Widespread regulatory specificities between transcriptional corepressors and enhancers in Drosophila

Animal development and homeostasis critically depend on the accurate regulation of gene transcription, which includes the silencing of genes that should not be expressed. Repression is mediated by a specific class of transcription factors (TFs) termed repressors that, via the recruitment of co-repressors (CoRs), can dominantly prevent transcription, even in the presence of activating cues. However, the relationship between specific CoRs and enhancers has remained unclear. Here, we used functional genomics to uncover regulatory specificities between CoRs and enhancers. We show that enhancers can typically be repressed by only a subset of CoRs. Enhancers classified by CoR sensitivity also show distinct biological functions and endogenous chromatin features. Moreover, enhancers that are sensitive or resistant to silencing by specific CoRs differ in TF motif content, and their sensitivity to CoRs can be predicted based on TF motif content. Finally, we identified and validated specific TF motifs that have a direct impact on enhancers sensitivity or resistance towards specific CoRs, using large scale motif mutagenesis and addition experiments. This study reveals the existence of TF motif-based regulatory rules that coordinate CoRs-enhancer compatibilities. These specificities between repressors and activators not only suggest that repression occurs via distinct mechanisms, but also provide an additional layer in transcriptional regulation that allows for differential repression at close genomic distances and offers multiple ways for de-repression.