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Need-based activation of ammonium uptake in Escherichia coli.

Kim M, Zhang Z, Okano H, Yan D, Groisman A, Hwa T - Mol. Syst. Biol. (2012)

Bottom Line: We find that as the ambient ammonium concentration is reduced, E. coli cells first maximize their ability to assimilate the gaseous NH3 diffusing into the cytoplasm and then abruptly activate ammonium transport.Quantitative modeling of known interactions reveals an integral feedback mechanism by which this need-based uptake strategy is implemented.This novel strategy ensures that the expensive cost of upholding the internal ammonium concentration against back diffusion is kept at a minimum.

View Article: PubMed Central - PubMed

Affiliation: Department of Physics, University of California at San Diego, La Jolla, CA, USA.

ABSTRACT
The efficient sequestration of nutrients is vital for the growth and survival of microorganisms. Some nutrients, such as CO2 and NH3, are readily diffusible across the cell membrane. The large membrane permeability of these nutrients obviates the need of transporters when the ambient level is high. When the ambient level is low, however, maintaining a high intracellular nutrient level against passive back diffusion is both challenging and costly. Here, we study the delicate management of ammonium (NH4+/NH3) sequestration by E. coli cells using microfluidic chemostats. We find that as the ambient ammonium concentration is reduced, E. coli cells first maximize their ability to assimilate the gaseous NH3 diffusing into the cytoplasm and then abruptly activate ammonium transport. The onset of transport varies under different growth conditions, but always occurring just as needed to maintain growth. Quantitative modeling of known interactions reveals an integral feedback mechanism by which this need-based uptake strategy is implemented. This novel strategy ensures that the expensive cost of upholding the internal ammonium concentration against back diffusion is kept at a minimum.

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Coordination of AmtB activity with the cell's growth status. The growth rate of the culture in ammonium-replete condition was changed by using different carbon sources: glucose-6-phosphate (g6p) plus gluconate (squares), and glucose (triangles). Data from Figures 1 and 2 (glycerol as the carbon source) are re-plotted for comparison (circles). Solid, dashed, and dotted arrows are associated with glycerol, glucose, and g6p plus gluconate. Solid and open symbols indicate the wild-type and ΔamtB strain (EQ66 and EQ130), respectively. (A) The growth rate of the wild-type and ΔamtB strain for all carbon sources tested. The onset of the growth defect of the ΔamtB strain (black arrows) was at successively higher ambient NH4+ concentrations for the carbon sources supporting faster growth. (B) The amtB promoter activities (reported by GFP) of the wild-type and ΔamtB strains deviated (green arrows) at slightly higher ambient NH4+ concentrations than where growth defect set in for the ΔamtB strain (gray zone), indicating that ammonium transport in the wild type occurred barely above where it would be needed to maintain the growth, in coordination with the different growth conditions. (C) The maintenance of the internal NH4+ concentration for the different carbon sources. (D) The growth rate (black symbols) and ammonium transport flux (green symbols) plotted against the internal NH4+ concentrations for the different carbon sources. The abrupt increase in the ammonium flux occurred above the respective onset of growth defect of the ΔamtB strain (right of the gray zone). The black lines in Figures 3C and D show the linear fits of our model (Supplementary Equations S36–S38), from which the onset of AmtB activation N*int is determined (green arrows); see Supplementary Table 5 for the parameters. All the data plotted here are given in Supplementary Tables 8–11. a.u., arbitrary units.
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f3: Coordination of AmtB activity with the cell's growth status. The growth rate of the culture in ammonium-replete condition was changed by using different carbon sources: glucose-6-phosphate (g6p) plus gluconate (squares), and glucose (triangles). Data from Figures 1 and 2 (glycerol as the carbon source) are re-plotted for comparison (circles). Solid, dashed, and dotted arrows are associated with glycerol, glucose, and g6p plus gluconate. Solid and open symbols indicate the wild-type and ΔamtB strain (EQ66 and EQ130), respectively. (A) The growth rate of the wild-type and ΔamtB strain for all carbon sources tested. The onset of the growth defect of the ΔamtB strain (black arrows) was at successively higher ambient NH4+ concentrations for the carbon sources supporting faster growth. (B) The amtB promoter activities (reported by GFP) of the wild-type and ΔamtB strains deviated (green arrows) at slightly higher ambient NH4+ concentrations than where growth defect set in for the ΔamtB strain (gray zone), indicating that ammonium transport in the wild type occurred barely above where it would be needed to maintain the growth, in coordination with the different growth conditions. (C) The maintenance of the internal NH4+ concentration for the different carbon sources. (D) The growth rate (black symbols) and ammonium transport flux (green symbols) plotted against the internal NH4+ concentrations for the different carbon sources. The abrupt increase in the ammonium flux occurred above the respective onset of growth defect of the ΔamtB strain (right of the gray zone). The black lines in Figures 3C and D show the linear fits of our model (Supplementary Equations S36–S38), from which the onset of AmtB activation N*int is determined (green arrows); see Supplementary Table 5 for the parameters. All the data plotted here are given in Supplementary Tables 8–11. a.u., arbitrary units.

