Figure 7
Quantification of LDs from THG images of Phycomyces blaekseneanus hypha. Hypha were cultured without nitrogen or in standard liquid media for 2–6 h (or longer up to 8 h) after the start of treatment. Obtained THG images of LDs were analyzed by PA. n = 6 independent cultures. (a) N-starvation increases number of LDs per unit area. LD number obtained from the individual hypha is normalized to hypha area (in 103 µm2). Control (n = 44), N-starved group (n = 17). The box and whisker plots, enclosed by the 25th and 75th percentile range, median line with whiskers extending minimal to maximal value. Unpaired t test with Welch's correction, two tail, p = 0.0038. (b) Time course of LD number/unit area, showing that the increase of LD number by N-starvation is significant at 4.5 h (p = p = 0.0006) and later times (p = 0.0045), compared to corresponding control. Two-way ANOVA, with Holm-Sidac correction. Mean ± SE, n(Control) = 8; 7; 11; 21 for time points (in h), respectively: 2; 3; 4.5; 6. n(N-starved) = 6; 3; 7 for time points (in h), respectively: 3; 4.5; 6. (c) N-starvation decreases diameter of LDs. LD diameters from Control (n = 1205) and N-starved group (n = 431). The box and whisker plots, enclosed by the 25th and 75th percentile range, median line with whiskers extending minimal to maximal value. Mann–Whitney (p = 0.0008), two-tailed. (d) Time course of LD diameter changes, showing that the decrease by N-starvation is significant only at long starvation times. Two-way ANOVA, Holm-Sidac correction (p < 0.0001), compared to corresponding control. Mean ± SE, n(Control) = 176; 124; 302; 571 for time points (in h), respectively: 2; 3; 4.5; 6. n(N-starved) = 100; 118; 214 for time points (in h), respectively:3; 4.5; 6. (e) Differential distribution of increased number of LDs after 4.5 h and 6 h N-starvation. The largest LDs are lost at longest starvation times. Histograms of LD diameter distributions, 0.3 µm binning, for Control and N-starved group. Number of LDs in each bin of the histogram is divided by sum of hypha area of the appropriate group. Errors are calculated as stated in Methods section. Numbers on x axes represent the upper bin limit.
