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Carbon catabolite repression correlates with the maintenance of near invariant molecular crowding in proliferating E. coli cells.

Zhou Y, Vazquez A, Wise A, Warita T, Warita K, Bar-Joseph Z, Oltvai ZN - BMC Syst Biol (2013)

Bottom Line: We also find that forced transient increase of intracellular crowding or transient perturbation of CCR delay cell growth, the latter leading to associated cell density-and volume alterations.CCR is activated at an increased bacterial cell growth rate when it is required for optimal cell growth while intracellular macromolecular density is maintained within a narrow physiological range.In addition to CCR, there are likely to be other regulatory mechanisms of cell metabolism that have evolved to ensure optimal cell growth in the context of the fundamental biophysical constraint imposed by intracellular molecular crowding.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Pathology, University of Pittsburgh, School of Medicine, S701 Scaife Hall, 3550 Terrace Street, Pittsburgh, PA 15213, USA. oltvai@pitt.edu.

ABSTRACT

Background: Carbon catabolite repression (CCR) is critical for optimal bacterial growth, and in bacterial (and yeast) cells it leads to their selective consumption of a single substrate from a complex environment. However, the root cause(s) for the development of this regulatory mechanism is unknown. Previously, a flux balance model (FBAwMC) of Escherichia coli metabolism that takes into account the crowded intracellular milieu of the bacterial cell correctly predicted selective glucose uptake in a medium containing five different carbon sources, suggesting that CCR may be an adaptive mechanism that ensures optimal bacterial metabolic network activity for growth.

Results: Here, we show that slowly growing E. coli cells do not display CCR in a mixed substrate culture and gradual activation of CCR correlates with an increasing rate of E. coli cell growth and proliferation. In contrast, CCR mutant cells do not achieve fast growth in mixed substrate culture, and display differences in their cell volume and density compared to wild-type cells. Analyses of transcriptome data from wt E. coli cells indicate the expected regulation of substrate uptake and metabolic pathway utilization upon growth rate change. We also find that forced transient increase of intracellular crowding or transient perturbation of CCR delay cell growth, the latter leading to associated cell density-and volume alterations.

Conclusions: CCR is activated at an increased bacterial cell growth rate when it is required for optimal cell growth while intracellular macromolecular density is maintained within a narrow physiological range. In addition to CCR, there are likely to be other regulatory mechanisms of cell metabolism that have evolved to ensure optimal cell growth in the context of the fundamental biophysical constraint imposed by intracellular molecular crowding.

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Continuous-feed, mixed carbon medium chemostat culture of E. coli cells.E. coli cells were inoculated into the fermenter at an initial OD ~ = 0.04. The flow rate of the continuous-feed culture was adjusted every 24 hr and samples were collected after the cell density has stabilized. (A) At the indicated dilution rates samples were tested for culture density (OD600nm), pH, cell volume, and cell buoyant density; (B) displays the concentration of the indicated substrates of the mixed carbon growth medium at the indicated dilution rates, together with the concentration of the secreted acetate. In (C) the consumption ratios, which are calculated as the ratio between individual substrate uptake rate and the total carbon uptake rate, are shown at the indicated dilution rates.
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Figure 3: Continuous-feed, mixed carbon medium chemostat culture of E. coli cells.E. coli cells were inoculated into the fermenter at an initial OD ~ = 0.04. The flow rate of the continuous-feed culture was adjusted every 24 hr and samples were collected after the cell density has stabilized. (A) At the indicated dilution rates samples were tested for culture density (OD600nm), pH, cell volume, and cell buoyant density; (B) displays the concentration of the indicated substrates of the mixed carbon growth medium at the indicated dilution rates, together with the concentration of the secreted acetate. In (C) the consumption ratios, which are calculated as the ratio between individual substrate uptake rate and the total carbon uptake rate, are shown at the indicated dilution rates.

Mentions: As in glucose-limited culture [20], the cell concentration in the culture medium (i.e., the culture density) decreases with the increased exchange rate of the culture medium (Figure 3A, red curve). Also, the pH of the culture medium decreases slightly to pH ~ 6.8 at 0.2/hr dilution rate but then returns to pH ~ 6.9 and above at growth rates higher than 0.3/hr. This is likely due to the faster dilution rate of the pH 7.0 growth medium (Figure 3A, green curve). From a cell volume of 0.89 fL at the 0.1/hr dilution rate there is an initial increase in the volume of E. coli cells with increasing dilution (and cell growth) rate that reaches its peak (1.064 fL) at 0.4/hr, and then decreases and levels off with ~0.98 fL at the highest dilution rate of 0.7/hr (Figure 3A, purple curve). However, the buoyant density of E. coli cells displays much less variation. It is lowest (1.11 g/ml) at the lowest dilution/ growth rate at 0.1/hr, and then reaches a medium density (~1.13 g/ml) at growth rates 0.2 and 0.3/hr, respectively. Cell density reaches its highest value of ~1.14 g/ml at 0.4/hr dilution rate that is then maintained till 0.7/hr. These data indicate that while cell volumes change dynamically to match the faster biomass accumulation rate brought on by faster cell growth, cell buoyant density remains remarkably stable and does not increase above a threshold level.


