Limits...
Large-scale 13C-flux analysis reveals distinct transcriptional control of respiratory and fermentative metabolism in Escherichia coli.

Haverkorn van Rijsewijk BR, Nanchen A, Nallet S, Kleijn RJ, Sauer U - Mol. Syst. Biol. (2011)

Bottom Line: Despite our increasing topological knowledge on regulation networks in model bacteria, it is largely unknown which of the many co-occurring regulatory events actually control metabolic function and the distribution of intracellular fluxes.While 2/3 of the regulators directly or indirectly affected absolute flux rates, the partitioning between different pathways remained largely stable with transcriptional control focusing primarily on the acetyl-CoA branch point.Five further transcription factors affected this flux only indirectly through cAMP and Crp by increasing the galactose uptake rate.

View Article: PubMed Central - PubMed

Affiliation: Institute of Molecular Systems Biology, ETH Zurich, Zurich, Switzerland.

ABSTRACT
Despite our increasing topological knowledge on regulation networks in model bacteria, it is largely unknown which of the many co-occurring regulatory events actually control metabolic function and the distribution of intracellular fluxes. Here, we unravel condition-dependent transcriptional control of Escherichia coli metabolism by large-scale (13)C-flux analysis in 91 transcriptional regulator mutants on glucose and galactose. In contrast to the canonical respiro-fermentative glucose metabolism, fully respiratory galactose metabolism depends exclusively on the phosphoenol-pyruvate (PEP)-glyoxylate cycle. While 2/3 of the regulators directly or indirectly affected absolute flux rates, the partitioning between different pathways remained largely stable with transcriptional control focusing primarily on the acetyl-CoA branch point. Flux distribution control was achieved by nine transcription factors on glucose, including ArcA, Fur, PdhR, IHF A and IHF B, but was exclusively mediated by the cAMP-dependent Crp regulation of the PEP-glyoxylate cycle flux on galactose. Five further transcription factors affected this flux only indirectly through cAMP and Crp by increasing the galactose uptake rate. Thus, E. coli actively limits its galactose catabolism at the expense of otherwise possible faster growth.

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Related in: MedlinePlus

Absolute (A) and substrate uptake normalized (B) flux changes of key metabolic pathways in the 91 mutants compared with the wild type during growth on glucose (Δ) and galactose (□). The dashed line indicates the wild-type reference fluxes.
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f3: Absolute (A) and substrate uptake normalized (B) flux changes of key metabolic pathways in the 91 mutants compared with the wild type during growth on glucose (Δ) and galactose (□). The dashed line indicates the wild-type reference fluxes.

Mentions: Next we focussed on flux changes that occurred for key metabolic pathways in the 91 mutants (Figure 3). On glucose, transcription factor deletions altered many fluxes, including those of hexose uptake, glycolysis, pyruvate dehydrogenase, TCA cycle and acetate secretion (Figure 3A). On galactose, these flux changes were even larger and additionally included the glyoxylate shunt. These variations in absolute flux values may be caused by either a change of the overall flux magnitude (due to or resulting in increased carbon uptake) or changes in the distribution of flux within the network; hence, affecting specific flux magnitudes. To distinguish between these two possibilities, we also compared fluxes normalized to the uptake rate between wild type and the 91 mutants (Figure 3B). Many transcription factors affected absolute uptake rates. About 60 of the 91 transcription factor mutants exhibited more than a 10% deviation in uptake rates compared with the wild type under either condition, indicating the importance of transcriptional regulation in controlling metabolic flux. The distribution of key fluxes through glycolysis, PP and Entner–Doudoroff (ED) pathway in upper metabolism, however, remained constant under both conditions (Figure 3B). Thus, flux variations in upper metabolism were caused by an altered magnitude of overall fluxes, but our data do not allow differentiating between a putative-specific transcriptional control of pathways in upper metabolism and indirect, growth-related effects. In key fluxes of lower metabolism (pyruvate dehydrogenase, anaplerosis, TCA cycle, glyoxylate cycle, acetate secretion), not only absolute fluxes but also their distribution changed (Figure 3B). TCA cycle, glyoxylate shunt and acetate secretion fluxes normalized to uptake rates were significantly different in several mutants, indicating that in lower metabolism also the distribution of flux is transcriptionally controlled.


