the general stress response in Escherichia coli. Sci. Prog. 90, 103–127 (2007).

  17. 17.

    , & Modulating RssB activity: IraP, a novel regulator of σS stability in Escherichia coli. Genes Dev. 20, 884–897 (2006). A molecular genetics investigation which finds that stabilization of σS during phosphate starvation is mediated by SpoT-induced activation of iraP transcription.

  18. 18.

    & ppGpp regulation of RpoS degradation via anti-adaptor protein IraP. Proc. Natl Acad. Sci. USA 104, 12896–12901 (2007).

  19. 19.

    , & Growth phase and (p)ppGpp control of IraD, a regulator of RpoS stability, in Escherichia coli. J. Bacteriol. 191, 7436–7446 (2009).

  20. 20.

    , , , & A DNA damage response in Escherichia coli involving the alternative sigma factor, RpoS. Proc. Natl Acad. Sci. USA 106, 611–616 (2009).

  21. 21.

    Regulation by destruction: design of the σE envelope stress response. Curr. Opin. Microbiol. 11, 535–540 (2008).

  22. 22.

    , , , & Transcription profiling of the stringent response in Escherichia coli. J. Bacteriol. 190, 1084–1096 (2008).

  23. 23.

    & Growth phase-dependent regulation of the extracytoplasmic stress factor, σE, by guanosine 3′,5′-bispyrophosphate (ppGpp). J. Bacteriol. 188, 4627–4634 (2006).

  24. 24.

    et al. ppGpp and DksA likely regulate the activity of the extracytoplasmic stress factor σE in Escherichia coli by both direct and indirect mechanisms. Mol. Microbiol. 67, 619–632 (2008). A biochemical experiment determining that ppGpp directly stimulates σE activity, together with an in vivo analysis that indicates enhanced competition for RNAP also occurs.

  25. 25.

    “Magic Spot” Conjures Transmission of Legionella pneumophila. Thesis, Univ. Michigan (2010).

  26. 26.

    et al. Circuitry linking the Csr and stringent response global regulatory systems. Mol. Microbiol. 80, 1561–1580 (2011).

  27. 27.

    et al. Biofilm formation and dispersal under the influence of the global regulator CsrA of Escherichia coli. J. Bacteriol. 184, 290–301 (2002).

  28. 28.

    et al. Positive regulation of motility and flhDC expression by the RNA-binding protein CsrA of Escherichia coli. Mol. Microbiol. 40, 245–256 (2001).

  29. 29.

    & Legionella pneumophila CsrA is a pivotal repressor of transmission traits and activator of replication. Mol. Microbiol. 50, 445–461 (2003).

  30. 30.

    et al. The RNA binding protein CsrA is a pleiotropic regulator of the locus of enterocyte effacement pathogenicity island of enteropathogenic Escherichia coli. Infect. Immun. 77, 3552–3568 (2009).

  31. 31.

    & The LetA-RsmYZ-CsrA regulatory cascade, together with RpoS and PmrA, post-transcriptionally regulates stationary phase activation of Legionella pneumophila Icm/Dot effectors. Mol. Microbiol. 72, 995–1010 (2009).

  32. 32.

    & Determination of the regulon and identification of novel mRNA targets of Pseudomonas aeruginosa RsmA. Mol. Microbiol. 72, 612–632 (2009).

  33. 33.

    & Differentiate to thrive: lessons from the Legionella pneumophila life cycle. Mol. Microbiol. 53, 29–40 (2004).

  34. 34.

    , , & Gac/Rsm signal transduction pathway of γ-proteobacteria: from RNA recognition to regulation of social behaviour. Mol. Microbiol. 67, 241–253 (2008).

  35. 35.

    & CsrB sRNA family: sequestration of RNA-binding regulatory proteins. Curr. Opin. Microbiol. 10, 156–163 (2007).

  36. 36.

    , , , & Regulatory interactions of Csr components: the RNA binding protein CsrA activates csrB transcription in Escherichia coli. J. Bacteriol. 183, 6017–6027 (2001).

  37. 37.

    et al. A novel sRNA component of the carbon storage regulatory system of Escherichia coli. Mol. Microbiol. 48, 657–670 (2003).

  38. 38.

    et al. σS controls multiple pathways associated with intracellular multiplication of Legionella pneumophila. J. Bacteriol. 191, 2461–2473 (2009).

  39. 39.

    et al. Two small ncRNAs jointly govern virulence and transmission in Legionella pneumophila. Mol. Microbiol. 72, 741–762 (2009).

  40. 40.

