Comprehensive Genomic Analysis Reveals that the Pioneering Function of FOXA1 Is Independent of Hormonal Signaling.

Considerable work has linked hormone receptors, such as estrogen receptor-alpha (ER), with the pioneer factor FOXA1. Altered FOXA1 levels contribute to endocrine-resistant breast cancer, where it maintains ER-chromatin interactions, even in contexts in which cells are refractory to ER-targeted drugs. A recent study controversially suggests that FOXA1 binding can be induced by hormonal pathways, including the estrogen-ER complex. We now show that the vast majority (>99%) of FOXA1 binding events are unaffected by steroid activation. A small number (<1%) of FOXA1 binding sites appear to be induced by estrogen, but these are created from chromatin interactions between ER binding sites and adjacent FOXA1 binding sites and do not represent genuine new FOXA1-pioneering elements. FOXA1 is therefore not regulated by estrogen and remains a bone fide pioneer factor that is entirely upstream of the ER complex.

Identification of ChIP-seq and RIME grade antibodies for Estrogen Receptor alpha.

Estrogen Receptor alpha (ERα) plays a major role in most breast cancers, and it is the target of endocrine therapies used in the clinic as standard of care for women with breast cancer expressing this receptor. The two methods ChIP-seq (chromatin immunoprecipitation coupled with deep sequencing) and RIME (Rapid Immunoprecipitation of Endogenous Proteins) have greatly improved our understanding of ERα function during breast cancer progression and in response to anti-estrogens. A critical component of both ChIP-seq and RIME protocols is the antibody that is used against the bait protein. To date, most of the ChIP-seq and RIME experiments for the study of ERα have been performed using the sc-543 antibody from Santa Cruz Biotechnology. However, this antibody has been discontinued, thereby severely impacting the study of ERα in normal physiology as well as diseases such as breast cancer and ovarian cancer. Here, we compare the sc-543 antibody with other commercially available antibodies, and we show that 06-935 (EMD Millipore) and ab3575 (Abcam) antibodies can successfully replace the sc-543 antibody for ChIP-seq and RIME experiments.