DOI: 10.5061/dryad.8cz8w9gk9
title: Diversity in lac Operon Regulation among Diverse Escherichia coli Isolates Depends on the Broader Genetic Background but Is Not Explained by Genetic Relatedness

description: Transcription of bacterial genes is controlled by the coordinated
 action of cis- and trans-acting regulators. The activity and mode
 of action of these regulators can reflect different requirements
 for gene products in different environments. A well-studied
 example is the regulatory function that integrates the
 environmental availability of glucose and lactose to control the
 Escherichia coli lac operon. Most studies of lac operon
 regulation have focused on a few closely related strains.
 To determine the range of natural variation in lac regulatory
 function, we introduced a reporter construct into 23 diverse E.
 coli strains and measured expression with combinations of inducer
 concentrations. We found a wide range of regulatory
 functions. Several functions were similar to the one observed in
 a reference lab strain, whereas others depended weakly on the
 presence of cAMP. Some characteristics of the regulatory function
 were explained by the genetic relatedness of strains, indicating
 that differences varied on relatively short time scales. The
 regulatory characteristics explained by genetic relatedness were
 among those that best predicted the initial growth of strains
 following transition to a lactose environment, suggesting a role
 for selection. Finally, we transferred the lac operon, with the
 lacI regulatory gene, from five natural isolate strains into a
 reference lab strain. The regulatory function of these
 hybrid strains revealed the effect of local and global regulatory
 elements in controlling expression. Together, this work
 demonstrates that regulatory functions can be varied within a
 species and that there is variation within a species to best match a
 function to particular environments.
description: A gfp reporter driven by the lac promoter (Plac-GFP) was
 integrated into the attTn7 site of strains as indicated in file
 names. Regulatory input functions were characterized by measuring the
 expression of a Plac-GFP reporter at different combinations of cAMP and
 IPTG in DM supplemented with 2,000 g/ml glucose. This environment was used
 because glucose inhibits the production of cAMP, allowing measurement of
 the regulatory input function from as close to the basal level of
 Plac-GFP expression as possible. Strains containing the Plac-GFP reporter
 were preconditioned in DM medium supplemented with 2,000 g/ml (DM2000)
 glucose for 24 h and then transferred at a 1:1,000 dilution to the test
 environments containing combinations of DM2000 supplemented with cAMP and
 IPTG. cAMP was added at eight concentrations (0, 0.625, 1.25, 2.5, 5, 10,
 20, and 40 mM), and IPTG was added at 10 or 6 concentrations (0, 0.78,
 1.56, 3.125, 6.25, 12.5, 25, 50, 100, and 200 uM, or 0, 6.25, 12.5, 25,
 50, and 100 uM). Strains were grown in these environments for 16 h to an
 optical density at 450 nm (OD450) of 0.1 to 0.2, which corresponded to
 mid-log growth phase, as determined by tracking changes in population OD
 using a VersaMax spectrophotometer.  
description: File names give the name of the measured strains followed by the
 replicate number (if any). Files contain a matrix of expression values
 determined using a flow cytometer to sample induced cell populations and
 reported as the median value over the measured reporter expression
 distribution for each inducer combination. cAMP concentrations increase
 through rows 1-8 and IPTG concentrations increase therough columns 1-6 or
 1-10 as relevant to individual files. Concentrations are: cAMP 0, 0.625,
 1.25, 2.5, 5, 10, 20, and 40 mM and IPTG 0, 0.78, 1.56, 3.125, 6.25, 12.5,
 25, 50, 100, and 200 uM or 0, 6.25, 12.5, 25, 50, and 100 uM.