    , , , & Distinct roles of ppGpp and DksA in Legionella pneumophila differentiation. Mol. Microbiol. 76, 200–219 (2010).

  41. 41.

    , , , & Identification of YhdA as a regulator of the Escherichia coli carbon storage regulation system. FEMS Microbiol. Lett. 264, 232–237 (2006).

  42. 42.

    & The PhoQ/PhoP regulatory network of Salmonella enterica. Adv. Exp. Med. Biol. 631, 7–21 (2008).

  43. 43.

    , & PhoP-responsive expression of the Salmonella enterica serovar typhimurium slyA gene. J. Bacteriol. 185, 3508–3514 (2003).

  44. 44.

    , & A two-component regulatory system (phoP phoQ) controls Salmonella typhimurium virulence. Proc. Natl Acad. Sci. USA 86, 5054–5058 (1989).

  45. 45.

    et al. A cytolysin encoded by Salmonella is required for survival within macrophages. Proc. Natl Acad. Sci. USA 91, 489–493 (1994).

  46. 46.

    & Regulation of virulence by members of the MarR/SlyA family. Curr. Opin. Microbiol. 9, 153–159 (2006).

  47. 47.

    , , & Transcriptional control of the antimicrobial peptide resistance ugtL gene by the Salmonella PhoP and SlyA regulatory proteins. J. Biol. Chem. 279, 38618–38625 (2004).

  48. 48.

    , , , , & A dual-signal regulatory circuit activates transcription of a set of divergent operons in Salmonella typhimurium. Proc. Natl Acad. Sci. USA 105, 20924–20929 (2008). Genetic and biochemical experiments which demonstrate that ppGpp binds the transcription factor SlyA and promotes its dimerization, DNA binding and activity as a transcriptional activator.

  49. 49.

    , & Salmonella sensing of anti-microbial mechanisms to promote survival within macrophages. Immunol. Rev. 219, 55–65 (2007).

  50. 50.

    et al. MglA regulates transcription of virulence factors necessary for Francisella tularensis intraamoebae and intramacrophage survival. Proc. Natl Acad. Sci. USA 101, 4246–4249 (2004).

  51. 51.

    & Subunits of RNA polymerase in function and structure. IX. Regulation of RNA polymerase activity by stringent starvation protein (SSP). J. Mol. Biol. 129, 517–530 (1979).

  52. 52.

    , & Starvation-induced expression of SspA and SspB: the effects of a null mutation in sspA on Escherichia coli protein synthesis and survival during growth and prolonged starvation. Mol. Microbiol. 11, 1029–1043 (1994).

  53. 53.

    et al. MglA regulates Francisella tularensis subsp. novicida (Francisella novicida) response to starvation and oxidative stress. J. Bacteriol. 189, 6580–6586 (2007).

  54. 54.

    , , , & Small molecule control of virulence gene expression in Francisella tularensis. PLoS Pathog 5, e1000641 (2009). Molecular genetic, biochemical and cell biological analyses determining that ppGpp binding promotes assembly and activity of a transcriptional complex which is crucial for intracellular growth.

  55. 55.

    & in Escherichia coli and Salmonella typhimurium: Cellular and Molecular Biology (eds Neidhardt, F. C. et al.) 1458–1496 (American Society for Microbiology, Washington DC, 1996).

  56. 56.

    & Control of bacterial transcription, translation and replication by (p)ppGpp. Curr. Opin. Microbiol. 11, 100–105 (2008).

  57. 57.

    et al. Guanosine 5′-diphosphate 3′-diphosphate (ppGpp) as a negative modulator of polynucleotide phosphorylase activity in a 'rare' actinomycete. Mol. Microbiol. 77, 716–729 (2010). In vitro and in vivo experiments which establish that ppGpp binds and inhibits the activity of an enzyme that destabilizes mRNAs.

  58. 58.

    & (p)ppGpp inhibits polynucleotide phosphorylase from streptomyces but not from Escherichia coli and increases the stability of bulk mRNA in Streptomyces coelicolor. J. Bacteriol. 192, 4275–4280 (2010). A biochemical study demonstrating that direct binding by ppGpp inhibits enzymes that destabilize mRNA.

  59. 59.

    et al. Linkage between the bacterial acid stress and stringent responses: the structure of the inducible lysine decarboxylase. EMBO J. 30, 931–944 (2011). The crystalography and enzymatic assays that establish ppGpp as a direct regulator of enyzme activity.

  60. 60.

    , & Inorganic polyphosphate: essential for growth and survival. Annu. Rev. Biochem. 78, 605–647 (2009).

  61. 61.

