The HR97 (NR1L) group of nuclear receptors: A new group of nuclear receptors discovered in Daphnia species

Abstract

The recently sequenced Daphnia pulex genome revealed the NR1L nuclear receptor group consisting of three novel receptors. Phylogenetic studies show that this group is related to the NR1I group (CAR/PXR/VDR) and the NR1J group (HR96), and were subsequently named HR97a/b/g. Each of the HR97 paralogs from Daphnia magna, a commonly used crustacean in toxicity testing, was cloned, sequenced, and partially characterized. Phylogenetic analysis indicates that the HR97 receptors are present in primitive arthropods such as the chelicerates but lost in insects. qPCR and immunohistochemistry demonstrate that each of the receptors is expressed near or at reproductive maturity, and that HR97g, the most ancient of the HR97 receptors, is primarily expressed in the gastrointestinal tract, mandibular region, and ovaries, consistent with a role in reproduction. Transactivation assays using an HR97a/b/g-GAL4 chimera indicate that unlike Daphnia HR96 that is promiscuous with respect to ligand recognition, the HR97 receptors do not respond to many of the ligands that activate CAR/PXR/HR96 nuclear receptors. Only three putative ligands of HR97 receptors were identified in this study: pyriproxyfen, methyl farnesoate, and arachidonic acid. Only arachidonic acid, which acts as an inverse agonist, alters HR97g activity at concentrations that would be considered within physiologically relevant ranges. Overall, this study demonstrates that, although closely related to the promiscuous receptors in the NR1I and NR1J groups, the HR97 receptors are mostly likely not multi-xenobiotic sensors, but rather may perform physiological functions, potentially in reproduction, unique to crustaceans and other non-insect arthropod groups.

Annotation of the Nuclear Receptors in an Estuarine Fish species, Fundulus heteroclitus

“…Because of whole genome duplication in teleosts (42), it is most likely that an ancient relative lost PPARβ/δ in the Percomorphs and PPARα in the early teleosts. However, it cannot be completely ruled out that separate duplication events occurred in the Percomorphs and early teleosts as individual gene duplication events have occurred multiple times including cytochrome P450s, opsins, and NRs (20,35,61–63). The organ distribution of PPARs differs from mammals.…”

“…These domains were identified and isolated using the pfam00105 (Zf-C4) and pfam00104 (hormone receptor) designations on the conserved domain database CDD (34) as described previously (20,23,35). The DBD and LBD from 246 NRs from three fish species and humans were aligned using ClustalX default parameters (36) (Additional File 2).…”

Annotation of the Daphnia magna nuclear receptors: Comparison to Daphnia pulex

“…NRs in this group include those activated by ecdysteroids and responsible for molting and development (EcR) (Fahrbach et al, 2012), the newly discovered HR97 group, RAR- and THR-like receptors (RARL_10, THRL_11), and HR96, a receptor involved in cholesterol and triacylglycerol homeostasis that is also promiscuous and involved in xenobiotic stress responses (King-Jones et al, 2006; Horner et al, 2009; Karimullina et al, 2012; Sieber and Thummel, 2012; Li et al, 2014). Many NR1 subfamily members are involved in resource allocation or energy metabolism, including LXR/FXR (NR1H) (Schultz et al, 2000; Zhang et al, 2012), CAR/PXR/VDR/HR96 (NR1I/J) (Dong et al, 2009; Gao and Xie, 2010; Karimullina et al, 2012; Sieber and Thummel, 2012), PPARs (NR1C) (Semple et al, 2006; Jiang et al, 2012; LeBlanc et al, 2012), and THR (NR1A) (Duntas and Brenta, 2012; Cordeiro et al, 2013).…”

“…There have been several duplication events within the large Daphnia NR1 clade as the 1H group (EcRα/β) has two paralogs and the new 1L group (HR97a/b/g), first discovered in D. pulex, has three paralogs (Thomson et al, 2009; Li et al, 2014). HR97a and b show high conservation between D. pulex and D. magna , but HR97g, the most ancient of the three receptors is weakly to moderately conserved relative to most NRs ( Table 1 ).…”

“…HR97a and b show high conservation between D. pulex and D. magna , but HR97g, the most ancient of the three receptors is weakly to moderately conserved relative to most NRs ( Table 1 ). Interestingly, new data indicates that the HR97 receptors are not as promiscuous as the HR96 receptors (Li et al, 2014), and therefore may represent an example of the evolution of specificity from promiscuity (Eick et al, 2012). …”