What are Alarmones?

Alarmones are nucleotide-based second messengers that respond to environmental changes in bacteria and plant chloroplasts. The nucleotides guanosine tetraphosphate (ppGpp) and guanosine pentaphosphate (pppGpp) are called alarmones, collectively known as (p)ppGpp.

In bacteria and plant chloroplasts, alarmones globally reprogram cellular physiology during cellular stress. Special enzymes belonging to the RelA/SpoT homology (RSH) family synthesize (p)ppGpp by transferring pyrophosphate from ATP to GDP or GTP. Alarmones are known as regulatory metabolites of the “stringent response.” The stringent response is characterized by growth arrest and modulation of gene expression during nutritional stresses.

Figure 1: Structure and model of (p)ppGpp.

Historically, the stringent response was identified by the rapid downregulation of stable RNA (rRNA and tRNA) genes when cells encountered amino acid starvation, resulting in global genetic and physiological changes in cellular metabolism.

(p)ppGpp has two major effects:

(i)     Modification of gene transcription, and

(ii)    Direct interaction with target proteins. 

The alarmone 3’,5’-(bis)pyrophosphate (ppGpp) shuts down transcription in starving bacteria. This stringent response helps them to conserve energy and allows survival of bacteria in adverse conditions. In recent years the molecular mechanisms of (p)ppGpp metabolism and (p)ppGpp-mediated regulation have been studied in more detail. More recently Kamarthapu et al. showed that the alarmone ppGpp is also essential for DNA repair. The researchers reported that ppGpp couples transcription elongation to the nucleotide excision repair pathway by backtracking RNA polymerase away from the DNA damage site and by also inhibiting DNA replication. The final effect is that ppGpp prompts transitions between repair and recovery states in bacteria.

Bacteria have sensory systems for monitoring their environment allowing adaption to stressful conditions. External stimuli are converted into changes in intracellular concentrations of secondary messenger molecules. Bacteria contain three common nucleotide-based secondary messengers: cAMP, c-di-GMP and (p)ppGpp. Various stress conditions are mediated by amino acid starvation, iron and fatty acid starvation, heat shock, and others that induce the stringent response in bacteria and chloroplasts.

RelA/SpoT Homologue (RSH) proteins modulate intracellular concentration of the ppGpp alarmone nucleotide thereby mediating the stringent response.  ppGpp binds and modulates activities of several targets including RNA polymerase, the translational GTPases EF‐G and IF2, lysine decarboxylase Ldc1, polynucleotide phosphorylase, DnaG primase and others, for its regulatory role. 

For sensing amino acid starvation RelA directly interacts with the 70S ribosome and inspects the aminoacylation status of the A‐site tRNA. If it senses the presence of deacylated tRNA ppGpp synthesis is induced.

The enzyme SpoT functions as a bifunctional enzyme. It has ppGpp synthetic and hydrolytic activities and senses several cues that modulate its net activity.

According to Atkinson and Hauryliuk (2012), the RSH protein family is divided into 30 subgroups comprising three groups: 

(i)    long RSHs (such as RelA and SpoT), 

(ii)   small alarmone synthetases (SASs), and 

(iii)  small alarmone hydrolases (SAHs), as revealed by phylogenetic analysis. 

Furthermore, RSH proteins have also been identified in eukaryotes and isolated species of archaea.  However, the ppGpp‐mediated stringent response has not yet been identified in these organisms.

Eukaryotes have a general amino acid control (GAAC) system not homologous to the RSH system but with similar function.

Other protein can bind ppGpp as well. These proteins fall into five main categories:

(i)     Cellular GTPases,

(ii)    Proteins involved in nucleotide metabolism,

(iii)   Proteins involved in lipid metabolism,

(iv)   General metabolic proteins, and

(v)    PLP-dependent basic, aliphatic amino acid decarboxylases.

In E. coli, gene expression profiles are altered during the stringent response as a result of interactions between the RNA polymerase (RNAP), ppGpp, and a specific transcription factor DksA. ppGpp and DksA facilitate opposing effects on transcription:

(i)    Downregulation of highly expressed stable RNA (rRNA and tRNA0, and cell proliferation genes, and

(ii)    Up-regulation of stress and starvation genes. 

These observations suggest that ppGpp may have a bigger role in the cellular metabolism of bacteria. However, to define the role and functions of (p)ppGpp more clearly, further structural and biochemical research is needed.

Reference

Gemma C Atkinson, Vasili Hauryliuk; Evolution and Function of the RelA/SpoT Homologue (RSH) Proteins. Published online: February 2012. DOI: 10.1002/9780470015902.a0023959.

Vasili Hauryliuk, Gemma C. Atkinson, Katsuhiko S. Murakami, Tanel Tenson & Kenn Gerdes; Recent functional insights into the role of (p)ppGpp in bacterial physiology. Nature Reviews Microbiology 13, 298–309 (2015), doi:10.1038/nrmicro3448.

Venu Kamarthapu, Vitaly Epshtein, Bradley Benjamin, Sergey Proshkin, Alexander Mironov, Michael Cashel, Evgeny Nudler; ppGpp couples transcription to DNA repair in E. coli. Science  20 May 2016: Vol. 352, Issue 6288, pp. 993-996, DOI: 10.1126/science.aad6945. http://science.sciencemag.org/content/352/6288/993

Kanjee U, Ogata K, Houry WA.; Direct binding targets of the stringent response alarmone (p)ppGpp. Mol Microbiol. 2012 Sep;85(6):1029-43. doi: 10.1111/j.1365-2958.2012.08177.x. Epub 2012 Aug 2.

Steinchen W, Schuhmacher JS, Altegoer F, et al. Catalytic mechanism and allosteric regulation of an oligomeric (p)ppGpp synthetase by an alarmone. Proceedings of the National Academy of Sciences of the United States of America. 2015;112(43):13348-13353. doi:10.1073/pnas.1505271112.

Crystal structure of the small alarmone synthetase 1: http://www.rcsb.org/pdb/explore/explore.do?structureId=5DEC

(p)ppGpp metabolism: http://www.nature.com/nrmicro/journal/v13/n5/fig_tab/nrmicro3448_F1.html

Evolution and Function of RSH proteins: http://www.els.net/WileyCDA/ElsArticle/refId-a0023959.html

RelA info:  http://www.ncbi.nlm.nih.gov/medgen/?term=RelA

Solved structures of alarmone binding enzymes

x