Effects of nutrition and growth rate on Lrp levels in Escherichia coli.
Lrp (leucine-responsive regulatory protein) activates some Escherichia coli operons that function in anabolism and represses others involved in catabolism (for a review, see J. M. Calvo and R. G. Matthews, Microbiol. Rev. 58:466-490, 1994). This overall pattern suggests that Lrp may help cells adapt to changes in the nutritional environment. Here, we tested the idea that the nutritional richness of the medium determines the amount of Lrp in cells. Lrp was measured directly by Western blotting (immunoblotting) in cells grown in a chemically defined rich medium or in a minimal medium. In addition, transcription from the lrp promoter was assessed with a lacZ reporter gene. The results with these two different measurements were nearly the same, indicating that under the conditions employed, beta-galactosidase measurements can accurately reflect Lrp levels. For cells in a minimal medium, Lrp levels were consistently lowest during the logarithmic phase of growth, but overall, there was not much variation in levels as a function of growth phase (1.3-fold difference between highest and lowest values). However, for cells in a rich medium, Lrp levels dropped 3- to 4-fold during the lag phase, remained constant during the log phase, and then rose to starting levels upon entry into the stationary phase. When cells in the log phase were compared, Lrp levels were 3- to 4-fold higher in cells growing in a minimal medium than those in a rich medium. The levels of lrp expression were the same or slightly higher in strains containing mutations in rpoS, cya, or crp compared with wild-type strains, suggesting that neither RpoS nor the cyclic AMP (cAMP) receptor protein-cAMP complex is required for expression. On the other hand, lrp expression was severely restricted in cells that could not make ppGpp because of mutations in relA and spoT. The reduced expression of lrp during logarithmic growth in a rich medium may be due to low ppGpp levels under these conditions. The repressive effects of rich medium and the stimulatory effects of ppGpp were also observed with a construct having only a minimal lrp promoter (-57 to +21). The results of other experiments suggest that Lrp levels vary inversely with the growth rate of cells instead of being determined by some component of the medium.
Landgraf
J R
JR
Section of Biochemistry, Molecular and Cell Biology, Cornell University, Ithaca, New York 14853, USA.
Wu
J
J
Calvo
J M
JM
D000262:Adenylyl Cyclases / Q000235:genetics; D001426:Bacterial Proteins / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*; D003470:Culture Media; D002373:Cyclic AMP Receptor Protein / Q000235:genetics; D004268:DNA-Binding Proteins / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*; D004926:Escherichia coli / Q000254:growth & development / Q000378:metabolism*; D029968:Escherichia coli Proteins; D015870:Gene Expression; D015964:Gene Expression Regulation, Bacterial; D017930:Genes, Reporter; D006159:Guanosine Tetraphosphate / Q000494:pharmacology; D051756:Leucine-Responsive Regulatory Protein; D011401:Promoter Regions, Genetic; D012808:Sigma Factor / Q000235:genetics; D014157:Transcription Factors
D001426:Bacterial Proteins; D003470:Culture Media; D002373:Cyclic AMP Receptor Protein; D004268:DNA-Binding Proteins; D029968:Escherichia coli Proteins; C497278:Lrp protein, E coli; D012808:Sigma Factor; D014157:Transcription Factors; C069567:sigma factor KatF protein, Bacteria; D051756:Leucine-Responsive Regulatory Protein; D006159:Guanosine Tetraphosphate; D000262:Adenylyl Cyclases