Evolution and Function of the RelA/SpoT Homologue (RSH) Proteins

Abstract

RelA/SpoT Homologue (RSH) proteins comprise a superfamily of enzymes that synthesise and/or hydrolyse the nucleotide alarmone ppGpp, mediator of the ‘stringent’ response and regulator of cellular metabolism in response to changing environmental conditions. Most of what we know about RSHs comes from the Escherichia coli proteins RelA and SpoT, paralogues that appear to have evolved from duplication of an ancestral Rel gene in the lineage to β‐ and γ‐proteobacteria. Recently, more details have come to light on the evolution of the RSH superfamily, revealing that there is a much greater diversity of RSHs than previously thought.

Key Concepts:

  • Various stress conditions (amino acid, iron and fatty acid starvation, heat shock, etc.) induce the so‐called ‘stringent response’ in bacteria and chloroplasts.

  • The stringent response is mediated by RelA/SpoT Homologue (RSH) proteins that modulate intracellular concentration of the ppGpp alarmone nucleotide.

  • ppGpp exerts its regulatory role by binding and modulating activity of several targets: the RNA polymerase, translational GTPases EF‐G and IF2, lysine decarboxylase Ldc1, polynucleotide phosphorylase, DnaG primase and others.

  • RelA senses amino acid starvation by directly interacting with the 70S ribosome and inspecting the aminoacylation status of the A‐site tRNA, and responds to the presence of deacylated tRNA by synthesising ppGpp.

  • SpoT is a bifunctional enzyme that has both ppGpp synthetic and hydrolytic activities and senses several cues that modulate its net activity.

  • Phylogenetic analyses divide the RSH protein family into 30 subgroups comprising three groups: long RSHs (such as RelA and SpoT), small alarmone synthetases (SASs) and small alarmone hydrolases (SAHs).

  • In addition to bacteria and chloroplasts, RSH proteins have been identified in eukaryotes and isolated species of archaea, however the ppGpp‐mediated stringent response has not yet been identified in these organisms.

  • In eukaryotes, amino acid starvation is sensed by the general amino acid control (GAAC) system that is nonhomologous to the RSH system but is functionally analogous.

Keywords: RelA; SpoT; Rel; ppGpp; stringent response; ribosome; SAS; SAH; RSH; amino acid starvation

Figure 1.

Domain structure of RSHs. In Rsh4, cTP and EFh stand for chloroplast transit peptide and EF hand domain, respectively. Abbreviations for the other RSHs are as discussed in the text.

Figure 2.

Schematic diagram for the evolution of long RSHs in bacteria. Thick grey branches indicate the divergence of bacterial groups, whereas the inner line shows the divergence of long RSH proteins and their functionality, as per the inset box. Reproduced with permission from Atkinson et al..

Figure 3.

Phylogenetic trees of the RSH ppGpp synthetase and hydrolase domains. Trees were generated from maximum likelihood phylogenetic analyses: (a) ppGpp hydrolase (HD) domain‐containing RSHs, and (b) the ppGpp synthetase (SYNTH) domain‐containing RSHs. In both trees, subgroups are labelled and shading behind the branches shows the most common domain structure observed for those groups, as per the legend in the inset box. Symbols on branches indicate bootstrap support, as per the inset box. Branch length is proportional to the number of substitutions per site (see scale bar). Reproduced with permission from Atkinson et al..

close

References

Alon U (2007a) An Introduction to Systems Biology: Design Principles of Biological Circuits. London: Chapman & Hall.

Alon U (2007b) Network motifs: theory and experimental approaches. Nature Reviews Genetics 8: 450–461.

Atkinson GC, Tenson T and Hauryliuk V (2011) The RelA/SpoT homolog (RSH) superfamily: distribution and functional evolution of ppGpp synthetases and hydrolases across the tree of life. PLoS ONE 6: e23479.

Battesti A and Bouveret E (2006) Acyl carrier protein/SpoT interaction, the switch linking SpoT‐dependent stress response to fatty acid metabolism. Molecular Microbiology 62: 1048–1063.

Braeken K, Moris M, Daniels R, Vanderleyden J and Michiels J (2006) New horizons for (p)ppGpp in bacterial and plant physiology. Trends in Microbiology 14: 45–54.

