The Hippo kinase LATS1 controls CTCF chromatin occupancy and the hormonal response of three-dimensionally grown breast cancer cells

The cancer epigenome has been studied in cells cultured in 2D monolayers on plastic surfaces, but recent studies highlight the impact of the extracellular matrix (ECM) and the 3D environment on multiple cellular functions. Here, we report the physical, biochemical and genomic differences between T47D breast cancer cells cultured in 2D monolayer and as 3D spheroids in Matrigel. Cells within 3D spheroids exhibit a rounder nucleus with less accessible, more compacted chromatin, and altered expression of over 2,000 genes, the majority of which become repressed. Hi-C analysis reveals that cells grown in 3D exhibit enrichment in regions belonging to the B compartment, decrease on chromatin bound CTCF and increased fusion of Topologically Associating Domains (TADs). Upregulation of the Hippo pathway in 3D spheroids results in the activation of the LATS1 kinase, which promotes phosphorylation and displacement of CTCF from DNA, likely responsible for the observed TAD fusions. Cells grown in 3D exhibit higher progesterone receptor (PR) binding to chromatin, leading to an increase in the number hormone-regulated genes. This effect is in part mediated by the LATS1 activation, which favors cytoplasmic retention of YAP and CTCF removal.