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Regulatory and metabolic rewiring during laboratory evolution of ethanol tolerance in E. coli.

Goodarzi H, Bennett BD, Amini S, Reaves ML, Hottes AK, Rabinowitz JD, Tavazoie S - Mol. Syst. Biol. (2010)

Bottom Line: However, revealing the underlying molecular mechanisms has been challenging as changes in fitness may result from perturbations to many pathways, any of which may contribute relatively little.A module-level computational analysis was then used to reveal the organization of the contributing loci into cellular processes and regulatory pathways (e.g. osmoregulation and cell-wall biogenesis) whose modifications significantly affect ethanol tolerance.Remarkably, these laboratory-evolved strains, by and large, follow the same adaptive paths as inferred from our coarse-grained search of the fitness landscape.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Biology, Princeton University, Princeton, NJ 08544, USA.

ABSTRACT
Understanding the genetic basis of adaptation is a central problem in biology. However, revealing the underlying molecular mechanisms has been challenging as changes in fitness may result from perturbations to many pathways, any of which may contribute relatively little. We have developed a combined experimental/computational framework to address this problem and used it to understand the genetic basis of ethanol tolerance in Escherichia coli. We used fitness profiling to measure the consequences of single-locus perturbations in the context of ethanol exposure. A module-level computational analysis was then used to reveal the organization of the contributing loci into cellular processes and regulatory pathways (e.g. osmoregulation and cell-wall biogenesis) whose modifications significantly affect ethanol tolerance. Strikingly, we discovered that a dominant component of adaptation involves metabolic rewiring that boosts intracellular ethanol degradation and assimilation. Through phenotypic and metabolomic analysis of laboratory-evolved ethanol-tolerant strains, we investigated naturally accessible pathways of ethanol tolerance. Remarkably, these laboratory-evolved strains, by and large, follow the same adaptive paths as inferred from our coarse-grained search of the fitness landscape.

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Acid stress response and ethanol tolerance. (A) The fitness scores of the genes active in acid tolerance are shown here for all the samples. Disrupting the activity of these genes is beneficial for ethanol tolerance, whereas their overexpression is deleterious. (B) We used kill curves (i.e. the number of CFUs at each timepoint) to compare the ethanol tolerance of the wild-type strain and Δafi strain (containing a partial deletion in the acid fitness island) in 7% v/v ethanol (left). Δafi, an acid-sensitive strain, showed a significant increase in ethanol tolerance (P-value<1e−3). The experiments were performed in triplicate, and the error bars mark the minimum and maximum for each point.
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f3: Acid stress response and ethanol tolerance. (A) The fitness scores of the genes active in acid tolerance are shown here for all the samples. Disrupting the activity of these genes is beneficial for ethanol tolerance, whereas their overexpression is deleterious. (B) We used kill curves (i.e. the number of CFUs at each timepoint) to compare the ethanol tolerance of the wild-type strain and Δafi strain (containing a partial deletion in the acid fitness island) in 7% v/v ethanol (left). Δafi, an acid-sensitive strain, showed a significant increase in ethanol tolerance (P-value<1e−3). The experiments were performed in triplicate, and the error bars mark the minimum and maximum for each point.

Mentions: Remarkably, we also discovered that the acid stress response pathway (Foster, 2004) antagonizes ethanol tolerance (Figure 2). We observed that the overexpression of the genes in this pathway increases ethanol sensitivity (Figure 3A). To further validate this effect, we made a partial deletion of the acid fitness island (Δafi: b3506–b3511), which includes four of the genes presented in Figure 3A (Mates et al, 2007), and found that in comparison with wild type, the resulting strain shows a significantly increased survival rate in 7% (v/v) ethanol (P-value<0.001; Figure 3B).


Regulatory and metabolic rewiring during laboratory evolution of ethanol tolerance in E. coli.

Goodarzi H, Bennett BD, Amini S, Reaves ML, Hottes AK, Rabinowitz JD, Tavazoie S - Mol. Syst. Biol. (2010)

Acid stress response and ethanol tolerance. (A) The fitness scores of the genes active in acid tolerance are shown here for all the samples. Disrupting the activity of these genes is beneficial for ethanol tolerance, whereas their overexpression is deleterious. (B) We used kill curves (i.e. the number of CFUs at each timepoint) to compare the ethanol tolerance of the wild-type strain and Δafi strain (containing a partial deletion in the acid fitness island) in 7% v/v ethanol (left). Δafi, an acid-sensitive strain, showed a significant increase in ethanol tolerance (P-value<1e−3). The experiments were performed in triplicate, and the error bars mark the minimum and maximum for each point.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2913397&req=5

f3: Acid stress response and ethanol tolerance. (A) The fitness scores of the genes active in acid tolerance are shown here for all the samples. Disrupting the activity of these genes is beneficial for ethanol tolerance, whereas their overexpression is deleterious. (B) We used kill curves (i.e. the number of CFUs at each timepoint) to compare the ethanol tolerance of the wild-type strain and Δafi strain (containing a partial deletion in the acid fitness island) in 7% v/v ethanol (left). Δafi, an acid-sensitive strain, showed a significant increase in ethanol tolerance (P-value<1e−3). The experiments were performed in triplicate, and the error bars mark the minimum and maximum for each point.
Mentions: Remarkably, we also discovered that the acid stress response pathway (Foster, 2004) antagonizes ethanol tolerance (Figure 2). We observed that the overexpression of the genes in this pathway increases ethanol sensitivity (Figure 3A). To further validate this effect, we made a partial deletion of the acid fitness island (Δafi: b3506–b3511), which includes four of the genes presented in Figure 3A (Mates et al, 2007), and found that in comparison with wild type, the resulting strain shows a significantly increased survival rate in 7% (v/v) ethanol (P-value<0.001; Figure 3B).

Bottom Line: However, revealing the underlying molecular mechanisms has been challenging as changes in fitness may result from perturbations to many pathways, any of which may contribute relatively little.A module-level computational analysis was then used to reveal the organization of the contributing loci into cellular processes and regulatory pathways (e.g. osmoregulation and cell-wall biogenesis) whose modifications significantly affect ethanol tolerance.Remarkably, these laboratory-evolved strains, by and large, follow the same adaptive paths as inferred from our coarse-grained search of the fitness landscape.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Biology, Princeton University, Princeton, NJ 08544, USA.

ABSTRACT
Understanding the genetic basis of adaptation is a central problem in biology. However, revealing the underlying molecular mechanisms has been challenging as changes in fitness may result from perturbations to many pathways, any of which may contribute relatively little. We have developed a combined experimental/computational framework to address this problem and used it to understand the genetic basis of ethanol tolerance in Escherichia coli. We used fitness profiling to measure the consequences of single-locus perturbations in the context of ethanol exposure. A module-level computational analysis was then used to reveal the organization of the contributing loci into cellular processes and regulatory pathways (e.g. osmoregulation and cell-wall biogenesis) whose modifications significantly affect ethanol tolerance. Strikingly, we discovered that a dominant component of adaptation involves metabolic rewiring that boosts intracellular ethanol degradation and assimilation. Through phenotypic and metabolomic analysis of laboratory-evolved ethanol-tolerant strains, we investigated naturally accessible pathways of ethanol tolerance. Remarkably, these laboratory-evolved strains, by and large, follow the same adaptive paths as inferred from our coarse-grained search of the fitness landscape.

Show MeSH
Related in: MedlinePlus