Mentions: Cells coordinate nitrogen and carbon metabolism through various mechanisms (Commichau et al, 2006; Doucette et al, 2011). Since cells growing on different carbon sources exhibit different growth rates in ammonium-replete conditions and hence have different demands for nitrogen, we varied the carbon source in the growth medium to ones that provided successively faster growth than what glycerol can provide. We expect this would force the cells to activate AmtB at higher ammonium concentrations if the faster growth rates are to be maintained as the external ammonium concentration is reduced. Indeed, the data on growth rates (Figure 3A) show that the wild type (solid symbols) was able to maintain growth throughout the range of NH4+ concentrations tested. In contrast, the ΔamtB strain (open symbols) exhibited growth reduction as the ambient NH4+ concentration was reduced, and the onset of the growth defect (black arrows) occurred at successively higher ambient NH4+ concentrations for the medium supporting faster growth. Also, Figure 3B shows that the NH4+ concentrations at which the amtB promoter activity (GFP) of the wild-type and ΔamtB strain deviate (green arrows) are always just slightly above the respective onset of the growth defect for the three carbon sources tested.


Need-based activation of ammonium uptake in Escherichia coli.

Kim M, Zhang Z, Okano H, Yan D, Groisman A, Hwa T - Mol. Syst. Biol. (2012)

Coordination of AmtB activity with the cell's growth status. The growth rate of the culture in ammonium-replete condition was changed by using different carbon sources: glucose-6-phosphate (g6p) plus gluconate (squares), and glucose (triangles). Data from Figures 1 and 2 (glycerol as the carbon source) are re-plotted for comparison (circles). Solid, dashed, and dotted arrows are associated with glycerol, glucose, and g6p plus gluconate. Solid and open symbols indicate the wild-type and ΔamtB strain (EQ66 and EQ130), respectively. (A) The growth rate of the wild-type and ΔamtB strain for all carbon sources tested. The onset of the growth defect of the ΔamtB strain (black arrows) was at successively higher ambient NH4+ concentrations for the carbon sources supporting faster growth. (B) The amtB promoter activities (reported by GFP) of the wild-type and ΔamtB strains deviated (green arrows) at slightly higher ambient NH4+ concentrations than where growth defect set in for the ΔamtB strain (gray zone), indicating that ammonium transport in the wild type occurred barely above where it would be needed to maintain the growth, in coordination with the different growth conditions. (C) The maintenance of the internal NH4+ concentration for the different carbon sources. (D) The growth rate (black symbols) and ammonium transport flux (green symbols) plotted against the internal NH4+ concentrations for the different carbon sources. The abrupt increase in the ammonium flux occurred above the respective onset of growth defect of the ΔamtB strain (right of the gray zone). The black lines in Figures 3C and D show the linear fits of our model (Supplementary Equations S36–S38), from which the onset of AmtB activation N*int is determined (green arrows); see Supplementary Table 5 for the parameters. All the data plotted here are given in Supplementary Tables 8–11. a.u., arbitrary units.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3472687&req=5

