Phospho-Nbs1 and Mre11 proteins which recognize DSBs co-localize with γH2AX in the nucleus after heat treatment

Abstract

Background: Critical hyperthermia targets in cancer cells have to be clearly identified.
Method: To determine if the DNA double strand break (DSB)-recognizing proteins, phospho-Nbs1 and Mre11, co-localize with phospho-H2AX (γH2AX), immunohistochemical methods were used with multiple antibodies. After a heat treatment at 45.5°C for 20 min, human normal fibroblasts were analyzed with a laser scanning confocal microscope at 0.5 h and 8 h post-heat treatment.
Results: At 0.5 h after a heat treatment, γH2AX formed foci in the nucleus, but phospho-Nbs1 and Mre11 were scattered over the nucleus and cytoplasm, and did not co-localize with γH2AX. At 8 h after a heat-treatment, and also after X-irradiation, both phospho-Nbs1 and Mre11 co-localized with γH2AX foci, which were already present, in the nucleus. Moreover, both phospho-Nbs1 and Mre11 which were observed at 0.5 h after a heat treatment in the cytoplasm, were no longer observed in the cytoplasm at 8 h after a heat treatment.
Conclusion: These findings provide support for the concept that heat, like X-rays, may lead to the induction of DSBs.