Differential regulation by ppGpp versus pppGpp in Escherichia coli

Undine Mechold, Katarzyna Potrykus, Helen Murphy, Katsuhiko S. Murakami, Michael Cashel

Research output: Contribution to journalArticle

98 Citations (Scopus)

Abstract

Both ppGpp and pppGpp are thought to function collectively as second messengers for many complex cellular responses to nutritional stress throughout biology. There are few indications that their regulatory effects might be different; however, this question has been largely unexplored for lack of an ability to experimentally manipulate the relative abundance of ppGpp and pppGpp. Here, we achieve preferential accumulation of either ppGpp or pppGpp with Escherichia coli strains through induction of different Streptococcal (p)ppGpp synthetase fragments. In addition, expression of E. coli GppA, a pppGpp 5 0 -gamma phosphate hydrolase that converts pppGpp to ppGpp, is manipulated to fine tune differential accumulation of ppGpp and pppGpp. In vivo and in vitro experiments show that pppGpp is less potent than ppGpp with respect to regulation of growth rate, RNA/DNA ratios, ribosomal RNA P1 promoter transcription inhibition, threonine operon promoter activation and RpoS induction. To provide further insights into regulation by (p)ppGpp, we have also determined crystal structures of E. coli RNA polymerase-p 70 holoenzyme with ppGpp and pppGpp. We find that both nucleotides bind to a site at the interface between β′ and ω subunits.

Original languageEnglish (US)
Pages (from-to)6175-6189
Number of pages15
JournalNucleic acids research
Volume41
Issue number12
DOIs
StatePublished - Jul 1 2013

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Escherichia coli
Holoenzymes
Ribosomal RNA
Second Messenger Systems
Hydrolases
Threonine
DNA-Directed RNA Polymerases
Operon
Nucleotides
Phosphates
RNA
DNA
Growth
guanosine 3',5'-polyphosphate synthetases
In Vitro Techniques

All Science Journal Classification (ASJC) codes

  • Genetics

Cite this

Mechold, Undine ; Potrykus, Katarzyna ; Murphy, Helen ; Murakami, Katsuhiko S. ; Cashel, Michael. / Differential regulation by ppGpp versus pppGpp in Escherichia coli. In: Nucleic acids research. 2013 ; Vol. 41, No. 12. pp. 6175-6189.
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Mechold, U, Potrykus, K, Murphy, H, Murakami, KS & Cashel, M 2013, 'Differential regulation by ppGpp versus pppGpp in Escherichia coli', Nucleic acids research, vol. 41, no. 12, pp. 6175-6189. https://doi.org/10.1093/nar/gkt302

Differential regulation by ppGpp versus pppGpp in Escherichia coli. / Mechold, Undine; Potrykus, Katarzyna; Murphy, Helen; Murakami, Katsuhiko S.; Cashel, Michael.

In: Nucleic acids research, Vol. 41, No. 12, 01.07.2013, p. 6175-6189.

Research output: Contribution to journalArticle

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