Fig. 1: Inhibiting Arp2/3 activity lowers DSB mobility.
A Stills from a 20-min time course of a budding yeast cell 3 h after DSB induction with Gal-HO. Green focus is Ddc2-GFP, and red foci are Spc42-mCherry. B Diagram of experimental setup to measure the MSD of a DSB labeled with Ddc2-GFP. CK-666 (100 µM) or CK-689 (100 μM) was added 2 h 40 min after adding galactose. C Mean-squared displacement (MSD) analysis (∆t = 30 s) of the DSB relative to the SPB in FZ015 (WT) strain (n = 23), WT strain with CK-689 (100 μM) (n = 11), and WT strain with CK-666 (100 µM) (n = 28) 3 h after galactose induction of an HO endonuclease-induced irreparable DSB at the MAT locus on chromosome III. Resected DSBs were bound by Ddc2-GFP, and the SPBs were labeled with Spc42-mCherry. Cells were imaged with a 488 nm and 561 nm laser, taking optical stacks of 1.5 µm with a step size of 300 nm every 30 s for 20 min. See “Methods” section for MSD analysis. Mean ± SEM is shown. Source data are provided as a Source Data file. D Radius of confinement (Rc) from MSD analysis of strains in (C). Statistical analysis for the radius of confinement derived in PRISM using a one-way Anova (ns p ≥ 0.05, *p < 0.05, **p < 0.01, and ***p < 0.001). WT vs. WT + CK-666 (p = 0.0082) and WT vs. WT + CK-689 (p = 0.4112). See Supplementary Data 1 and 2 for significance and RC values. Mean ± SEM is shown. Source data are provided as a Source Data file. E MSD analysis (∆t = 30 s) of the DSB relative to the SPB in FZ015 (n = 10) and FZ077 (sla2∆) (n = 20) 3 h after Gal-HO induction. Imaging and analysis done as described in (C). Mean ± SEM is shown. Source data are provided as a Source Data file. F Rc from MSD analysis of strains in (E). Statistical analysis was done with a two-sided unpaired t-test. WT vs. sla2∆ (p = 0.8251). See Supplementary Data 1, 2 for significance and RC values. Mean ± SEM is shown. Source data are provided as a Source Data file.