Large-scale 13C-flux analysis reveals distinct transcriptional control of respiratory and fermentative metabolism in Escherichia coli.

Haverkorn van Rijsewijk BR, Nanchen A, Nallet S, Kleijn RJ, Sauer U - Mol. Syst. Biol. (2011)

Absolute (A) and substrate uptake normalized (B) flux changes of key metabolic pathways in the 91 mutants compared with the wild type during growth on glucose (Δ) and galactose (□). The dashed line indicates the wild-type reference fluxes.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3094070&req=5

f3: Absolute (A) and substrate uptake normalized (B) flux changes of key metabolic pathways in the 91 mutants compared with the wild type during growth on glucose (Δ) and galactose (□). The dashed line indicates the wild-type reference fluxes.
Mentions: Next we focussed on flux changes that occurred for key metabolic pathways in the 91 mutants (Figure 3). On glucose, transcription factor deletions altered many fluxes, including those of hexose uptake, glycolysis, pyruvate dehydrogenase, TCA cycle and acetate secretion (Figure 3A). On galactose, these flux changes were even larger and additionally included the glyoxylate shunt. These variations in absolute flux values may be caused by either a change of the overall flux magnitude (due to or resulting in increased carbon uptake) or changes in the distribution of flux within the network; hence, affecting specific flux magnitudes. To distinguish between these two possibilities, we also compared fluxes normalized to the uptake rate between wild type and the 91 mutants (Figure 3B). Many transcription factors affected absolute uptake rates. About 60 of the 91 transcription factor mutants exhibited more than a 10% deviation in uptake rates compared with the wild type under either condition, indicating the importance of transcriptional regulation in controlling metabolic flux. The distribution of key fluxes through glycolysis, PP and Entner–Doudoroff (ED) pathway in upper metabolism, however, remained constant under both conditions (Figure 3B). Thus, flux variations in upper metabolism were caused by an altered magnitude of overall fluxes, but our data do not allow differentiating between a putative-specific transcriptional control of pathways in upper metabolism and indirect, growth-related effects. In key fluxes of lower metabolism (pyruvate dehydrogenase, anaplerosis, TCA cycle, glyoxylate cycle, acetate secretion), not only absolute fluxes but also their distribution changed (Figure 3B). TCA cycle, glyoxylate shunt and acetate secretion fluxes normalized to uptake rates were significantly different in several mutants, indicating that in lower metabolism also the distribution of flux is transcriptionally controlled.

Bottom Line: Despite our increasing topological knowledge on regulation networks in model bacteria, it is largely unknown which of the many co-occurring regulatory events actually control metabolic function and the distribution of intracellular fluxes.While 2/3 of the regulators directly or indirectly affected absolute flux rates, the partitioning between different pathways remained largely stable with transcriptional control focusing primarily on the acetyl-CoA branch point.Five further transcription factors affected this flux only indirectly through cAMP and Crp by increasing the galactose uptake rate.

View Article: PubMed Central - PubMed

Affiliation: Institute of Molecular Systems Biology, ETH Zurich, Zurich, Switzerland.

ABSTRACT
Despite our increasing topological knowledge on regulation networks in model bacteria, it is largely unknown which of the many co-occurring regulatory events actually control metabolic function and the distribution of intracellular fluxes. Here, we unravel condition-dependent transcriptional control of Escherichia coli metabolism by large-scale (13)C-flux analysis in 91 transcriptional regulator mutants on glucose and galactose. In contrast to the canonical respiro-fermentative glucose metabolism, fully respiratory galactose metabolism depends exclusively on the phosphoenol-pyruvate (PEP)-glyoxylate cycle. While 2/3 of the regulators directly or indirectly affected absolute flux rates, the partitioning between different pathways remained largely stable with transcriptional control focusing primarily on the acetyl-CoA branch point. Flux distribution control was achieved by nine transcription factors on glucose, including ArcA, Fur, PdhR, IHF A and IHF B, but was exclusively mediated by the cAMP-dependent Crp regulation of the PEP-glyoxylate cycle flux on galactose. Five further transcription factors affected this flux only indirectly through cAMP and Crp by increasing the galactose uptake rate. Thus, E. coli actively limits its galactose catabolism at the expense of otherwise possible faster growth.

Show MeSH
Related in: MedlinePlus