Figure 4. DNA2 depletion does not affect formation of FANCD2 foci after cisplatin exposure. (A) U2OS cells with endogenous DNA2 depletion were treated with cisplatin (15 μM, 24 h) and stained with anti-FANCD2 or γH2Ax antibodies. A representative immunofluorescence image of FANCD2 foci is shown. (B) Quantification of FANCD2 foci-positive cells that are also positive for γH2Ax foci. (C) FANCD2 is monoubiquitylated in DNA2-depleted A549 cells treated with or without formaldehyde. (D) PD20 FANCD2^−/− complemented with vector, FANCD2 wild-type, or FANCD2-K561R cDNAs were treated with formaldehyde and western blots prepared. Formaldehyde-treated cells showed FANCD2 monoubiquitylation, and FANCD2-K561R mutants were not monoubiquitylated. Error bars indicate mean ± SEM for n ≥ 2 independent experiments.