Heat shock inhibits radiation-induced activation of NF-κB via inhibition of I-κB kinase

Radiation stimulates signaling cascades that result in the activation of several transcription factors that are believed to play a central role in protective response(s) to ionizing radiation (IR). It is also well established that heat shock alters the regulation of signaling cascades and transcription factors and is a potent radiosensitizing agent. To explore the hypothesis that heat disrupts or alters the regulation of signaling factors activated by IR, the effect of heat shock on IR-induced activation of NF-kappa B was determined. Irradiated HeLa cells demonstrated transient increases in NF-kappa B DNA binding activity and NF-kappa B protein nuclear localization. In addition, irradiated cells demonstrated increased I-kappa B phosphorylation and decreased I-kappa B alpha cytoplasmic protein levels, corresponding temporally with the increase of NF-kappa B DNA binding. Heat shock prior to IR inhibited the increase in NF-kappa B DNA binding activity, nuclear localization of NF-kappa B, and the phosphorylation and subsequent degradation of I-kappa B. I-kappa B kinase (IKK) immunoprecipitation assays demonstrated an increase in IKK catalytic activity in response to IR that was inhibited by pretreatment with heat. Kinetic experiments determined that heat-induced inhibition of NF-kappa B activation in response to LR decayed within 5 h after heating. Furthermore, pretreatment with cycloheximide, to block de novo protein synthesis, did not alter heat shock inhibition of IR induction of NF-kappa B, These experiments demonstrate that heat shock transiently inhibits IR induction of NF-kappa B DNA binding activity by preventing IKK activation and suggests a mechanism independent of protein synthesis.