Cashel M and Gallant J (1969) Two compounds implicated in the function of the RC gene of E. coli. Nature 221: 838–841.

Cellini A, Scoarughi GL, Poggiali P et al. (2004) Stringent control in the archaeal genus Sulfolobus. Research in Microbiology 155: 98–104.

Chandrangsu P, Lemke JJ and Gourse RL (2011) The dksA promoter is negatively feedback regulated by DksA and ppGpp. Molecular Microbiology 80: 1337–1348.

Christiansen L and Neirhaus KH (1976) Ribosomal proteins of Escherichia coli that stimulate stringent‐factor‐mediated pyrophosphoryl transfer in vitro. Proceedings of the National Academy of Sciences of the USA 73: 1839–1843.

Dalebroux ZD, Svensson SL, Gaynor EC and Swanson MS (2010) ppGpp conjures bacterial virulence. Microbiology and Molecular Biology Reviews 74: 171–199.

Das B, Pal RR, Bag S and Bhadra RK (2009) Stringent response in Vibrio cholerae: genetic analysis of spoT gene function and identification of a novel (p)ppGpp synthetase gene. Molecular Microbiology 72: 380–398.

Edwards AN, Patterson‐Fortin LM, Vakulskas CA et al. (2011) Circuitry linking the Csr and stringent response global regulatory systems. Molecular Microbiology 80: 1561–1580.

English BP, Hauryliuk V, Sanamrad A et al. (2011) Single‐molecule investigations of the stringent response machinery in living bacterial cells. Proceedings of the National Academy of Sciences of the USA 108: E359–364.

Flardh K, Axberg T, Albertson NH and Kjelleberg S (1994) Stringent control during carbon starvation of marine Vibrio sp. strain S14: molecular cloning, nucleotide sequence, and deletion of the relA gene. Journal of Bacteriology 176: 5949–5957.

Gallant J, Palmer L and Pao CC (1977) Anomalous synthesis of ppGpp in growing cells. Cell 11: 181–185.

Gatewood ML and Jones GH (2010) (p)ppGpp inhibits polynucleotide phosphorylase from Streptomyces but not from E. coli and increases the stability of bulk mRNA in Streptomyces coelicolor. Journal of Bacteriology 192: 4275–4280.

Givens RM, Lin MH, Taylor DJ et al. (2004) Inducible expression, enzymatic activity, and origin of higher plant homologues of bacterial RelA/SpoT stress proteins in Nicotiana tabacum. Journal of Biological Chemistry 279: 7495–7504.

Haseltine W and Block R (1973) Synthesis of guanosine tetra‐ and pentaphosphate requires the presence of a codon‐specific, uncharged transfer ribonucleic acid in the acceptor site of ribosomes. Proceedings of the National Academy of Sciences of the USA 70: 1564–1568.

Hinnebusch AG (2005) Translational regulation of GCN4 and the general amino acid control of yeast. Annual Review of Microbiology 59: 407–450.

Hogg T, Mechold U, Malke H, Cashel M and Hilgenfeld R (2004) Conformational antagonism between opposing active sites in a bifunctional RelA/SpoT homolog modulates (p)ppGpp metabolism during the stringent response [corrected]. Cell 117: 57–68.

Innan H and Kondrashov F (2010) The evolution of gene duplications: classifying and distinguishing between models. Nature Reviews Genetics 11: 97–108.

Irr JD, Kaulenas MS and Unsworth BR (1974) Synthesis of ppGpp by mouse embryonic ribosomes. Cell 3: 249–253.

Jain R, Rivera M and Lake J (1999) Horizontal gene transfer among genomes: the complexity hypothesis. Proceedings of the National Academy of Sciences of the USA 96: 3801–3806.

Jain V, Saleem‐Batcha R and Chatterji D (2007) Synthesis and hydrolysis of pppGpp in Mycobacteria: a ligand mediated conformational switch in Rel. Biophysical Chemistry 127: 41–50.

Jiang M, Sullivan SM, Wout PK and Maddock JR (2007) G‐protein control of the ribosome‐associated stress response protein SpoT. Journal of Bacteriology 189: 6140–6147.

Kanjee U, Gutsche I, Alexopoulos E et al. (2011) Linkage between the bacterial acid stress and stringent responses: the structure of the inducible lysine decarboxylase. EMBO Journal 30: 931–944.

