Heat induces γH2AX foci formation in mammalian cells

Abstract

H2AX is a histone variant which is present and ubiquitously distributed throughout the genome. An immunocytochemical assay using antibodies capable of recognizing histone H2AX phosphorylated at serine 139 (γH2AX) is very sensitive and is a specific indicator for the existence of a DNA double strand break. Although heat stress has been reported to induce the formation of γH2AX foci, no γH2AX foci formation was observed in several mammalian cell lines after heat shock. Since this was in contrast to earlier reports, the work described here was intended to verify that heat-induced γH2AX foci do form in mammalian cell lines other than the cell lines used in earlier reports concerning γH2AX foci formation. The cell lines used in this work includes cell lines with differing p53-gene status (H1299, H1299/ neo, H1299/m p53 and H1299/wt p53 cells), various cancer cell lines (HeLa, HepG2, U2-OS cells), normal human cells (HEK-293 and AG1522), and cell lines established from other species (MEF normal mouse cells and CHL normal Chinese hamster cells). Exponentially growing cells were exposed to heat shock (42 °C for 6 h or 45.5 °C for 20 min) or to X-rays (3 Gy). The presence of γH2AX was examined with immunocytochemistry and flow cytometry. Induction of γH2AX foci formation was observed in all of the mammalian cell lines used here after heat-treatment as well as after X-irradiation. However, the intensity of γH2AX was different in the different cell lines used. These results confirm that heat can induce γH2AX foci formation in many mammalian cell lines.