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Adaptation to sustained nitrogen starvation by Escherichia coli requires the eukaryote-like serine/threonine kinase YeaG.

Figueira R, Brown DR, Ferreira D, Eldridge MJ, Burchell L, Pan Z, Helaine S, Wigneshweraraj S - Sci Rep (2015)

Bottom Line: The mechanism by which yeaG acts, involves the transcriptional repression of two toxin/antitoxin modules, mqsR/mqsA and dinJ/yafQ.This, consequently, has a positive effect on the expression of rpoS, the master regulator of the general bacterial stress response.Overall, results indicate that yeaG is required to fully execute the rpoS-dependent gene expression program to allow E. coli to adapt to sustained N starvation and unravels a novel facet to the regulatory basis that underpins adaptive response to N stress.

View Article: PubMed Central - PubMed

Affiliation: MRC Centre for Molecular Microbiology and Infection, Imperial College London, UK.

ABSTRACT
The Escherichia coli eukaryote-like serine/threonine kinase, encoded by yeaG, is expressed in response to diverse stresses, including nitrogen (N) starvation. A role for yeaG in bacterial stress response is unknown. Here we reveal for the first time that wild-type E. coli displays metabolic heterogeneity following sustained periods of N starvation, with the metabolically active population displaying compromised viability. In contrast, such heterogeneity in metabolic activity is not observed in an E. coli ∆yeaG mutant, which continues to exist as a single and metabolically active population and thus displays an overall compromised ability to survive sustained periods of N starvation. The mechanism by which yeaG acts, involves the transcriptional repression of two toxin/antitoxin modules, mqsR/mqsA and dinJ/yafQ. This, consequently, has a positive effect on the expression of rpoS, the master regulator of the general bacterial stress response. Overall, results indicate that yeaG is required to fully execute the rpoS-dependent gene expression program to allow E. coli to adapt to sustained N starvation and unravels a novel facet to the regulatory basis that underpins adaptive response to N stress.

No MeSH data available.


Related in: MedlinePlus

∆yeaG mutant strain displays increased metabolic activity than the wild-type strain following sustained N starvation.(A) Representative histograms of GFP fluorescence in wild-type (blue) and ∆yeaG mutant strain (red) populations at selected time points following induction of GFP expression during ‘recovery’ growth (after 24 h N starvation). The insert is a schematic representation of the experiment. (B) Quantification of GFP production over 8 h of ‘recovery’ growth (insert) was obtained from the geometric means of GFP fluorescence. Error bars represent sd (n = 3).
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f4: ∆yeaG mutant strain displays increased metabolic activity than the wild-type strain following sustained N starvation.(A) Representative histograms of GFP fluorescence in wild-type (blue) and ∆yeaG mutant strain (red) populations at selected time points following induction of GFP expression during ‘recovery’ growth (after 24 h N starvation). The insert is a schematic representation of the experiment. (B) Quantification of GFP production over 8 h of ‘recovery’ growth (insert) was obtained from the geometric means of GFP fluorescence. Error bars represent sd (n = 3).

Mentions: Since many bacteria adjust their metabolism as an adaptive response to prolonged conditions of nutritional adversity, we considered whether the short lag phase displayed by the ∆yeaG mutant strain during ‘recovery’ growth was indicative of an altered metabolic state ergo adaptive response between ∆yeaG mutant and wild-type strains to sustained N starvation. Accumulation of GFP has been widely used as a reporter for bacterial metabolic status1415. Therefore, to investigate how E. coli adjust their metabolic state in response to N sustained starvation and to determine if this differs between the wild-type and ∆yeaG mutant strains, we assessed the rate of production of green fluorescent protein (GFP) at the single-cell level as a measure for metabolic activity during recovery growth in the presence of the inducer. Bacterial cells were recovered at hourly time points and fluorescence levels were determined by flow cytometry. Results in Fig. 4A,B, show that GFP production in the wild-type strain first became apparent at 3 h, although, strikingly, only in a subset (25%) of the population. By 4 h into ‘recovery’ growth the majority (77%) of the wild-type population contained GFP-producing bacteria and the increase in fluorescence levels continued progressively. However, a small proportion of the population remained non-fluorescent even after 5 h into ‘recovery’ growth. In marked contrast to the wild-type strain, GFP production could be detected in the ∆yeaG mutant strain population after only 2 h into ‘recovery’ growth and fluorescence levels increased at a constant rate over time. Strikingly, in the ∆yeaG population, the N starvation-induced metabolic heterogeneity clearly evident in the wild-type population was not detected. Overall, results are consistent with the ‘recovery’ growth characteristics of both strains (Fig. 1D), and indicate that the short lag phase observed for the ∆yeaG mutant strain during ‘recovery’ growth is likely due to an increased metabolic activity despite the mutant cells having experienced sustained N starvation. Importantly, the results also reveal differences between ∆yeaG mutant and wild-type strains at the population level: Whereas the wild-type population displayed heterogeneity in its metabolic activity and contained a small sub-population that appeared to be metabolically inactive, this was clearly not the case in the ∆yeaG mutant population.


