The Role of the Mre11 Complex in Heat-radiosensitization

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DESCRIPTION (provided by applicant): Several phase III clinical trials have validated the use of hyperthermia as an adjuvant to radiation therapy. Hyperthermia sensitizes mammalian cells to ionizing radiation. Therefore, the potential benefit of using heat in the clinic may be maximized if the molecular mechanisms for heat-radiosensitization are better understood. Radiation induces the formation of DNA double strand breaks (DSBs), and thermal radiosensitization is believed to be caused by an inhibition of repair of radiation-induced DSBs by heat. Normally, radiationinduced DSB repair is believed to occur via two pathways: non-homologous end-joining (NHEJ) and homologous recombination (HR). The Mre11 complex has been implicated to function in both of these pathways. Our preliminary data suggest that hyperthermia induces alterations to Mrell, RadSO and Nbsl, the proteins comprising the Mre11 complex. We have also shown that heat induces the translocation of these proteins from the nucleus to the cytoplasm. Heat-induced alterations to, or redistribution of the components of the Mre11 complex could result in a decrease in DSB repair efficiency and potentiation of radiationinduced cell killing. We will test the-hypothesis that heat-radiosensitization is caused by heat-induced alterations to, or redistribution of proteins of the Mre11 DSB repair complex. We will determine the role of Mre11 and Rad50 in heat-radiosensitization, and determine whether alterations of these proteins lead to an inhibition or decrease in the efficiency of DSB repair and heat-radiosensitization after cells are exposed to moderate temperature (41.5¿C) hyperthermia treatments. Using Western blotting and confocal microscopy, we will characterize heat-induced thermal damage to, or relocalization of Mre11, RadSO, and Nbsl and attempt to identify the mechanisms by which these alterations occur. We will also determine the role of HR in heat-radiosensitization using silencing RNA (siRNA) targeting technology or conditional mutants deficient in specific NHEJ or HR proteins.