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. Author manuscript; available in PMC: 2011 Jul 30.
Published in final edited form as: Mol Cell. 2010 Jul 30;39(2):259–268. doi: 10.1016/j.molcel.2010.07.005

Figure 2. FANCM and FAAP24 are selectively required for MMC-induced RPA foci formation and RPA2 phosphorylation.

Figure 2

(A) Representative images showing the requirement of FANCM and FAAP24 for MMC-induced RPA foci assembly but not other replication stress. HeLa cells were treated with MMC (1 μM, 6 hr), HU (2 mM, 6 hr), CPT (500 nM, 6 hr) or UV (30 J/m2, 6 hr post irradiation) respectively at 48 hr after transfection with either FANCM or FAAP24 siRNAs. RPA foci were detected using anti-RPA2 antibody. (B) Quantification for RPA foci shown in (A). The percentage of cells containing RPA foci was determined by counting at least 200 cells from each sample. Data are represented as mean ± SD from three independent experiments. (C) Immunoblot showing that depletion of either FANCM or FAAP24 more severely reduced RPA2 phosphorylation following MMC treatment. HeLa cells were treated as described in (A). Different mobility of RPA2 caused by phosphorylation is indicated as “B” (baseline) and “H” (hyperphosphorylated), respectively. (D) Time-course of RPA2 phosphorylation in FAAP24- or FANCM-depleted cells. HeLa cells transfected with indicated siRNAs were harvested at 0, 6, or 12 hr post treatment with MMC (1 μM) or HU (2 mM). (E) Dose-response of RPA2 phosphorylation in FAAP24- or FANCM-depleted cells. HeLa cells transfected with siRNAs were treated with MMC or HU at indicated concentrations for 8 hr.