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Constitutive expression of the sRNA GadY decreases acetate production and improves E. coli growth.

Negrete A, Shiloach J - Microb. Cell Fact. (2015)

Bottom Line: It was reported that at mild acidic conditions (pH 5.8), the Hfq-associated sRNA GadY is activated.After analyzing 17 genes associated with acid stress, it was found that at pH 7.0 LDS was expressed in the early exponential phase and GDS was expressed in the late exponential phase in both strains.The expression of GadY also decreases acetate production regardless of pH, which decreases the inhibitory effect of this acid on bacterial growth.

View Article: PubMed Central - PubMed

Affiliation: Biotechnology Core Laboratory, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Building 14A Room 173, Bethesda, MD, 20892, USA.

ABSTRACT

Background: Escherichia coli responds to acid stress by applying various physiological, metabolic, and proton-consuming mechanisms depending on the growth media composition, cell density, growth phase, pH, and aerobic or anaerobic growth conditions. It was reported that at mild acidic conditions (pH 5.8), the Hfq-associated sRNA GadY is activated. It was also reported that the two decarboxylase systems-the lysine decarboxylase system (LDS) and the glutamate decarboxylase system (GDS)-are activated to maintain intracellular balance of protons. The purpose of this study was to establish the role of GadY in high density growth of E. coli and to evaluate the possibility of using this small RNA to create an acid-resistant strain suitable for industrial applications.

Results: Parental E. coli K-12 and constitutively expressing GadY strains were grown to high cell densities in a bioreactor at pH 7.0 and pH 6.0. At pH 7.0, both strains grew to similar cell densities of 43 OD, but the constitutively expressing GadY strain produced around 6 g/L acetate compared with 10 g/L by the parental strain. At pH 6.0, the parental strain grew to an OD of 20 and produced 10 g/L of acetate while the GadY strain grew to an average OD of 31 and produced 4 g/L acetate. After analyzing 17 genes associated with acid stress, it was found that at pH 7.0 LDS was expressed in the early exponential phase and GDS was expressed in the late exponential phase in both strains. However, at pH.6.0, GDS was expressed in the late exponential phase only in the parental strain and not in the constitutively expressing GadY strain, while there was no difference in the LDS expression pattern; it was expressed in the early exponential phase in both strains. This indicates that GadY affects GDS expression at low pH since the GDS was not detected in the GadY strain at pH 6.0.

Conclusions: The constitutive expression of GadY improves E. coli growth at pH 6.0 by deactivating the expression of the GDS in the late exponential growth phase. The expression of GadY also decreases acetate production regardless of pH, which decreases the inhibitory effect of this acid on bacterial growth.

No MeSH data available.


Related in: MedlinePlus

Regulatory network of the glutamate-dependent acid resistance. Modified from Foster, 2004. Large arrows represent genes, small arrows indicate positive control, and dotted lines denote negative control. The genes for the GDS are represented as solid black arrows
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Fig8: Regulatory network of the glutamate-dependent acid resistance. Modified from Foster, 2004. Large arrows represent genes, small arrows indicate positive control, and dotted lines denote negative control. The genes for the GDS are represented as solid black arrows

Mentions: In the GadY strain grown at pH 7.0, the LDS was activated in the early exponential growth phase and the GDS in the late exponential phase, suggesting that both the LDS and the GDS have a role in maintaining intracellular homeostasis linked to acetate accumulation at pH 7.0. This differs from the induction of the GDS and the LDS described previously as acid stress response in E. coli grown at pH 5.8 [5, 7–9, 28, 29]. As observed in this work, the expression of the LDS and the GDS in the GadY strain grown at pH 7.0 is linked to acetate accumulation and not to low pH. The interaction between GadY and the GDS has been previously observed in an acid response regulatory network described in Fig. 8 [5, 15]. In that network, GadY induced GadX, which then activated GadW and the expression of the GDS directly or via gadE. In this study, GadY activated the GDS at pH 7.0 independent of gadE, gadX, and gadW as these three genes were not expressed. Also, it has been reported that the GDS can be induced either by acetate accumulation or by GadW via GadE in the presence of acetate [29–33]. Our results indicated that GadY activated GDS at pH 7.0 possibly, by an alternative mechanism to GadE, GadX, and GadW or by acetate accumulation.Fig. 8


Constitutive expression of the sRNA GadY decreases acetate production and improves E. coli growth.