Adaptation to sustained nitrogen starvation by Escherichia coli requires the eukaryote-like serine/threonine kinase YeaG.

Figueira R, Brown DR, Ferreira D, Eldridge MJ, Burchell L, Pan Z, Helaine S, Wigneshweraraj S - Sci Rep (2015)

∆yeaG mutant strain displays increased metabolic activity than the wild-type strain following sustained N starvation.(A) Representative histograms of GFP fluorescence in wild-type (blue) and ∆yeaG mutant strain (red) populations at selected time points following induction of GFP expression during ‘recovery’ growth (after 24 h N starvation). The insert is a schematic representation of the experiment. (B) Quantification of GFP production over 8 h of ‘recovery’ growth (insert) was obtained from the geometric means of GFP fluorescence. Error bars represent sd (n = 3).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4664914&req=5

f4: ∆yeaG mutant strain displays increased metabolic activity than the wild-type strain following sustained N starvation.(A) Representative histograms of GFP fluorescence in wild-type (blue) and ∆yeaG mutant strain (red) populations at selected time points following induction of GFP expression during ‘recovery’ growth (after 24 h N starvation). The insert is a schematic representation of the experiment. (B) Quantification of GFP production over 8 h of ‘recovery’ growth (insert) was obtained from the geometric means of GFP fluorescence. Error bars represent sd (n = 3).
Mentions: Since many bacteria adjust their metabolism as an adaptive response to prolonged conditions of nutritional adversity, we considered whether the short lag phase displayed by the ∆yeaG mutant strain during ‘recovery’ growth was indicative of an altered metabolic state ergo adaptive response between ∆yeaG mutant and wild-type strains to sustained N starvation. Accumulation of GFP has been widely used as a reporter for bacterial metabolic status1415. Therefore, to investigate how E. coli adjust their metabolic state in response to N sustained starvation and to determine if this differs between the wild-type and ∆yeaG mutant strains, we assessed the rate of production of green fluorescent protein (GFP) at the single-cell level as a measure for metabolic activity during recovery growth in the presence of the inducer. Bacterial cells were recovered at hourly time points and fluorescence levels were determined by flow cytometry. Results in Fig. 4A,B, show that GFP production in the wild-type strain first became apparent at 3 h, although, strikingly, only in a subset (25%) of the population. By 4 h into ‘recovery’ growth the majority (77%) of the wild-type population contained GFP-producing bacteria and the increase in fluorescence levels continued progressively. However, a small proportion of the population remained non-fluorescent even after 5 h into ‘recovery’ growth. In marked contrast to the wild-type strain, GFP production could be detected in the ∆yeaG mutant strain population after only 2 h into ‘recovery’ growth and fluorescence levels increased at a constant rate over time. Strikingly, in the ∆yeaG population, the N starvation-induced metabolic heterogeneity clearly evident in the wild-type population was not detected. Overall, results are consistent with the ‘recovery’ growth characteristics of both strains (Fig. 1D), and indicate that the short lag phase observed for the ∆yeaG mutant strain during ‘recovery’ growth is likely due to an increased metabolic activity despite the mutant cells having experienced sustained N starvation. Importantly, the results also reveal differences between ∆yeaG mutant and wild-type strains at the population level: Whereas the wild-type population displayed heterogeneity in its metabolic activity and contained a small sub-population that appeared to be metabolically inactive, this was clearly not the case in the ∆yeaG mutant population.

Bottom Line: The mechanism by which yeaG acts, involves the transcriptional repression of two toxin/antitoxin modules, mqsR/mqsA and dinJ/yafQ.This, consequently, has a positive effect on the expression of rpoS, the master regulator of the general bacterial stress response.Overall, results indicate that yeaG is required to fully execute the rpoS-dependent gene expression program to allow E. coli to adapt to sustained N starvation and unravels a novel facet to the regulatory basis that underpins adaptive response to N stress.

View Article: PubMed Central - PubMed

Affiliation: MRC Centre for Molecular Microbiology and Infection, Imperial College London, UK.

ABSTRACT
The Escherichia coli eukaryote-like serine/threonine kinase, encoded by yeaG, is expressed in response to diverse stresses, including nitrogen (N) starvation. A role for yeaG in bacterial stress response is unknown. Here we reveal for the first time that wild-type E. coli displays metabolic heterogeneity following sustained periods of N starvation, with the metabolically active population displaying compromised viability. In contrast, such heterogeneity in metabolic activity is not observed in an E. coli ∆yeaG mutant, which continues to exist as a single and metabolically active population and thus displays an overall compromised ability to survive sustained periods of N starvation. The mechanism by which yeaG acts, involves the transcriptional repression of two toxin/antitoxin modules, mqsR/mqsA and dinJ/yafQ. This, consequently, has a positive effect on the expression of rpoS, the master regulator of the general bacterial stress response. Overall, results indicate that yeaG is required to fully execute the rpoS-dependent gene expression program to allow E. coli to adapt to sustained N starvation and unravels a novel facet to the regulatory basis that underpins adaptive response to N stress.

No MeSH data available.


Related in: MedlinePlus