Kasai K, Kanno T, Endo Y, Wakasa K and Tozawa Y (2004) Guanosine tetra‐ and pentaphosphate synthase activity in chloroplasts of a higher plant: association with 70S ribosomes and inhibition by tetracycline. Nucleic Acids Research 32: 5732–5741.

Laffler T and Gallant JA (1974) Stringent control of protein synthesis in E. coli. Cell 3: 47–49.

Lemke JJ, Sanchez‐Vazquez P, Burgos HL et al. (2011) Direct regulation of Escherichia coli ribosomal protein promoters by the transcription factors ppGpp and DksA. Proceedings of the National Academy of Sciences of the USA 108(14): 5712–5717.

Lemos JA, Lin VK, Nascimento MM, Abranches J and Burne RA (2007) Three gene products govern (p)ppGpp production by Streptococcus mutants. Molecular Microbiology 65: 1568–1581.

Ma W and Berkowitz GA (2011) Ca2+ conduction by plant cyclic nucleotide gated channels and associated signaling components in pathogen defense signal transduction cascades. New Phytology 190: 566–572.

Maciag M, Kochanowska M, Lyzen R, Wegrzyn G and Szalewska‐Palasz A (2010) ppGpp inhibits the activity of E. coli DnaG primase. Plasmid 63: 61–67.

Martini O, Irr J and Richter D (1977) Questioning of reported evidence for guanosine tetraphosphate synthesis in a ribosome system from mouse embryos. Cell 12: 1127–1131.

Mechold U, Murphy H, Brown L and Cashel M (2002) Intramolecular regulation of the opposing (p)ppGpp catalytic activities of Rel(Seq), the Rel/Spo enzyme from Streptococcus equisimilis. Journal of Bacteriology 184: 2878–2888.

Mitkevich VA, Ermakov A, Kulikova AA et al. (2010) Thermodynamic characterization of ppGpp binding to EF‐G or IF2 and of initiator tRNA binding to free IF2 in the presence of GDP, GTP, or ppGpp. Journal of Molecular Biology 402: 838–846.

Mittenhuber G (2001) Comparative genomics and evolution of genes encoding bacterial (p)ppGpp synthetases/hydrolases (the Rel, RelA and SpoT proteins). Journal of Molecular Microbiology and Biotechnology 3: 585–600.

Nanamiya H, Kasai K, Nozawa A et al. (2008) Identification and functional analysis of novel (p)ppGpp synthetase genes in Bacillus subtilis. Molecular Microbiology 67: 291–304.

Ohno S (1970) Evolution by Gene Duplication. New York: Springer.

Ooga T, Ohashi Y, Kuramitsu S et al. (2009) Degradation of ppGpp by nudix pyrophosphatase modulates the transition of growth phase in the bacterium Thermus thermophilus. Journal of Biological Chemistry 284: 15549–15556.

Parker J, Watson RJ and Friesen JD (1976) A relaxed mutant with an altered ribosomal protein L11. Molecular and General Genetics 144: 111–114.

Paul BJ, Barker MM, Ross W et al. (2004) DksA: a critical component of the transcription initiation machinery that potentiates the regulation of rRNA promoters by ppGpp and the initiating NTP. Cell 118: 311–322.

Paul BJ, Berkmen MB and Gourse RL (2005) DksA potentiates direct activation of amino acid promoters by ppGpp. Proceedings of the National Academy of Sciences of the USA 102: 7823–7828.

Pesavento C and Hengge R (2009) Bacterial nucleotide‐based second messengers. Current Opinion in Microbiology 12: 170–176.

Potrykus K, Murphy H, Philippe N and Cashel M (2011) ppGpp is the major source of growth rate control in E. coli. Environmental Microbiology 13: 563–575.

Ray JC, Tabor JJ and Igoshin OA (2011) Non‐transcriptional regulatory processes shape transcriptional network dynamics. Nature Reviews Microbiology 9: 817–828.

Sarubbi E, Rudd KE, Xiao H et al. (1989) Characterization of the spoT gene of E. coli. Journal of Biological Chemistry 264: 15074–15082.

