Research Article | Applied and Environmental Science

Availability of the Molecular Switch XylR Controls Phenotypic Heterogeneity and Lag Duration during Escherichia coli Adaptation from Glucose to Xylose

Manon Barthe, Josué Tchouanti, Pedro Henrique Gomes, Carine Bideaux, Delphine Lestrade, Carl Graham, Jean-Philippe Steyer, Sylvie Meleard, Jérôme Harmand, Nathalie Gorret, Muriel Cocaign-Bousquet, Brice Enjalbert
Pablo Ivan Nikel, Invited Editor, Sang Yup Lee, Editor
DOI: 10.1128/mBio.02938-20

Article Figures & Data

Figures

  • FIG 1
    FIG 1

    The length of the glucose-xylose lags in E. coli is background dependent. Thirteen different strains of E. coli were cultivated by switching strains previously grown on M9 glucose medium to 96-well microplates filled with 200 μl of fresh M9 xylose or M9 glucose medium. (A) Lag times (bars) and maximum growth rates (μmax) (circles) after the switch to glucose (white) or xylose (gray) for the 13 E. coli strains (n = 9 with 3 technical replicates for each of the 3 biological replicates). (B) Scatterplot of the regrowth lag time on xylose or glucose as a function of the maximum growth rate on xylose or glucose for each strain (gray circles for xylose, white circles for glucose). (C) Scatterplot of glucose-xylose diauxic lag times versus regrowth lag times (cells grown overnight on glucose switched to xylose). For diauxic growth, strains previously grown on glucose were transferred to a glucose-xylose M9 medium (12.5% glucose and 87.5% xylose). Diauxic lags were calculated as explained in Materials and Methods. The negative regrowth lag time calculated for the BL21 strain was due to a higher growth rate when this strain resumed growth on xylose than its maximum growth rate during its exponential growth on xylose. For diauxic lags, n = 9 for strains BW25113, 1404, LF82, and S5Vir, and n = 6 for strains MG1655, E2348/69, E22, BL21(DE3), BW3070, CA244, M1/5, Nissle 1917, and SP15 strains.

  • FIG 2
    FIG 2

    XylR overexpression reduces regrowth lag time during the glucose-xylose shift of different E. coli strains. Strains carrying the pMET219 plasmid as a control (white symbols) or the same backbone plasmid with xylR expression under the control of the cysG constitutive promoter (pMET219_xylR; gray symbols), were grown like those shown in Fig. 1A (9 biological replicates but for strain E22 without xylR overexpression where n = 6; ** indicates a significant P value of <0.01 compared to the control strain not overexpressing xylR). The bars represent the lag time (in hours) before growth resumed. The circles represent the growth rates.

  • FIG 3
    FIG 3

    XylR titration modifies regrowth lag time. The ability of the E. coli BW25113 strain to resume growth with different plasmid constructs was assessed by switching cells after overnight culture on glucose to a 96-well microplate filled with 200 μl of fresh M9 xylose (A) or M9 glucose (B) medium. The BW25113 strain (in black) with the empty pSB1C3 plasmid as a control, the BW25113 strain (in blue) with a pSB1C3 plasmid carrying the xylA promoter, the BW25113 strain (in red) with a pSB1C3 plasmid expressing the red fluorescent protein mRFP1 under the control of the xylA promoter, and the BW25113 strain (in green) with a pSB1C3 expressing mRFP1 under the control of a xylA promoter deprived of its XylR binding sites. n = 6 for all conditions.

  • FIG 4
    FIG 4

    The length of the transition in the modified BW25113 strain is related to the emergence of a new subpopulation. Batch fermentations of the BW25113 strain transformed with the pSB1C3 plasmid carrying the PihfB-BFP PxylA-mRFP1 construct were carried out in minimum medium of 90 mMeqC and sampled at 30-min intervals. (A) Kinetics of substrate consumption and production and growth of the strain in M9 medium supplemented with a 40% glucose and 60% xylose mix. (B) Cytometric profiles of the 40% glucose−60% xylose batch over time. The y axis displays the blue fluorescence levels in arbitrary fluorescence units (a.f.u.), and the x axis displays the red fluorescence levels (a.f.u.). The blue gate represents the glucose population type as seen with the 100% glucose control. The red gate represents the xylose population type as seen with the 100% xylose control. (C) The theoretical biomass of each subpopulation was extrapolated from the percentages resulting from the flow cytometry analyses during growth on the 40% glucose and 60% xylose mix (glucose type cells in blue, xylose type cells in red, and the whole population in gray). (D) Same representation with 100% glucose growth. (E) Same representation with 100% xylose growth. Others substrate ratios are presented in Fig. S2 in the supplemental material.

  • FIG 5
    FIG 5

    Previous exposure to xylose creates a memory effect. The strain with the titration plasmid was grown on xylose and switched to a glucose-xylose mix on microplates with different concentrations of inoculant so that the number of generations on glucose differs between two growth phases on xylose. The duration of the diauxic lags are represented as a function of the number of generations on glucose. For each condition, four replicates were performed. n = 4 for all generation numbers but the highest (n = 2).

  • FIG 6
    FIG 6

    Modeling subpopulation behavior. (A) Scheme of the stochastic model. The diagram shows individual growth and transitions between cell types in the population. X1 corresponds to glucose consumers with the basal level of XylR, Y to xylose consumers with a high level of XylR, and X2 to glucose consumers that previously grew on xylose with an initially high but decreasing level of XylR. We assume that these mechanisms occur after a random time distributed according to an exponential law with the corresponding rates. Indeed, an individual that grows on glucose (class X1 or X2) divides at rate b1(S1) or switches to xylose consumption at rate λ(S) and (1+θ)λ(S), respectively. Specifically, in the X2 compartment, where cells have many xylR copies, each individual can give birth to a cell of the compartment X1 with probability 0 < α < 1 because of xylR dilution. In addition, an individual growing on xylose (class Y) divides at rate b2(S2) or switches to glucose consumption (class X2) at rate η(S1), if glucose is abundant. (B) Validation and predictions of the model (solid lines) compared to the experimental data (crosses) during growth on six glucose-xylose mixes (glucose cell type in blue, xylose cell type in red, glucose in green, and xylose in violet). The conditions 80% glucose−20% xylose and 20% glucose−80% xylose were not used to estimate the model parameters. The lines represent predictions, whereas in the other conditions, they represent validation.

Tables

  • TABLE 1

    Escherichia coli strains used in the study

    TABLE 1
  • TABLE 2

    Plasmids used in this study

    TABLE 2

Supplemental Material

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KEYWORDS

adaptation
Escherichia coli
subpopulations
heterogeneity
metabolic transition

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