The Binding Landscape of an Essential Nuclear Hormone Receptor in C. elegans

Transcription factors (TFs) control gene expression by binding to response elements but distinguishing functional binding sites from non-functional ones remains a major challenge in the field of transcriptional regulation. To probe transcription factor binding and function in a multicellular organism we characterized the binding and transcriptional landscape of NHR-25, a highly-conserved and essential transcription factor in C. elegans. Using CRISPR/Cas9, we tagged the essential nuclear hormone receptor, nhr-25, with GFP and FLAG at its endogenous locus in C. elegans and performed ChIP-seq. We found that NHR-25 binds with two distinct modes: proximal promoter binding and distal enhancer-type binding. Motif analysis of the ChIP-seq peaks showed that the NHR-25 binding motif depended on chromatin context and identified additional enriched motifs, suggesting other putative regulatory partners at composite response elements. By combining ChIP-seq with conditional RNAi knockdown, we characterized nhr-25's transcriptome and identified putative direct targets of NHR-25 binding. Through an 11-bp mutation of the summit of an identified NHR-25 occupied region, we demonstrated that a single response element was responsible for regulating the expression of multiple genes. Our findings provide a comprehensive view of an essential transcription factor's functional binding landscape at native protein levels, highlighting the intricate nature of response element logic and demonstrating the critical role of a single regulatory element in regulating gene expression.