Expression and ecdysteroid responsiveness of the nuclear receptors HR3 and E75 in the crustacean Daphnia magna

Abstract

Ecdysteroids initiate signaling along multiple pathways that regulate various aspects of development, maturation, and reproduction in arthropods. Signaling often involves the induction of downstream transcription factors that either positively or negatively regulate aspects of the pathway. We tested the hypothesis that crustaceans express the nuclear receptors HR3 (ortholog to vertebrate ROR) and E75 (ortholog to vertebrate rev-erb) in response to ecdysteroid signaling. HR3 and E75 cDNAs were cloned from the crustacean Daphnia magna. The DNA binding domain and ligand binding domain of the daphnid HR3 was 95% and 61% identical to those of Drosophila melanogaster. The DNA binding domain and ligand binding domain of the daphnid E75 was 100% and 71% identical to those of D. melanogaster. Both receptors exhibited structural characteristics of binding to DNA as a monomer. The expression of these receptor mRNAs was evaluated through the adult molt cycle and during embryo development. E75 levels were relatively constant throughout the adult molt cycle and through embryo development. HR3 levels were comparable to those of E75 during the initial phases of the adult molt cycle but were elevated ~30-fold at a time in the cycle co-incident with the pre-molt surge in ecdysteroid levels. HR3 mRNA levels in embryos also varied coincident with ecdysteroids levels. To substantiate a role of ecdysteroids in the expression of HR3, daphnids were continuously exposed to 20-hydroxyecdysone and changes in gene expression were measured. HR3 levels were significantly induced by 20-hydroxyecdysone; while E75 levels were minimally affected. These results are consistent with the premise that transcription of HR3 is regulated by ecdysteroids in the crustacean Daphnia magna and that HR3 likely serves as a mediator of ecdysteroid regulatory action in crustaceans. The marginal induction of E75 by 20-hydroxyecdysone may represent limited, tissue or cell-type-specific induction of this transcription factor.

Alternative splicing in the fiddler crab cognate ecdysteroid receptor: Variation in receptor isoform expression and DNA binding properties in response to hormone

“…4 (continued) through September), and the Indo-Pacific species, U. annulipes and U. triangularis, breed continuously for 9 months of the year (i.e., August through the succeeding May), releasing as many as 16-18 broods (Crane, 1975). Although both ecdysteroids and methyl farnesoate have been implicated in the control of oogenesis in crustaceans, identification of responsive genes is just beginning (Shechter et al, 2007;Soetaert et al, 2007;Hannas and LeBlanc, 2010;Nagaraju et al, 2011;Tiu et al, 2012). Whether the differences in expression pattern for the two components of the ecdysteroid receptor in the ovaries of U. annulipes and U. triangularis is related to the ability of these species to be breed continuously awaits studies of the effects of receptor gene knockdown on oogenesis, and what specific genes and gene networks are under hormonal control.…”

Agonist-mediated assembly of the crustacean methyl farnesoate receptor

“…Three hundred adult female D. magna were reared as described by Hannas et al 37. for use in MfR subunit gene expression analysis.…”

Review of the reproductive biology of amphipods and their endocrine regulation: Identification of mechanistic pathways for reproductive toxicants

“…Displacement of zinc in the DNA binding protein domain by Pb and Cd has been proposed as a mechanism causing toxicity of these metals [120,121], but whether these metals cause reproductive toxicity in amphipods by this mechanism has not been determined. Other nuclear receptor genes have been identified in the Daphnia pulex genome, but little is known of the function of the nuclear receptors of crustaceans [122]. Some of these nuclear receptors possess ligand-binding sites and may also serve as targets for disruption by environmental contaminants.…”