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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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E. coli batch cultures in mixed carbon- and single carbon limited media. Each carbon substrate concentration was at 0.2% in single substrate cultures and 0.04% in the mixed substrate experiment. A. Culture growth rates as measured by tracing OD600; B. Relative cell growth = ln(OD600t – OD600t-1); C. measured acetic acid secretion rates; D, E. substrate consumption rates in D./ mixed substrate culture and E./ single substrate cultures; F. the FBAwMC modeling prediction of substrate consumption kinetics in single substrate culture conditions. G, H, The substrate consumption curves (D and E) were normalized to the OD600nm data (A) in individual culture conditions (shown by the dotted points). Curve fitting with one phase decay equation was applied to the normalized data in (G) mixed substrate culture and (H) single substrate cultures (shown in the continuous lines). K is the rate constant and a higher K value indicates a faster substrate uptake. Half-life shows the time points when the normalized substrates concentration decreases to 50%. The data for the black tracings in panels A, B, and the data for panel D are from Ref. [7]).
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Figure 1: E. coli batch cultures in mixed carbon- and single carbon limited media. Each carbon substrate concentration was at 0.2% in single substrate cultures and 0.04% in the mixed substrate experiment. A. Culture growth rates as measured by tracing OD600; B. Relative cell growth = ln(OD600t – OD600t-1); C. measured acetic acid secretion rates; D, E. substrate consumption rates in D./ mixed substrate culture and E./ single substrate cultures; F. the FBAwMC modeling prediction of substrate consumption kinetics in single substrate culture conditions. G, H, The substrate consumption curves (D and E) were normalized to the OD600nm data (A) in individual culture conditions (shown by the dotted points). Curve fitting with one phase decay equation was applied to the normalized data in (G) mixed substrate culture and (H) single substrate cultures (shown in the continuous lines). K is the rate constant and a higher K value indicates a faster substrate uptake. Half-life shows the time points when the normalized substrates concentration decreases to 50%. The data for the black tracings in panels A, B, and the data for panel D are from Ref. [7]).

Mentions: We previously characterized the culture density-, growth rate- (Figure 1A, B, black lines, respectively) and substrate uptake kinetics (Figure 1D) of E. coli cells in mixed substrate culture, and also determined the level of acetate, a well-known metabolic byproduct of rapidly dividing E. coli cells, in that culture’s supernatant (Figure 1C, black line) [7]. The carbon source consumption profiles we observed [7] were compatible with the presence of carbon catabolite repression (CCR) in the culture, in which the sole consumption of glucose preceded the concomitant utilization of all other substrates (Figure 1D). To better understand the root cause of the observed substrate uptake patterns, we grew E. coli cells separately in the individual components of the mixed culture medium (i.e., in single carbon-limited media), the experiments being terminated upon substrate exhaustion from the growth media or when cells entered the stationary phase.


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)

E. coli batch cultures in mixed carbon- and single carbon limited media. Each carbon substrate concentration was at 0.2% in single substrate cultures and 0.04% in the mixed substrate experiment. A. Culture growth rates as measured by tracing OD600; B. Relative cell growth = ln(OD600t – OD600t-1); C. measured acetic acid secretion rates; D, E. substrate consumption rates in D./ mixed substrate culture and E./ single substrate cultures; F. the FBAwMC modeling prediction of substrate consumption kinetics in single substrate culture conditions. G, H, The substrate consumption curves (D and E) were normalized to the OD600nm data (A) in individual culture conditions (shown by the dotted points). Curve fitting with one phase decay equation was applied to the normalized data in (G) mixed substrate culture and (H) single substrate cultures (shown in the continuous lines). K is the rate constant and a higher K value indicates a faster substrate uptake. Half-life shows the time points when the normalized substrates concentration decreases to 50%. The data for the black tracings in panels A, B, and the data for panel D are from Ref. [7]).
© Copyright Policy - open-access
Related In: Results  -  Collection

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

Figure 1: E. coli batch cultures in mixed carbon- and single carbon limited media. Each carbon substrate concentration was at 0.2% in single substrate cultures and 0.04% in the mixed substrate experiment. A. Culture growth rates as measured by tracing OD600; B. Relative cell growth = ln(OD600t – OD600t-1); C. measured acetic acid secretion rates; D, E. substrate consumption rates in D./ mixed substrate culture and E./ single substrate cultures; F. the FBAwMC modeling prediction of substrate consumption kinetics in single substrate culture conditions. G, H, The substrate consumption curves (D and E) were normalized to the OD600nm data (A) in individual culture conditions (shown by the dotted points). Curve fitting with one phase decay equation was applied to the normalized data in (G) mixed substrate culture and (H) single substrate cultures (shown in the continuous lines). K is the rate constant and a higher K value indicates a faster substrate uptake. Half-life shows the time points when the normalized substrates concentration decreases to 50%. The data for the black tracings in panels A, B, and the data for panel D are from Ref. [7]).
Mentions: We previously characterized the culture density-, growth rate- (Figure 1A, B, black lines, respectively) and substrate uptake kinetics (Figure 1D) of E. coli cells in mixed substrate culture, and also determined the level of acetate, a well-known metabolic byproduct of rapidly dividing E. coli cells, in that culture’s supernatant (Figure 1C, black line) [7]. The carbon source consumption profiles we observed [7] were compatible with the presence of carbon catabolite repression (CCR) in the culture, in which the sole consumption of glucose preceded the concomitant utilization of all other substrates (Figure 1D). To better understand the root cause of the observed substrate uptake patterns, we grew E. coli cells separately in the individual components of the mixed culture medium (i.e., in single carbon-limited media), the experiments being terminated upon substrate exhaustion from the growth media or when cells entered the stationary phase.

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