NirD curtails the stringent response by inhibiting RelA activity in Escherichia coli.

Loïc Léger,Elsa Germain,Deborah Byrne,Etienne Maisonneuve,Paul Guiraud

doi:10.7554/elife.64092

Abstract

Bacteria regulate their metabolism to adapt and survive adverse conditions, in particular to stressful downshifts in nutrient availability. These shifts trigger the so-called stringent response, coordinated by the signaling molecules guanosine tetra and pentaphosphate collectively referred to as (p)ppGpp. In Escherichia coli, accumulation of theses alarmones depends on the (p)ppGpp synthetase RelA and the bifunctional (p)ppGpp synthetase/hydrolase SpoT. A tight regulation of these intracellular activities is therefore crucial to rapidly adjust the (p)ppGpp levels in response to environmental stresses but also to avoid toxic consequences of (p)ppGpp over-accumulation. In this study, we show that the small protein NirD restrains RelA-dependent accumulation of (p)ppGpp and can inhibit the stringent response in E. coli. Mechanistically, our in vivo and in vitro studies reveal that NirD directly binds the catalytic domains of RelA to balance (p)ppGpp accumulation. Finally, we show that NirD can control RelA activity by directly inhibiting the rate of (p)ppGpp synthesis.

Highlights

Bacteria have evolved numerous molecular mechanisms to detect and cope with environmental stress, including the use of nucleotide-based signaling pathways to efficiently coordinate cellular processes and provide a fast response
E. coli MG1655 cells harboring the pBbS2k-relA were transformed with a pooled collection of plasmids obtained from the ASKA library containing almost all E. coli K-12 genes (Kitagawa et al, 2005), each cloned into the high-copy-number vector pCA24N downstream of the isopropyl b-D-1-thiogalactopyranoside (IPTG)-inducible PT5-lac promoter (Figure 1A)
While SpoT is a bifunctional enzyme, RelA is monofunctional with a degenerated hydrolase domain, making SpoT the primary source of hydrolysis (Xiao et al, 1991)

Summary

Introduction

Bacteria have evolved numerous molecular mechanisms to detect and cope with environmental stress, including the use of nucleotide-based signaling pathways to efficiently coordinate cellular processes and provide a fast response. Among these signaling pathways, the stringent response is a general stress response that is mediated by the accumulation of the nucleotides guanosine 50diphosphate 30-diphosphate (ppGpp) and guanosine 50-triphosphate 30-diphosphate (pppGpp), collectively known as (p)ppGpp (Potrykus and Cashel, 2008). (p)ppGpp binds directly and alters the activity of several enzymes including DNA primase, translation factors, lysine decarboxylase, and polyphosphate kinase (Corrigan et al, 2016; Kanjee et al, 2012; Wang et al, 2019a; Zhang et al, 2018). This rewiring of cell physiology appears to play a critical role in the regulation of bacterial virulence, survival during host invasion, and antibiotic resistance and tolerance (Hauryliuk et al, 2015; Hengge, 2020; Irving et al, 2021; Potrykus and Cashel, 2008)

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Conclusion