Regulation of DNA-dependent protein kinase activity by ionizing radiation-activated Ab1 kinase is an ATM-dependent process

Ionizing radiation (IR) treatment results in activation of the nonreceptor tyrosine kinase c-Ab1 because of phosphorylation by ATM. In vitro evidence indicates that DNA-dependent protein kinase (DNA-PK) can also phosphorylate and thus potentially activate Ab1 kinase activity in response to IR exposure. To unravel the role of ATM and DNA-PK in the activation of Ab1, we assayed Ab1, ATM, and DNA-PK activity in ATM- and DNA-PKcs-deficient cells after irradiation. Our results show that despite the presence of higher than normal levels of DNA-PK kinase activity, c-Ab1 fails to become activated after IR exposure in ATM-deficient cells. Conversely, normal activation of both ATM and c-Ab1 occurs in DNA-PKcs-deficient cells, indicating that ATM but not DNA-PK is required for activation of Ab1 in response to IR treatment. Moreover, activation of Ab1 kinase activity by IR correlates well with activation of ATM activity in all phases of the cell cycle. These results indicate that ATM is primarily responsible for activation of Ab1 in response to IR exposure in a cell cycle-independent fashion. Examination of DNA-PK activity in response to IR treatment in Ab1-deficient cells expressing mutant forms of Ab1 or in normal cells exposed to an inhibitor of Ab1 suggests an in vivo role for Ab1 in the down-regulation of DNA-PK activity. Collectively, these results suggest a convergence of the ATM and DNA-PK pathways in the cellular response to IR through c-Ab1 kinase.