Fig 3.

PIN1 is required for HSF1-dependent HSP gene expression. (A) Schematic of the HSP70B promoter-driven luciferase construct. Wild type MEFs, PIN1^−/− MEFs, and PIN1-expressing-PIN1^−/− MEFs were exposed to heat shock and analyzed by luciferase assay. (B) Schematic of the luciferase construct containing three repeats of the HSE (3HSE). In the left panel, HeLa cells cotransfected with PIN1-HA and 3HSE constructs were exposed to heat shock and analyzed by luciferase assay. In the right panel, HeLa cells cotransfected with the PIN1 shRNA plasmid and 3HSE constructs were subjected to heat shock and analyzed by luciferase assay. Wild-type and PIN1^−/− MEFs were heat shock treated and allowed to recover for 3 h. Cytoplasmic RNA was extracted and analyzed by quantitative RT-PCR. Comparative HSP gene expression profiles for HSP70 (C), HSP90 (D), and HSP105 (E) are shown. (F) Wild-type and PIN1^−/− MEFs were subjected to heat shock treatment and allowed to recover for 10 h. Cells were lysed with RIPA buffer and analyzed by Western blotting. (G) Heat-induced HSP expression in MCF7 cells treated with scramble-shRNA or shPIN1 (PIN1 shRNA). Cells were exposed to heat shock treatment for 1 h at 43°C as indicated (HS) and analyzed by Western blotting after 10 h of recovery at 37°C. An actin Western blot serves as the loading control.