Figure 7.

Heat-shock factor 1 occupies a heat shock element on the fibronectin promoter and occupancy increases with geldanamycin treatment. (A) Schematic diagram showing the location of putative HSEs in the FN1 promoter and the location of primer sets used to amplify these regions from ChIP DNA. (B) Hs578T cells were treated with either 100 nM GA or DMSO. Chromatin immunoprecipitation with anti-HSF1 antibody or isotype control was conducted using the Imprint UltraChromatin Immunoprecipitation Kit (Sigma-Aldrich) as per the manufacturer’s instructions. The purified DNA was amplified by qPCR using primers F1 and R1 (designed to amplify the region containing HSE1) and the fold enrichment of the putative FN1 heat-shock element relative to the isotype control and input DNA from the anti-HSF1 immunoprecipitation under the two conditions was determined using the ∆Ct method (2^{−ΔΔCt [HSF1/Isotype]})⁴³. Data shown are average fold enrichments from 3 samples, and are representative of 2 independent biological experiments conducted in triplicate. Statistical analysis was conducted using one way ANOVA with Bonferroni post-test (*p < 0.05) comparing the HSF1 ChIP samples to the relevant isotype ChIP control.