Figure 1.

Single-cell Dynamics of a Cationic Dye Revealed a Gradual Increase on Forespores and Sudden Drop upon Mother-Cell Lysis

(A) Illustrative diagram showing the membrane polarities of a sporulating cell. Engulfment process results in forespore membranes with opposite polarities to each other. Quality control works during late sporulation (stage IV-VI) where protective layers are assembled and matured.

(B) Time series of the mean intensities of phase contrast (gray) and ThT (orange) in spore region. The data correspond to the panel (C). ROIs (region of interest) used for the measurements are shown in Figure S1. ThT was added at 10 μM.

(C) Film-strip images showing the phase-contrast (upper) and ThT (lower) of a live single-cell sporulating cell. Scale bar, 1 μm. Color scale for ThT intensity is shown in the left-end panel for ThT. Film-strip is a representative of successfully sporulating cells from twenty-nine independent experiments.

(D) Time series of single-cell ThT fluorescence dynamics from 122 forespores. Forespore regions of ThT intensity were measured until 1 h after mother-cell lysis. All time series qualitatively follow the pattern represented in panel (C) but a great degree of heterogeneity between cells. The dashed line represents the value used for alignment in panel (E).

(E) The dataset shown in panel (D) was aligned to the frame it first reaches ThT intensity value 1,000. Time series after reaching maximum are highlighted in red. ThT increases at a relatively finite rate. A great degree of heterogeneity in timing of mother-cell lysis was observed (red lines). Histograms of the late-sporulation duration with and without ThT are shown in Figures S3 and S4.