American Society of Hematology

Figure 2

$Figure 2. FANCA knockdown results in marked reduction of FANCD2 monoubiquitination and nuclear foci formation. (A) FANCAi and LUCi hESCs were treated with 2mM HU for 24 hours, and lysates were subjected to Western blotting for FANCD2. FANCD2-L is the active, monoubiquitinated form of FANCD2-S. The ratio of L/S was calculated using ImageJ software. (B) FANCD2 immunofluorescence of FANCAi and LUCi hESCs treated with 200nM MMC treatment for 24 hours. DAPI (4,6 diamidino-2-phenylindole) stains for DNA content and GFP marks infected cells. (C) Quantification of FANCD2 nuclear foci after 100, 200, and 400 nM MMC treatment for 24 hours. Percentage FANCD2 nuclear foci (y-axis) represents the fraction of cells with 4 or more nuclear foci, as assayed by FANCD2 immunofluorescence. Data are represented as the average ± SEM; each data point represents 3 independent sets of 50 cells. ***P < .01 by Student t test.$

FANCA knockdown results in marked reduction of FANCD2 monoubiquitination and nuclear foci formation. (A) FANCAi and LUCi hESCs were treated with 2mM HU for 24 hours, and lysates were subjected to Western blotting for FANCD2. FANCD2-L is the active, monoubiquitinated form of FANCD2-S. The ratio of L/S was calculated using ImageJ software. (B) FANCD2 immunofluorescence of FANCAi and LUCi hESCs treated with 200nM MMC treatment for 24 hours. DAPI (4,6 diamidino-2-phenylindole) stains for DNA content and GFP marks infected cells. (C) Quantification of FANCD2 nuclear foci after 100, 200, and 400 nM MMC treatment for 24 hours. Percentage FANCD2 nuclear foci (y-axis) represents the fraction of cells with 4 or more nuclear foci, as assayed by FANCD2 immunofluorescence. Data are represented as the average ± SEM; each data point represents 3 independent sets of 50 cells. ***P < .01 by Student t test.

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