Negrete A, Shiloach J - Microb. Cell Fact. (2015)

Regulatory network of the glutamate-dependent acid resistance. Modified from Foster, 2004. Large arrows represent genes, small arrows indicate positive control, and dotted lines denote negative control. The genes for the GDS are represented as solid black arrows
© Copyright Policy - OpenAccess
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4574537&req=5

Fig8: Regulatory network of the glutamate-dependent acid resistance. Modified from Foster, 2004. Large arrows represent genes, small arrows indicate positive control, and dotted lines denote negative control. The genes for the GDS are represented as solid black arrows
Mentions: In the GadY strain grown at pH 7.0, the LDS was activated in the early exponential growth phase and the GDS in the late exponential phase, suggesting that both the LDS and the GDS have a role in maintaining intracellular homeostasis linked to acetate accumulation at pH 7.0. This differs from the induction of the GDS and the LDS described previously as acid stress response in E. coli grown at pH 5.8 [5, 7–9, 28, 29]. As observed in this work, the expression of the LDS and the GDS in the GadY strain grown at pH 7.0 is linked to acetate accumulation and not to low pH. The interaction between GadY and the GDS has been previously observed in an acid response regulatory network described in Fig. 8 [5, 15]. In that network, GadY induced GadX, which then activated GadW and the expression of the GDS directly or via gadE. In this study, GadY activated the GDS at pH 7.0 independent of gadE, gadX, and gadW as these three genes were not expressed. Also, it has been reported that the GDS can be induced either by acetate accumulation or by GadW via GadE in the presence of acetate [29–33]. Our results indicated that GadY activated GDS at pH 7.0 possibly, by an alternative mechanism to GadE, GadX, and GadW or by acetate accumulation.Fig. 8

Bottom Line: It was reported that at mild acidic conditions (pH 5.8), the Hfq-associated sRNA GadY is activated.After analyzing 17 genes associated with acid stress, it was found that at pH 7.0 LDS was expressed in the early exponential phase and GDS was expressed in the late exponential phase in both strains.The expression of GadY also decreases acetate production regardless of pH, which decreases the inhibitory effect of this acid on bacterial growth.

View Article: PubMed Central - PubMed

Affiliation: Biotechnology Core Laboratory, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Building 14A Room 173, Bethesda, MD, 20892, USA.

ABSTRACT

Background: Escherichia coli responds to acid stress by applying various physiological, metabolic, and proton-consuming mechanisms depending on the growth media composition, cell density, growth phase, pH, and aerobic or anaerobic growth conditions. It was reported that at mild acidic conditions (pH 5.8), the Hfq-associated sRNA GadY is activated. It was also reported that the two decarboxylase systems-the lysine decarboxylase system (LDS) and the glutamate decarboxylase system (GDS)-are activated to maintain intracellular balance of protons. The purpose of this study was to establish the role of GadY in high density growth of E. coli and to evaluate the possibility of using this small RNA to create an acid-resistant strain suitable for industrial applications.

Results: Parental E. coli K-12 and constitutively expressing GadY strains were grown to high cell densities in a bioreactor at pH 7.0 and pH 6.0. At pH 7.0, both strains grew to similar cell densities of 43 OD, but the constitutively expressing GadY strain produced around 6 g/L acetate compared with 10 g/L by the parental strain. At pH 6.0, the parental strain grew to an OD of 20 and produced 10 g/L of acetate while the GadY strain grew to an average OD of 31 and produced 4 g/L acetate. After analyzing 17 genes associated with acid stress, it was found that at pH 7.0 LDS was expressed in the early exponential phase and GDS was expressed in the late exponential phase in both strains. However, at pH.6.0, GDS was expressed in the late exponential phase only in the parental strain and not in the constitutively expressing GadY strain, while there was no difference in the LDS expression pattern; it was expressed in the early exponential phase in both strains. This indicates that GadY affects GDS expression at low pH since the GDS was not detected in the GadY strain at pH 6.0.

Conclusions: The constitutive expression of GadY improves E. coli growth at pH 6.0 by deactivating the expression of the GDS in the late exponential growth phase. The expression of GadY also decreases acetate production regardless of pH, which decreases the inhibitory effect of this acid on bacterial growth.

No MeSH data available.


Related in: MedlinePlus