Estrogen-receptor-α exchange and chromatin dynamics are ligand- and domain-dependent

摘要: We report a mammalian-based promoter chromosomal array system developed for single-cell studies of transcription-factor function. Designed after the prolactin promoter-enhancer, it allows direct visualization estrogen receptor α (ERα) and/or Pit-1 interactions at physiologically regulated transcription locus. ERα- and ligand-dependent cofactor recruitment, large-scale chromatin modifications transcriptional activity identified distinct fingerprint responses each condition. Ligand-dependent (more than threefold activation compared with vehicle, or complete repression by mRNA fluorescent in situ hybridization) correlated its state condensation, which was assayed using novel high throughput microscopy approach. In support nuclear hit-and-run model, photobleaching provided evidence very transient ER-array interactions, revealed changes k off. ERα-truncation mutants indicated that helix-12 co-regulators influenced both modeling recovery times. These data also showed ERα DNA-binding domain insufficient targeting. Collectively, quantitative observations from this relevant biosensor suggest stochastic-based dynamics influence gene regulation level.