A CRISPR-Cas9 screen identifies essential CTCF anchor sites for Estrogen Receptor-driven breast cancer cell proliferation

Estrogen receptor a (ERa) is an enhancer activating transcription factor, a key driver of breast cancer, and a main target for cancer therapy. ER?-mediated gene regulation requires proper chromatin-conformation to facilitate interactions between ER?-bound enhancers and their target promoters. A major determinant of chromatin structure is the CCCTC-binding factor (CTCF), that dimerizes and together with cohesin stabilizes chromatin loops and forms the boundaries of topologically associated domains. However, whether CTCF-binding elements (CBEs) are essential for ER?-driven cell proliferation is unknown. To address this question in a global manner, we implemented a CRISPR-based functional genetic screening approach targeting CBEs located in the vicinity of ERa-bound enhancers. We identified four functional CBEs and demonstrated the role of one in inducing chromatin conformation changes in favor of activation of PREX1, a key ERa target gene in breast cancer. Indeed, high PREX1 expression is a bona-fide marker of ER-dependency in cell lines, and associated with good outcome after anti-hormonal treatment. Altogether, our data show that distinct CTCF-mediated chromatin structures are required for ER?- driven breast cancer cell proliferation. Overall design: The analysis of CTCF, ERalpha, and FOXA1 in MCF-7 cells was done by ChIP-seq data infected either with indicated sgRNA or non-targeting as a control.