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. 2019 Sep 15;30(20):2584–2597. doi: 10.1091/mbc.E18-10-0650

FIGURE 2:

FIGURE 2:

Damage-induced increase of the bound to free NADH ratio is PARP1-dependent. (A) Immunofluorescence staining for PAR (left) and γH2AX (right) in HeLa cells fixed at various time points up to 12 h postdamage following low, medium, and high input laser power. Scale bar = 5 µm. (B) The fraction of bound NADH and intensity of NADH over time in the cytoplasmic and nuclear compartments of HeLa cells treated with 0.1% DMSO (top), 20 µM PARP inhibitor (olaparib) (middle), or 10 µM ATM inhibitor (KU55933) and 10 µM DNA-PK inhibitor (NU7026) (bottom). (C) The percent change in the fraction of bound NADH (top) and the intensity of NADH (bottom) at 2 h (left) and 12 h (right) postdamage relative to basal conditions. N = 20. *p < 0.05, **p < 0.01, ***p < 0.001. (D) The change in fraction of bound NADH over time in the cytoplasmic (left) and nuclear (middle) compartments of HeLa cells with siPARP1 or control siRNA. N = 15. Western blot analysis (right) of siControl and siPARP1 transfected HeLa cells. The whole cell extracts were run on SDS–PAGE and blotted with anti-PARP1 antibody. Anti-actin antibody served as loading control.