Figure 4.

CHD3 is recruited to sites of DNA damage and is required for DNA damage induced chromatin relaxation. (A) Recruitment kinetics of GFP-CHD3 (red) and GFP-CHD4 (black) to sites of DNA damage. (B) Recruitment of GFP-CHD3 to sites of microirradiation 120 seconds after DNA damage in the presence (PARPi) or absence (CTRL) of PARP inhibitor. Microirradiation is highlighted with green arrows. Scale bars are 5 μm. (C) GFP-CHD3 is tethered to the LacO array and co-expressed with mCherry-PARP1. Upon DNA damage, PARP1 recruits to microirradiation induced DNA breaks but does not accumulate at the tethered GFP-CHD3, indicating that CHD3 does not bind PAR. Scale bars are 5 μm. Inset shows the magnified LacO array. (D) Effect of two independent CHD3 (siCHD3) knockdowns on DNA damage-induced chromatin relaxation as compared to scrambled siRNA treatment (siScr). Boxplots show chromatin relaxation 120 seconds after microirradiation (E) Western blot showing knockdown efficiency of CHD3 and expression levels of CHD4, PARP1 and Alc1. Tubulin is used as a loading control.