Fig. 1. Heat-induced localization of HSF1 and HSF2 in nuclear granules. Fluorescence micrographs of HSF1 and HSF2 localization in untreated (C) and 1-hour heat-shocked (HS) HeLa cells, using polyclonal antibodies against HSF1 (left) and HSF2 (right). The phase contrast and DAPI-stained DNA of the same fields are also shown. (B) Cells were untreated (C), heat-shocked for 30 minutes to 6 hours (30'-6h) or, after 1 hour of heat shock, allowed to recover for 1 or 3 hours (HS+1R, HS+3R). (C) The graph represents the means ± SEM of three independent experiments in which the number of granule positive cells was analyzed by counting nine groups of 50 cells. (D) The HSF DNA-binding activity and HSF1 hyperphosphorylation were analyzed by gel mobility shift assay with radiolabeled HSE (top) and by western blotting (bottom), respectively. The arrow indicates the inducibly phosphorylated HSF1. Scale bars, 5 µm.