Carbon catabolite repression correlates with the maintenance of near invariant molecular crowding in proliferating E. coli cells.

Zhou Y, Vazquez A, Wise A, Warita T, Warita K, Bar-Joseph Z, Oltvai ZN - BMC Syst Biol (2013)

Continuous-feed, mixed carbon medium chemostat culture of E. coli cells.E. coli cells were inoculated into the fermenter at an initial OD ~ = 0.04. The flow rate of the continuous-feed culture was adjusted every 24 hr and samples were collected after the cell density has stabilized. (A) At the indicated dilution rates samples were tested for culture density (OD600nm), pH, cell volume, and cell buoyant density; (B) displays the concentration of the indicated substrates of the mixed carbon growth medium at the indicated dilution rates, together with the concentration of the secreted acetate. In (C) the consumption ratios, which are calculated as the ratio between individual substrate uptake rate and the total carbon uptake rate, are shown at the indicated dilution rates.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3924228&req=5

Figure 3: Continuous-feed, mixed carbon medium chemostat culture of E. coli cells.E. coli cells were inoculated into the fermenter at an initial OD ~ = 0.04. The flow rate of the continuous-feed culture was adjusted every 24 hr and samples were collected after the cell density has stabilized. (A) At the indicated dilution rates samples were tested for culture density (OD600nm), pH, cell volume, and cell buoyant density; (B) displays the concentration of the indicated substrates of the mixed carbon growth medium at the indicated dilution rates, together with the concentration of the secreted acetate. In (C) the consumption ratios, which are calculated as the ratio between individual substrate uptake rate and the total carbon uptake rate, are shown at the indicated dilution rates.
Mentions: As in glucose-limited culture [20], the cell concentration in the culture medium (i.e., the culture density) decreases with the increased exchange rate of the culture medium (Figure 3A, red curve). Also, the pH of the culture medium decreases slightly to pH ~ 6.8 at 0.2/hr dilution rate but then returns to pH ~ 6.9 and above at growth rates higher than 0.3/hr. This is likely due to the faster dilution rate of the pH 7.0 growth medium (Figure 3A, green curve). From a cell volume of 0.89 fL at the 0.1/hr dilution rate there is an initial increase in the volume of E. coli cells with increasing dilution (and cell growth) rate that reaches its peak (1.064 fL) at 0.4/hr, and then decreases and levels off with ~0.98 fL at the highest dilution rate of 0.7/hr (Figure 3A, purple curve). However, the buoyant density of E. coli cells displays much less variation. It is lowest (1.11 g/ml) at the lowest dilution/ growth rate at 0.1/hr, and then reaches a medium density (~1.13 g/ml) at growth rates 0.2 and 0.3/hr, respectively. Cell density reaches its highest value of ~1.14 g/ml at 0.4/hr dilution rate that is then maintained till 0.7/hr. These data indicate that while cell volumes change dynamically to match the faster biomass accumulation rate brought on by faster cell growth, cell buoyant density remains remarkably stable and does not increase above a threshold level.

Bottom Line: We also find that forced transient increase of intracellular crowding or transient perturbation of CCR delay cell growth, the latter leading to associated cell density-and volume alterations.CCR is activated at an increased bacterial cell growth rate when it is required for optimal cell growth while intracellular macromolecular density is maintained within a narrow physiological range.In addition to CCR, there are likely to be other regulatory mechanisms of cell metabolism that have evolved to ensure optimal cell growth in the context of the fundamental biophysical constraint imposed by intracellular molecular crowding.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Pathology, University of Pittsburgh, School of Medicine, S701 Scaife Hall, 3550 Terrace Street, Pittsburgh, PA 15213, USA. oltvai@pitt.edu.

ABSTRACT

Background: Carbon catabolite repression (CCR) is critical for optimal bacterial growth, and in bacterial (and yeast) cells it leads to their selective consumption of a single substrate from a complex environment. However, the root cause(s) for the development of this regulatory mechanism is unknown. Previously, a flux balance model (FBAwMC) of Escherichia coli metabolism that takes into account the crowded intracellular milieu of the bacterial cell correctly predicted selective glucose uptake in a medium containing five different carbon sources, suggesting that CCR may be an adaptive mechanism that ensures optimal bacterial metabolic network activity for growth.

Results: Here, we show that slowly growing E. coli cells do not display CCR in a mixed substrate culture and gradual activation of CCR correlates with an increasing rate of E. coli cell growth and proliferation. In contrast, CCR mutant cells do not achieve fast growth in mixed substrate culture, and display differences in their cell volume and density compared to wild-type cells. Analyses of transcriptome data from wt E. coli cells indicate the expected regulation of substrate uptake and metabolic pathway utilization upon growth rate change. We also find that forced transient increase of intracellular crowding or transient perturbation of CCR delay cell growth, the latter leading to associated cell density-and volume alterations.

Conclusions: CCR is activated at an increased bacterial cell growth rate when it is required for optimal cell growth while intracellular macromolecular density is maintained within a narrow physiological range. In addition to CCR, there are likely to be other regulatory mechanisms of cell metabolism that have evolved to ensure optimal cell growth in the context of the fundamental biophysical constraint imposed by intracellular molecular crowding.

Show MeSH
Related in: MedlinePlus