Enhanced phosphorylation of Nbs1 on serine 343 in BCR/ABL-positive cells on DNA damage. (A) M07e and UT7 parental (P) and their BCR/ABL (B/A) counterparts were treated with 0.5 μg/mL MMC in the presence or absence of SCF. pNbs1, total Nbs1, Mre11, and RAD50 were examined by Western-blot analysis at the indicated time points during MMC treatment. (B) M07e (P) and their BCR/ABL (B/A) counterparts were treated with 0.3 μg/mL cisplatin (Cis) or with 10 mM hydroxyurea (HU) for 12 hours in the presence of SCF. pNbs1, Nbs1, Mre11, and RAD50 were examined by Western-blot analysis. (C) CD34⁺ cells were isolated from healthy donors (Normal) and CML-CP and CML-BC patients. Cells were treated or not with 0.5 μg/mL MMC for 12 hours in the presence of GM-CSF and SCF. pNbs1, Nbs1, Mre11, RAD50, and tubulin were detected by Western blot analysis. Results are representative for 3 donors in each group. (D) M07e (■) and B/A-M07e (▩) cells and (E) CD34⁺ cells from healthy donors (■) and CML-BC patients (▩) were untreated (left panels) or treated (right panels) with 0.5 μg/mL MMC for 12 hours in the presence of GM-CSF and SCF. pNbs1 foci were detected in the nuclei by immunofluorescence; results represent mean (± SD) of the number of cells classified as negative (< 4 foci), low-positive (4-20 foci), or high-positive (> 20 foci). *P < .05, ▩ versus ■ in particular groups. (F) Representative nuclei scored as negative, low-positive, and high-positive for pNbs1 foci; nuclear borders are outlined in blue.