title: The relationship between external glucose concentration and cAMP levels inside Escherichia coli: implications for models of phosphotransferase-mediated regulation of adenylate cyclase.
abstract: The concentration of glucose in the medium influences the regulation of cAMP levels in Escherichia coli. Growth in minimal medium with micromolar glucose results in 8- to 10-fold higher intracellular cAMP concentrations than observed during growth with excess glucose. Current models would suggest that the difference in cAMP levels between glucose-rich and glucose-limited states is due to altered transport flux through the phosphoenolpyruvate: glucose phosphotransferase system (PTS), which in turn controls adenylate cyclase. A consequence of this model is that cAMP levels should be inversely related to the saturation of the PTS transporter. To test this hypothesis, the relationship between external glucose concentration and cAMP levels inside E. coli were investigated in detail, both through direct cAMP assay and indirectly through measurement of expression of cAMP-regulated genes. Responses were followed in batch, dialysis and glucose-limited continuous culture. A sharp rise in intracellular cAMP occurred when the nutrient concentration in minimal medium dropped to approximately 0.3 mM glucose. Likewise, addition of > 0.3 mM glucose, but not < 0.3 mM glucose, sharply reduced the intracellular cAMP level of starving bacteria. There was no striking shift in growth rate or [14C] glucose assimilation in bacteria passing through the 0.5 to 0.3 mM concentration threshold influencing cAMP levels, suggesting that neither metabolic flux nor transporter saturation influenced the sensing of nutrient levels. The (IIA/IIBC)Glc PTS is 96-97% saturated at 0.3 mM glucose so these results are not easily reconcilable with current models of cAMP regulation. Aside from the transition in cAMP levels initiated above 0.3 mM, a second shift occurred below 1 muM glucose. Approaching starvation, well below saturation of the PTS, cAMP levels either increased or decreased depending on unknown factors that differ between common E. coli K-12 strains.
authors.affiliation: Department of Microbiology G08, University of Sydney, Sydney, New South Wales 2006, Australia.
authors.forename: Lucinda
authors.initials: L
authors.lastname: Notley-McRobb
authors.affiliation: Department of Microbiology G08, University of Sydney, Sydney, New South Wales 2006, Australia.
authors.forename: Alison
authors.initials: A
authors.lastname: Death
authors.affiliation: Department of Microbiology G08, University of Sydney, Sydney, New South Wales 2006, Australia.
authors.forename: Thomas
authors.initials: T
authors.lastname: Ferenci
mesh_terms: D000262:Adenylyl Cyclases / Q000378:metabolism; D001693:Biological Transport, Active; D003470:Culture Media; D000242:Cyclic AMP / Q000378:metabolism*; D003956:Dialysis / Q000379:methods; D004789:Enzyme Activation; D004926:Escherichia coli / Q000737:chemistry / Q000201:enzymology / Q000378:metabolism*; D015964:Gene Expression Regulation, Bacterial / Q000235:genetics; D005947:Glucose / Q000493:pharmacokinetics*; D008954:Models, Biological; D010770:Phosphotransferases / Q000494:pharmacology; D013217:Starvation / Q000378:metabolism
chemical_list: D003470:Culture Media; D000242:Cyclic AMP; D010770:Phosphotransferases; D000262:Adenylyl Cyclases; D005947:Glucose