    , & Control of mRNA processing and decay in prokaryotes. Genetica 94, 157–172 (1994).

  62. 62.

    & Acid stress response in enteropathogenic gammaproteobacteria: an aptitude for survival. Biochem. Cell Biol. 88, 301–314 (2010).

  63. 63.

    & Regulation of Vibrio cholerae genes required for acid tolerance by a member of the “ToxR-like” family of transcriptional regulators. J. Bacteriol. 182, 5342–5350 (2000).

  64. 64.

    , , , & Internal pH crisis, lysine decarboxylase and the acid tolerance response of Salmonella typhimurium. Mol. Microbiol. 20, 605–611 (1996).

  65. 65.

    , , & Guanosine tetra- and pentaphosphate promote accumulation of inorganic polyphosphate in Escherichia coli. J. Biol. Chem. 272, 21240–21243 (1997).

  66. 66.

    et al. Role of inorganic polyphosphate in promoting ribosomal protein degradation by the Lon protease in E. coli. Science 293, 705–708 (2001).

  67. 67.

    , & The synthesis and function of the alarmone (p)ppGpp in firmicutes. Int. J. Med. Microbiol. 300, 142–147 (2010).

  68. 68.

    , & The mechanism of amino acid control of guanylate and adenylate biosynthesis. J. Biol. Chem. 246, 5812–5816 (1971). A seminal study demonstrating that ppGpp is a second messenger which accumulates during amino acid starvation and inhibits the activity of enzymes that initiate ATP and GTP biosynthesis.

  69. 69.

    , & Response of guanosine 5′-triphosphate concentration to nutritional changes and its significance for Bacillus subtilis sporulation. J. Bacteriol. 146, 605–613 (1981).

  70. 70.

    & An alternative strategy for bacterial ribosome synthesis: Bacillus subtilis rRNA transcription regulation. EMBO J. 23, 4473–4483 (2004). The molecular genetic and biochemical experiments which establish that the consumption of GTP during ppGpp synthesis indirectly decreases rRNA synthesis in B. subtiliis.

  71. 71.

    CodY, a global regulator of stationary phase and virulence in Gram-positive bacteria. Curr. Opin. Microbiol. 8, 203–207 (2005).

  72. 72.

    et al. Role of the (p)ppGpp synthase RSH, a RelA/SpoT homolog, in stringent response and virulence of Staphylococcus aureus. Infect. Immun. 78, 1873–1883 (2010).

  73. 73.

    , , & Linking the nutritional status of Streptococcus pyogenes to alteration of transcriptional gene expression: the action of CodY and RelA. Int. J. Med. Microbiol. 296, 259–275 (2006).

  74. 74.

    , , , & Global regulation by (p)ppGpp and CodY in Streptococcus mutans. J. Bacteriol. 190, 5291–5299 (2008).

  75. 75.

    et al. Characterization of relA and codY mutants of Listeria monocytogenes: identification of the CodY regulon and its role in virulence. Mol. Microbiol. 63, 1453–1467 (2007).

  76. 76.

    & Prevailing concepts of c-di-GMP signaling. Contrib. Microbiol. 16, 161–181 (2009).

  77. 77.

    Principles of c-di-GMP signalling in bacteria. Nature Rev. Microbiol. 7, 263–273 (2009).

  78. 78.

    et al. A host type I interferon response is induced by cytosolic sensing of the bacterial second messenger cyclic-di-GMP. J. Exp. Med. 206, 1899–1911 (2009). The first work to demonstrate that treatment with cyclic nucleotides stimulates innate immune mechanisms of mammalian cells.

  79. 79.

    , & c-di-AMP secreted by intracellular Listeria monocytogenes activates a host type I interferon response. Science 328, 1703–1705 (2010). An investigation which uses bacterial genetics to establish that the production of cyclic nucleotides by an intracellular pathogen stimulates host innate immune defence pathways.

  80. 80.

    , & Patterns of pathogenesis: discrimination of pathogenic and nonpathogenic microbes by the innate immune system. Cell Host Microbe 6, 10–21 (2009).

  81. 81.

    & Evolving concepts in biofilm infections. Cell. Microbiol. 11, 1034–1043 (2009).

  82. 82.

    et al. Aminoglycoside antibiotics induce bacterial biofilm formation. Nature 436, 1171–1175 (2005).

  83. 83.

    , , & Structural biochemistry of a bacterial checkpoint protein reveals diadenylate cyclase activity regulated by DNA recombination intermediates. Mol. Cell 30, 167–178 (2008).

  84. 84.

