Fig 1.
Experimentally tracking single cells during glucose depletion assays.
A) Cells were trapped within a microfluidic flow chamber and the environmental concentration of glucose was depleted as a function of time, while holding the galactose concentration constant. As glucose levels dropped, individual cells heterogeneously activated Gal1p production, and the resulting fluorescence trajectories were recorded. B) Glucose concentration as a function of time (red line). Here, the depletion time is 4 hrs. Also shown is the experimentally measured fluorescence trajectory of an individual cell to the 4 hr depletion time (green line). This cell first initiates Gal1p production and then accumulates protein (below). C) Images of yeast cells in the microfluidic device at successive events from a 4 hr glucose-depletion assay. [glu] = 2%, t = 0 (hr) indicates the beginning of glucose depletion; [glu] = 0% indicates total depletion of glucose; labels FI = 0 and 200 (AU) correspond to the times at which FI reaches these values; end of run, the end of glucose depletion assay.
Fig 2.
Experimentally measured single-cell responses to different glucose-depletion times.
Ten glucose-depletion times were tested. Red lines depict changes in glucose concentration from 2% to 0% (w/v). Gray curves depict individual fluorescence trajectories of Gal1-YFP. Green curves show mean FI. For reference, the fluorescence of cells grown in just galactose was approximately 8200 ± 2000 AU (S1 Dataset, sheet 14).
Fig 3.
The initiation of GAL1 expression depends linearly on the glucose depletion time.
A) The mean initiation time increases linearly with increasing depletion time. Shown are the mean (dots), standard deviations (error bars), and best fit line. B) The standard deviation of the initiation time is related to the variability of the threshold g* and the delay time τ according to Eq (2). Error bars represent the standard deviation of the measurement as calculated with a bootstrapping method. C) The dependence of both the mean and the standard deviation of the initiation time can be explained by a simple threshold model. As glucose depletes (red curves, left) a variable threshold (green distribution, right) is crossed that relieves repression of the galactose genes. The slower the transition through the repression thresholds (horizontal shaded region) the greater the variability in the initiation time (vertical shaded region).
Fig 4.
Gal1p accumulation changes non-monotonically with glucose depletion rate.
A) Average accumulation time as a function of the depletion time (error bars represent SD). B) The standard deviation of the accumulation time as a function of the depletion time. In fast and slow depletion schemes, the accumulation of Gal1p is highly variable, and achieves a minimum at intermediate depletion rates. Error bars represent the standard deviation of the measurement as calculated with a bootstrapping method.
Fig 5.
Cell cycle times during glucose depletion.
A) Mean and standard deviation (error bars) of cell cycle times during growth in glucose (before glucose depletion) for different depletion times. B- D) Histograms of the cell cycle times shown in (A) for three different depletion times (0 hr, 2 hr and 8 hr, respectively). E-H) Same as (A-D), but during growth in the diauxic transition between glucose and galactose (during glucose depletion). G-L) Same as (A-D), but for growth in galactose (after glucose depletion).
Fig 6.
An energy model for the glucose/galactose switch.
A) A schematic of the regulation in the heuristic stochastic model. Glucose inhibits expression of the GAL genes, but increases the cellular energy needed for protein production. Gal4p up-regulates transcription of gal2, whose gene product, Gal2p, imports galactose. The ability of the cell to metabolize galactose is therefore dependent on the availability of Gal2p. In addition, intercellular galactose increases cellular energy and up-regulates gal2. B) We used piecewise linear functions for Eglu([glu]) and Egal([Gal2p]). The Eglu term is assumed to only depend on the environmental glucose concentration. The Egal term does not depend on environmental galactose because it is held constant; what changes is the amount of galactose that is utilized by the cell, a good proxy for which is the concentration of Gal2p. The maximum (steady-state) energy levels and
are inferred from the cell-cycle lengths in the two conditions. The threshold thglu is the glucose threshold at which the cell is assumed to obtain maximal energy. The equivalent threshold for galactose is thgal; this is the threshold at which the cell has sufficient Gal2p to maximally utilize environmental galactose.
Fig 7.
Results of the mathematical model.
A) Simulated standard deviation of the accumulation time as a function of the depletion time for the full model (red dots) and a model lacking the energy scaling (blue dashed line). Note that the energy scaling recreates the non-monotonicity observed in the experiments. B) Box plots of the distributions of the different types of cell cycle lengths for the 3 hr depletion scheme. The red box plots are the experimental data, whereas the blue box plots are obtained from a simulation of Eq. (5). The distribution of the energy availability in the diauxic phase (ϵ) is obtained by fitting the model to the diauxic cell cycle data.