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

Abstract

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-25s 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 factors 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.