Nuclear receptors (NRs) are transcription factors that are implicated in several biological processes such as embryonic development, homeostasis, and metabolic diseases. To study the role of NRs in development, it is critically important to know when and where individual genes are expressed. Although systematic expression studies using reverse transcriptase PCR and/or DNA microarrays have been performed in classical model systems such as Drosophila and mouse, no systematic atlas describing NR involvement during embryonic development on a global scale has been assembled. Adopting a systems biology approach, we conducted a systematic analysis of the dynamic spatiotemporal expression of all NR genes as well as their main transcriptional coregulators during zebrafish development (101 genes) using whole-mount in situ hybridization. This extensive dataset establishes overlapping expression patterns among NRs and coregulators, indicating hierarchical transcriptional networks. This complete developmental profiling provides an unprecedented examination of expression of NRs during embryogenesis, uncovering their potential function during central nervous system and retina formation. Moreover, our study reveals that tissue specificity of hormone action is conferred more by the receptors than by their coregulators. Finally, further evolutionary analyses of this global resource led us to propose that neofunctionalization of duplicated genes occurs at the levels of both protein sequence and RNA expression patterns. Altogether, this expression database of NRs provides novel routes for leading investigation into the biological function of each individual NR as well as for the study of their combinatorial regulatory circuitry within the superfamily. NRs are key molecules controlling development, metabolism, and reproduction in metazoans. Since NRs are implicated in many human diseases such as cancer, metabolic syndrome, and hormone resistance, they are important pharmaceutical targets and are under intense scrutiny to better understand their biological functions. In the present study, we determined the expression patterns of all NR genes as well as their main transcriptional coregulators during zebrafish development. We used zebrafish because the transparency of its embryo allows us to perform whole-mount in situ hybridization from early development to late organogenesis. This complete developmental profiling offers an unprecedented view of NR expression during embryogenesis, uncovering their potential function during central nervous system and retina formation. We observed that in contrast to NR genes, only a few coregulators exhibit a restricted expression pattern, suggesting that tissue specificity of hormone action is conferred more by the receptors than by their coregulators. Lastly, by evolutionary analysis of expression pattern divergence of duplicated genes, we observed that neofunctionalization occurs at the levels of both protein sequence and mRNA expression patterns. Taken together, our data provide the starting point for functional analysis of an entire gene family during development and call for the study of the intersection between metabolism and development.

Retinoid X receptor (RXR) and Ultraspiracle (USP) play a central role as ubiquitous heterodimerization partners of many nuclear receptors. While it has long been accepted that a wide range of ligands can activate vertebrate/mollusc RXRs, the existence and necessity of specific endogenous ligands activating RXR‐USP in vivo is still matter of intense debate. Here we report the existence of a novel type of RXR‐USP with a ligand‐independent functional conformation. Our studies involved Tribolium USP (TcUSP) as representative of most arthropod RXR‐USPs, with high sequence homology to vertebrate/mollusc RXRs. The crystal structure of the ligand‐binding domain of TcUSP was solved in the context of the functional heterodimer with the ecdysone receptor (EcR). While EcR exhibits a canonical ligand‐bound conformation, USP adopts an original apo structure. Our functional data demonstrate that TcUSP is a constitutively silent partner of EcR, and that none of the RXR ligands can bind and activate TcUSP. These findings together with a phylogenetic analysis suggest that RXR‐USPs have undergone remarkable functional shifts during evolution and give insight into receptor–ligand binding evolution and dynamics.