Molecular Evidence for a Functional Ecdysone Signaling System in Brugia malayi

Abstract

BackgroundFilarial nematodes, including Brugia malayi, the causative agent of lymphatic filariasis, undergo molting in both arthropod and mammalian hosts to complete their life cycles. An understanding of how these parasites cross developmental checkpoints may reveal potential targets for intervention. Pharmacological evidence suggests that ecdysteroids play a role in parasitic nematode molting and fertility although their specific function remains unknown. In insects, ecdysone triggers molting through the activation of the ecdysone receptor: a heterodimer of EcR (ecdysone receptor) and USP (Ultraspiracle).Methods and FindingsWe report the cloning and characterization of a B. malayi EcR homologue (Bma-EcR). Bma-EcR dimerizes with insect and nematode USP/RXRs and binds to DNA encoding a canonical ecdysone response element (EcRE). In support of the existence of an active ecdysone receptor in Brugia we also cloned a Brugia rxr (retinoid X receptor) homolog (Bma-RXR) and demonstrate that Bma-EcR and Bma-RXR interact to form an active heterodimer using a mammalian two-hybrid activation assay. The Bma-EcR ligand-binding domain (LBD) exhibits ligand-dependent transactivation via a GAL4 fusion protein combined with a chimeric RXR in mammalian cells treated with Ponasterone-A or a synthetic ecdysone agonist. Furthermore, we demonstrate specific up-regulation of reporter gene activity in transgenic B. malayi embryos transfected with a luciferase construct controlled by an EcRE engineered in a B. malayi promoter, in the presence of 20-hydroxy-ecdysone.ConclusionsOur study identifies and characterizes the two components (Bma-EcR and Bma-RXR) necessary for constituting a functional ecdysteroid receptor in B. malayi. Importantly, the ligand binding domain of BmaEcR is shown to be capable of responding to ecdysteroid ligands, and conversely, ecdysteroids can activate transcription of genes downstream of an EcRE in live B. malayi embryos. These results together confirm that an ecdysone signaling system operates in B. malayi and strongly suggest that Bma-EcR plays a central role in it. Furthermore, our study proposes that existing compounds targeting the insect ecdysone signaling pathway should be considered as potential pharmacological agents against filarial parasites.

The genome of the hydatid tapeworm Echinococcus granulosus

“…We also found genes encoding five sodium-bile acid cotransporters and seven multidrug resistance proteins (MRPs) 23 , as well as genes associated with bile acid metabolism, including sterol regulatory element-binding protein 1 and bile acid β-glucosidase-related proteins (Supplementary Table 30). Ecdysone or other sex steroids might regulate molting in parasitic nematodes through nuclear hormone receptors 24,25 , and it is reasonable to assume that exogenous bile acid from the host has an important role in the development of E. granulosus in a similar fashion.…”

Sex Steroids in Insects and the Role of the Endosymbiont Wolbachia:A New Perspective

“…As previously discussed, arthropod moulting is induced by the steroid hormone 20E and a role for ecdysteroids in nematode ecdysis has also been observed. In filarial nematodes, moulting seems to be regulated by ecdysteroid-like hormones: in Dirofilaria immitis, for example, moulting from the third to the fourth larval stage can be induced in vitro by the 20E of insects (Wabrick et al, 1993), and orthologs of insects nuclear receptors involved in ecdysone response have been found (Crossgrove et al, 2008;Ghedin et al, 2007;Tzertzinis et al, 2010). In Caenorhabditis elegans these nuclear receptors are also involved in the regulation of sex determination and reproductive development (Höss & Weltje, 2007;Motola et al, 2006) and, interestingly, ecdysone has also a role in the fertility and microfilaria release in filarial worms (Barker et al, 1991).…”

Nuclear receptors in nematode development: Natural experiments made by a phylum

“…RXR homologues are also found in Onchocerca volvulus Ov-RXR [90] and in Dirofilaria immitis Di-RXR-1. Di-RXR-1 was shown to be able to functionally interact with Drosophila EcR [35].…”

“…An RXR homologue is found in the genome of Pristonchus pacificus [36]. The genomes of the parasitic nematode Dirofilaria immitis [35] and of Brugia malayi both encode RXR and EcR [21,90,91] supporting the presence of RXR in basal nematode species.…”

“…Brugia malayi RXR, Bma-RXR is expressed in L1 larvae and in adults. Bma-RXR is able to dimerize with Bma-EcR and the LBD of Bma-EcR was capable of transducing an ecdysteroid signal in a mammalian cell system [35,90].…”