RNA fine-tunes estrogen receptor-alpha binding on low-affinity DNA motifs for transcriptional regulation

Transcription factors (TFs) regulate gene expression by binding, with varying strength, to DNA via their DNA-binding domain. Additionally, some TFs also interact with RNA, which modulates transcription factor binding to chromatin. However, whether RNA-mediated TF binding results in differential transcriptional outcomes remains unknown. In this study, we demonstrate that estrogen receptor a (ERa), a ligand-activated TF, interacts with RNA in a ligand-dependent manner. Defects in RNA binding lead to genome-wide loss of ERa recruitment, particularly at weaker ERa-motifs. Furthermore, ERa mobility in the nucleus increases in the absence of its RNA-binding capacity. Unexpectedly, this increased mobility coincides with robust polymerase loading and transcription of ERa-regulated genes that harbor low-strength motifs. However, highly stable binding of ERa on chromatin negatively impacts ligand-dependent transcription. Collectively, our results suggest that RNA interactions spatially confine ERa on low-affinity sites to fine-tune gene transcription. Overall design: Chromatin Immunoprecipitation (ChIP-seq) for the FLAG tagged WT and RBM Estrogen receptor-alpha upon 72 hours of hormone deprivation and 1 hour of E2 signaling. RNAseA ChIP-seq of Estrogen receptor- alpha with and without pre-extraction after 72 hours of hormone deprivation and 1 hour of E2 siganling . ChIP-seq of total PolII upon 1 hour of E2 siganling with expression of either FLAG tagged WT or RBM Estrogen receptor alpha. Paired factor ChIP with antiFLAG and anti-CTCF upon expression of either FLAG tagged WT or RBM Estrogen receptor alpha.