Fig. 3. Coimmunoprecipitation of HSF1 and HSF2. (A) Coimmunoprecipitation of HSF2 with transiently (left) or stably (right) overexpressed Myc-tagged HSF1 (4H9) using monoclonal anti-Myc antibodies (IP). The lower blot shows the expression levels of HSF1 and HSF2 (WB). (B) Exogenous HSF1 was coimmunoprecipitated with endogenous HSF2 using monoclonal HSF2 antibodies (IP). Anti-Flag antibodies were used as an antibody control. Cells were either left untreated (C) or heat shocked for 1 hour (HS). (C) The dynamics of the interacting complex was analyzed by coimmunoprecipitation of endogenous HSF2 with endogenous HSF1 using monoclonal HSF1 antibodies in HeLa cells heat-shocked for indicated times (IP). Thermotolerance was acquired at 1 or 3 hours of recovery after a 1-hour heat shock (HS+1R, HS+3R). The bottom blots show the expression levels of HSF1 and HSF2 (WB). Hsc70 was an equal loading control. HSF2- and -ß isoforms are indicated. Arrows and asterisks mark the inducibly phosphorylated HSF1 and the endogenous HSF2, respectively.

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