    Great times for small molecules: c-di-AMP, a second messenger candidate in Bacteria and Archaea. Sci. Signal 1, pe39 (2008).

  85. 85.

    et al. Discretely calibrated regulatory loops controlled by ppGpp partition gene induction across the 'feast to famine' gradient in Escherichia coli. Mol. Microbiol. 79, 830–845 (2011). A transcriptional profile analysis identifying genes that require different amounts of ppGpp for their induction, thus illustrating a dynamic range for this regulatory pathway.

  86. 86.

    et al. Selective fluorescent chemosensor for the bacterial alarmone (p)ppGpp. J. Am. Chem. Soc. 130, 784–785 (2008).

  87. 87.

    et al. Two novel point mutations in clinical Staphylococcus aureus reduce linezolid susceptibility and switch on the stringent response to promote persistent infection. PLoS Pathog. 6, e1000944 (2010).

  88. 88.

    et al. Asymmetrical distribution of the second messenger c-di-GMP upon bacterial cell division. Science 328, 1295–1297 (2010).

  89. 89.

    et al. A metazoan ortholog of SpoT hydrolyzes ppGpp and functions in starvation responses. Nature Struct. Mol. Biol. 17, 1188–1194 (2010). The first molecular and functional studies to identify stringent response components in D. melanogaster and human cells.

  90. 90.

    & The bacterial signal molecule, ppGpp, regulates Salmonella virulence gene expression. Mol. Microbiol. 52, 1827–1844 (2004).

  91. 91.

    , , , & The role of relA and spoT in Yersinia pestis KIM5 pathogenicity. PLoS ONE 4, e6720 (2009).

  92. 92.

    , & Characterization of a Legionella pneumophila relA insertion mutant and toles of RelA and RpoS in virulence gene expression. J. Bacteriol. 184, 67–75 (2002).

  93. 93.

    , & SpoT governs Legionella pneumophila differentiation in host macrophages. Mol. Microbiol. 71, 640–658 (2009).

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The authors' research on virulence regulation in L. pneumophila has been supported by the US National Institutes of Health (grant 2 R01 AI44212 to M.S.S.) and the University of Michigan, Ann Arbor, USA (a Rackham Predoctoral Fellowship to Z.D.D.).

Author information


  1. Department of Microbiology, University of Washington, Health Sciences Building K116, 1959 NE Pacific St., Box 357710, Seattle, Washington 98195-7710, USA.

    • Zachary D. Dalebroux
  2. Department of Microbiology & Immunology, University of Michigan Medical School, 6733 Medical Science Building II, 1150 West Medical Center Drive, Ann Arbor, Michigan 48109–5620, USA.

    • Michele S. Swanson


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Competing interests

The authors declare no competing financial interests.

Corresponding author

Correspondence to Michele S. Swanson.


Stringent response

A stress response coordinated by guanosine tetraphosphate and guanosine pentaphosphate, in which cells rapidly inhibit synthesis of stable RNA, ribosomes and proteins, leading to growth arrest.


In the context of post-translational regulation of protein stability, a protein that binds another protein and chaperones it to the proteasome for degradation; an example of such an adaptor is regulator of σS protein RssB.


A cytoplasmic complex that unfolds and degrades proteins.


In the context of post-translational regulation of protein stability, a protein that binds an adaptor and blocks its chaperone activity; examples of anti-adaptors are IraP and IraD.


The DNA sequence between the −10-box hexamer of a promoter and the transcriptional start site at nucleotide + 1. Promoters that are activated by guanosine tetraphosphate and guanosine pentaphosphate acting with DnaK suppressor (DksA) are typically AT-rich in this region, whereas repressed targets are typically GC-rich.

Serine hydroxamate

A structural analogue of L-serine that induces the stringent response by inhibiting charging of seryl-tRNA synthetase.


The gene encoding the β-subunit of RNA polymerase; this subunit is involved in transcription initiation through its interaction with the σ-factor. The commonly used antibiotic rifampicin targets this β-subunit to inhibit transcription, and mutations in rpoB commonly confer rifampicin resistance.


All of the genes and operons for which expression is controlled by a particular regulatory protein.

Stringent starvation protein A

A protein that is synthesized in response to amino acid starvation and associates with RNA polymerase to control transcription. It promotes survival during acid stress and nutrient limitation in bacteria.

DNA primase

An enzyme that generates short RNA primers that are elongated by DNA polymerase during chromosome replication. DNA primase is essential for the replication of chromosomes and plasmids.

Second messenger

A molecule that relays signals from a receiver or receptor by exerting an effect on a downstream cellular factor or process.

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