f3: Coordination of AmtB activity with the cell's growth status. The growth rate of the culture in ammonium-replete condition was changed by using different carbon sources: glucose-6-phosphate (g6p) plus gluconate (squares), and glucose (triangles). Data from Figures 1 and 2 (glycerol as the carbon source) are re-plotted for comparison (circles). Solid, dashed, and dotted arrows are associated with glycerol, glucose, and g6p plus gluconate. Solid and open symbols indicate the wild-type and ΔamtB strain (EQ66 and EQ130), respectively. (A) The growth rate of the wild-type and ΔamtB strain for all carbon sources tested. The onset of the growth defect of the ΔamtB strain (black arrows) was at successively higher ambient NH4+ concentrations for the carbon sources supporting faster growth. (B) The amtB promoter activities (reported by GFP) of the wild-type and ΔamtB strains deviated (green arrows) at slightly higher ambient NH4+ concentrations than where growth defect set in for the ΔamtB strain (gray zone), indicating that ammonium transport in the wild type occurred barely above where it would be needed to maintain the growth, in coordination with the different growth conditions. (C) The maintenance of the internal NH4+ concentration for the different carbon sources. (D) The growth rate (black symbols) and ammonium transport flux (green symbols) plotted against the internal NH4+ concentrations for the different carbon sources. The abrupt increase in the ammonium flux occurred above the respective onset of growth defect of the ΔamtB strain (right of the gray zone). The black lines in Figures 3C and D show the linear fits of our model (Supplementary Equations S36–S38), from which the onset of AmtB activation N*int is determined (green arrows); see Supplementary Table 5 for the parameters. All the data plotted here are given in Supplementary Tables 8–11. a.u., arbitrary units.
Mentions: Cells coordinate nitrogen and carbon metabolism through various mechanisms (Commichau et al, 2006; Doucette et al, 2011). Since cells growing on different carbon sources exhibit different growth rates in ammonium-replete conditions and hence have different demands for nitrogen, we varied the carbon source in the growth medium to ones that provided successively faster growth than what glycerol can provide. We expect this would force the cells to activate AmtB at higher ammonium concentrations if the faster growth rates are to be maintained as the external ammonium concentration is reduced. Indeed, the data on growth rates (Figure 3A) show that the wild type (solid symbols) was able to maintain growth throughout the range of NH4+ concentrations tested. In contrast, the ΔamtB strain (open symbols) exhibited growth reduction as the ambient NH4+ concentration was reduced, and the onset of the growth defect (black arrows) occurred at successively higher ambient NH4+ concentrations for the medium supporting faster growth. Also, Figure 3B shows that the NH4+ concentrations at which the amtB promoter activity (GFP) of the wild-type and ΔamtB strain deviate (green arrows) are always just slightly above the respective onset of the growth defect for the three carbon sources tested.

Bottom Line: We find that as the ambient ammonium concentration is reduced, E. coli cells first maximize their ability to assimilate the gaseous NH3 diffusing into the cytoplasm and then abruptly activate ammonium transport.Quantitative modeling of known interactions reveals an integral feedback mechanism by which this need-based uptake strategy is implemented.This novel strategy ensures that the expensive cost of upholding the internal ammonium concentration against back diffusion is kept at a minimum.

View Article: PubMed Central - PubMed

Affiliation: Department of Physics, University of California at San Diego, La Jolla, CA, USA.

ABSTRACT
The efficient sequestration of nutrients is vital for the growth and survival of microorganisms. Some nutrients, such as CO2 and NH3, are readily diffusible across the cell membrane. The large membrane permeability of these nutrients obviates the need of transporters when the ambient level is high. When the ambient level is low, however, maintaining a high intracellular nutrient level against passive back diffusion is both challenging and costly. Here, we study the delicate management of ammonium (NH4+/NH3) sequestration by E. coli cells using microfluidic chemostats. We find that as the ambient ammonium concentration is reduced, E. coli cells first maximize their ability to assimilate the gaseous NH3 diffusing into the cytoplasm and then abruptly activate ammonium transport. The onset of transport varies under different growth conditions, but always occurring just as needed to maintain growth. Quantitative modeling of known interactions reveals an integral feedback mechanism by which this need-based uptake strategy is implemented. This novel strategy ensures that the expensive cost of upholding the internal ammonium concentration against back diffusion is kept at a minimum.

Show MeSH
Related in: MedlinePlus