Fig. 4: Altered molecular packing in the presence of ATP and post-translational methylation.
a Single-droplet steady-state anisotropy as a function of increasing ATP concentration measured within individual FUS-eGFP condensates (n = 50 individual droplets from 3 independent experiments). b Representative anisotropy images of FUS-eGFP droplets formed in the absence of ATP (control) (scale bar 3 µm) and presence of 10 mM ATP (scale bar 2 µm). c The respective FRET efficiency estimated from the single-droplet steady-state anisotropy values (a) in the presence of varying ATP concentrations (n = 50 individual droplets from 3 independent experiments). d Representative time-resolved anisotropy decay measured within FUS-eGFP droplets yielded biexponential (0 mM and 10 mM ATP) and triexponential decay kinetics (5 mM ATP). Data for 0 mM ATP for FUS-eGFP droplet is shown in Fig. 2f and included here for comparison. e–f Single-droplet steady-state anisotropy plots with representative anisotropy images (scale bar 4 µm and 2 µm for unmethylated and methylated droplets) (e) and estimated homoFRET efficiencies (f) obtained from unmethylated (control) and methylated FUS-eGFP droplets (n = 57 and 90 individual droplets for unmethylated and methylated FUS-eGFP respectively, from 3 independent experiments). g Representative time-resolved anisotropy decay obtained within unmethylated and methylated FUS-eGFP droplets yielded biexponential decay kinetics. h, i Changes in the amplitude of the unresolved ultrafast component (h) and the slower timescale of energy migration (i) recovered from anisotropy decay shown in (g) (n = 9 droplets from 3 independent experiments). Data for unmethylated FUS-eGFP droplets is shown here for comparison and is the same as data shown for 10 µM FUS-eGFP droplets in Fig. 2c, d, and f. Rotational correlation times and energy migration time constants are included in Supplementary Table 1. Data shown in box-and-whiskers plots indicates the 25th to 75th percentile (box), median (line), mean (white square), and 10th to 90th percentile (whiskers). The difference was statistically significant as determined by a one-way ANOVA test (for *** p ≤ 0.001 compared to the respective control). Source data are provided as a Source Data file.
