Heat Shock Factor 1 (HSF1) supports the ESR1 action in breast cancer (RNA-seq)

Heat shock factor 1 (HSF1) is a key regulator of transcriptional responses to proteotoxic stress. It has been recently linked to signaling of estrogen via ESR1. To study the cooperation of HSF1 and ESR1 in the transcriptional response to estrogen, we established estrogen receptor (ER)-positive breast cancer cell lines with reduced HSF1 levels using specific shRNA or CRISPR/Cas9 approach. HSF1 deficiency led to the inhibition of the mitogenic effect of estrogen in MCF7 and T47D cells. RNA-seq analyses revealed that the stimulatory effect of E2 on the transcriptome was smaller in HSF1-deficient MCF7 cells. This could partially result from the higher basal expression of E2-dependent genes in these cells as a consequence of the enhanced binding of unliganded ESR1 to chromatin, which was revealed by ChIP-seq analyses. Thus, we postulate that some fraction of ESR1 could be released from the inhibitory complex with HSP90 and gain transcriptional competence without E2-stimulation. Overall design: Total RNA was isolated from all MCF7 cell variants (untreated and treated with 10 nM E2 for 4 h or heat shocked for 1h at 43°C and recovered for 2 h) using the Direct-ZolTM RNA MiniPrep Kit (Zymo Research) and digested with DNase I (Worthington Biochemical Corporation). For each experimental point, RNAs from three biological replicates were first tested by RT-qPCR for the efficiency of treatments, then pooled and sequenced.