Defective radiation signal transduction in ataxia-telangiectasia cells

Abstract

Purpose : The product of the gene ATM mutated in the human genetic disorder ataxia-telangiectasia (A-T) is predominantly present in the nucleus compatible with a role in DNA-damage recognition and cell-cycle control. However, ATM is also present outside the nucleus in cytoplasmic and membrane associated vesicles, which may explain the more general signalling defect in A-T. This led us to investigate signalling events initiated by ionizing radiation, remote from the nucleus. Materials and Methods : A-T and control lymphoblastoid cells were employed to study radiation-induced signalling at the level of protein activation using immunoprecipitation and immunoblotting. Flow cytometry was used to determine mobilization of intracellular Ca 2. Results : Lymphoblastoid cells from A-T patients were found to be defective in the radiation-induced activation of protein tyrosine kinase p53/p56lyn. In control cells Ca 2 was mobilized in response to gamma-radiation largely from internal stores, and increased significantly over a 20 min period. This mobilization of Ca2 was either absent or increased very slowly in A-T cells post-irradiation. The same pattern of release was observed after treatment with the radiometric agent, streptonigrin. In addition the phospatidylinositol 3-kinase (PI3-kinase) inhibitor wortmannin suppressed the release of Ca 2. Conclusion : These data demonstrate that ionizing radiation activates lyn kinase and leads to the release of Ca 2, and-for the first time-that these steps are ATM-dependent.