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. Author manuscript; available in PMC: 2013 Jun 7.
Published in final edited form as: Cell. 2012 Nov 29;151(6):1270–1282. doi: 10.1016/j.cell.2012.10.046

Figure 6. Diffusion barriers are crucial for bacterial fitness.

Figure 6

(A) Rate of periplasmic accumulation of an inducible PstS-mCherry protein fusion in the wild-type (EK363) and ΔstpAB (EK389) background. Cells were grown in HIGG-30 μM phosphate with 0.3% glucose to induce long stalks while repressing the synthesis of PstS-mCherry. The cells were seeded on pads with 0.3% xylose, and PstS-mCherry accumulation was monitored by timelapse microscopy. Mean fluorescence/cell body area was measured for ~ 300 cells per strain. Error bars = SD. Fitting the data to an exponential function and solving the equations in the exponential region (t = 165 to 255 min) for equal fluorescence intensities yielded a time difference in the accumulation of PstS-mCherry of 22.4 (± 0.8) min.

(B) Competitive growth of wild-type and diffusion barrier-deficient cells. To analyze the effect of diffusion barriers on the rate of recovery from nutrient starvation, wild-type, ΔstpAB and ΔstpCD cells were differentially labeled with the fluorescent proteins YFP (EK392, EK417, EK486) or mCherry (EK416, EK393, EK487) and starved for either phosphate or nitrogen. Mutant and wild-type cells were combined at equal ratios, transferred to nutrient-replete medium and grown to late-exponential phase. More than 1000 cells were analyzed by fluorescence microscopy before and after recovery to determine shifts in the relative composition of the cultures. The differentials were then used to calculate the lag in the onset of cell division (see Figure S6B). Values represent the average of four experiments, including two in which the fluorescent marker was switched (error bars = SD; * p < 0.02; ** p < 0.002).

(C) Wild-type cells outcompete a diffusion barrier-deficient mutant. Wild-type and ΔstpAB cells (SW51) were grown in PYE and mixed at equal optical densities. Mixed cultures (n = 5) were repeatedly diluted into fresh PYE and cultured to stationary phase. At the indicated timepoints, cells were withdrawn and spread on PYE agar. The ratio of wild-type and barrier-deficient cells was determined by colony PCR. Error bars show SD (n ~ 450). See also Figure S6.