Figure 7.

Phosphorylation of ATM kinase and its substrates in PML-RARα-expressing cells. (a) Immunoblot analysis of pSer1981-ATM, pSer343-NBN, and pThr68-CHK2 phosphorylation in APL blasts, NB4, NB4-MR4, and U937/PR9 cells. Previous to IR, NB4 and NB4-MR4 cells were either untreated or exposed to 1 μM RA for 72 h, whereas U937/PR9 cells were either untreated or exposed for 8 h to 100 μM ZnSO₄, and then treated or not with 1 μM RA for 72 h. Cells were then exposed to 1 Gy of X-rays and lysed after 0.5, 3, and 24 h. (b) Immunoblot analysis of ATM phosphorylation at the Ser1981 residue after 24 h from IR in NB4 and U937/PR9 cells. (c) U937/PR9 cells were first treated with 100 μM of ZnSO₄, then exposed to 10 μg/ml cycloheximide, and finally irradiated with 1 Gy and lysed after 0.5 h. Immunoblots were performed using anti-γ-H2AX, anti-pSer343-NBN, anti-NBN, anti-pThr68-CHK2, and anti-CHK2 antibodies. Blots presented are exemplificative of the results obtained from two independent experiments