Heat Inactivation of DNA-Dependent Protein Kinase: Possible Mechanism of Hyperthermic Radiosensitization

Abstract

Ionizing radiation combined with hyper-thermic treatment increases cell killing as the result of hyperthermic radiosensitization. DNA-dependent protein kinase (DNA-PK) is involved in the repair of radiation-induced DNA double-strand breaks. DNAPK consists of the catalytic subunit of DNA-PK (DNAPKcs) and Ku70/Ku80 heterodimers. To elucidate the mechanism of hyperthermic radiosensitization, we analyzed the heat sensitivity of DNA-PK activity in hybrid cells and the possible restoration of this activity with extracts from scid (defective in DNA-PKcs) and sxi-3 (defective in Ku80) cells. When hybrid cells were heated at 44°C for 15 min, DNA-PK activity was reduced to undetectable levels. As the cell extract from scid cells was added to the cell extract of heat-treated hybrid cells, the decreased DNA-PK activity could be concentration dependently restored with the addition of scid cell extract but not sxi-3 cell extract. Ku70/Ku80, but not Ku70 alone, could restore heat-inactivated DNA-PK. The inactivation of DNA-PK may play an important role in the process of inhibited repair of DNA double-strand breaks caused by hyperthermic radiosensitization.