Smits WK, Kuipers OP and Veening JW (2006) Phenotypic variation in bacteria: the role of feedback regulation. Nature Reviews Microbiology 4: 259–271.

Spira B, Silberstein N and Yagil E (1995) Guanosine 3′,5′‐bispyrophosphate (ppGpp) synthesis in cells of E. coli starved for Pi. Journal of Bacteriology 177: 4053–4058.

Sprinzl M and Richter D (1976) Free 3′‐OH group of the terminal adenosine of the tRNA molecule is essential for the synthesis in vitro of guanosine tetraphosphate and pentaphosphate in a ribosomal system from E. coli. European Journal of Biochemistry 71: 171–176.

Sun D, Lee G, Lee JH et al. (2010) A metazoan ortholog of SpoT hydrolyzes ppGpp and functions in starvation responses. Nature Structural & Molecular Biology 17: 1188–1194.

Sun J, Hesketh A and Bibb M (2001) Functional analysis of relA and rshA, two relA/spoT homologues of Streptomyces coelicolor A3(2). Journal of Bacteriology 183: 3488–3498.

Sureka K, Ghosh B, Dasgupta A et al. (2008) Positive feedback and noise activate the stringent response regulator rel in mycobacteria. PLoS ONE 3: e1771.

Tozawa Y, Nozawa A, Kanno T et al. (2007) Calcium‐activated (p)ppGpp synthetase in chloroplasts of land plants. Journal of Biological Chemistry 282: 35536–35545.

van der Biezen EA, Sun J, Coleman MJ, Bibb MJ and Jones JD (2000) Arabidopsis RelA/SpoT homologs implicate (p)ppGpp in plant signaling. Proceedings of the National Academy of Sciences of the USA 97: 3747–3752.

Vazquez de Aldana CR, Marton MJ and Hinnebusch AG (1995) GCN20, a novel ATP binding cassette protein, and GCN1 reside in a complex that mediates activation of the eIF‐2 alpha kinase GCN2 in amino acid‐starved cells. EMBO Journal 14: 3184–3199.

Vercruysse M, Fauvart M, Jans A et al. (2011) Stress response regulators identified through genome‐wide transcriptome analysis of the (p)ppGpp‐dependent response in Rhizobium etli. Genome Biology 12: R17.

Vinella D, Albrecht C, Cashel M and D'Ari R (2005) Iron limitation induces SpoT‐dependent accumulation of ppGpp in E. coli. Molecular Microbiology 56: 958–970.

Vrentas CE, Gaal T, Berkmen MB et al. (2008) Still looking for the magic spot: the crystallographically defined binding site for ppGpp on RNA polymerase is unlikely to be responsible for rRNA transcription regulation. Journal of Molecular Biology 377: 551–564.

Wek RC, Jackson BM and Hinnebusch AG (1989) Juxtaposition of domains homologous to protein kinases and histidyl‐tRNA synthetases in GCN2 protein suggests a mechanism for coupling GCN4 expression to amino acid availability. Proceedings of the National Academy of Sciences of the USA 86: 4579–4583.

Wendrich T, Blaha G, Wilson D, Marahiel M and Nierhaus K (2002) Dissection of the mechanism for the stringent factor RelA. Molecular Cell 10: 779–788.

Xiao H, Kalman M, Ikehara K et al. (1991) Residual guanosine 3′,5′‐bispyrophosphate synthetic activity of relA null mutants can be eliminated by spoT null mutations. Journal of Biological Chemistry 266: 5980–5990.

Further Reading

Cashel M, Gentry DR, Hernandez VJ and Vinella D (1996) The stringent response. In: Neidhardt FC (ed.) E. coli and Salmonella: Cellular and Molecular Biology, 2nd edn, pp. 1458–1496. Washington, DC: ASM Press.

Potrykus K and Cashel M (2008) (p)ppGpp: still magical? Annual Review of Microbiology 62: 35–51.

Contact Editor close
Submit a note to the editor about this article by filling in the form below.

* Required Field

How to Cite close
Atkinson, Gemma C, and Hauryliuk, Vasili(Feb 2012) Evolution and Function of the RelA/SpoT Homologue (RSH) Proteins. In: eLS. John Wiley & Sons Ltd, Chichester. http://www.els.net [doi: 10.1002/9780470015902.a0023959]