FANCD2 mono-ubiquitination in FA-I and FA-J cells. Whole cell extract was analyzed by direct Western blotting, using affinity-purified FANCD2_1-292 antibody (A) or mouse anti-FANCD2_1-272 monoclonal antibody (B). (A) FANCD2-L is clearly detectable in FA-J cells, but apparently absent in FA-I cells; EUFA816 (FA-I) and EUFA543 (FA-J) cells (results not shown) were in agreement with the FA-I and -J cell lines shown in the figure. EUFA1289 (FA-D2) cell line was used as a negative control for FANCD2. (B) No induction of FANCD2-L in FA-I cells and normal induction in FA-J cells. Cells were either untreated or treated with MMC or hydroxyurea (HU) for 24 hours, before analysis by direct Western blotting.