Ecdysone receptor in the mud crab Scylla paramamosain: a possible role in promoting ovarian development

Jan 8, 2015The Journal of endocrinology

Ecdysone receptor in mud crabs and its possible role in supporting ovary development

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Abstract

Three cDNA isoforms of the ecdysone receptor (EcR) were cloned from the mud crab Scylla paramamosain.

SpEcR mRNA was found to be abundant in the eyestalk, ovary, and epidermis.
Levels of SpEcR transcripts increased during ovarian development, peaking at stage IV before declining at stage V.
The concentration of 20-hydroxyecdysone (20E) in the hemolymph mirrored the pattern of SpEcR transcript levels during ovarian development.
In vivo experiments showed that 20E significantly stimulated the expression of SpEcR and vitellogenin (SpVg) during the early vitellogenic stage.
In vitro experiments demonstrated that SpEcR and SpVg expression increased in response to 20E in early vitellogenic ovarian tissue but not in previtellogenic tissue.
Gene silencing of SpEcR resulted in a significant inhibition of SpEcR and SpVg expression in the ovary.

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In arthropods, it is known that ecdysteroids regulate molting, limb regeneration, and reproduction through activation of the ecdysone receptor (EcR). However, the ecdysteroid signaling pathway for promotion of ovarian development in crustaceans is still unclear. In this study, three cDNA isoforms of EcR were cloned from the mud crab Scylla paramamosain. qRT-PCR revealed that the SpEcR mRNA was abundant in the eyestalk, ovary and epidermis. During ovarian development, the SpEcR transcripts increased from stage I (undeveloped stage) and reached a peak at stage IV (late vitellogenic stage) before dropping to a lower level at stage V (mature stage). Meanwhile, levels of 20-hydroxyecdysone (20E) in the hemolymph, detected by HPLC-MS, displayed a similar pattern of increase with ovarian development. Results from in situ hybridization indicated that SpEcR mRNA was present in the follicular cells during vitellogenesis. Results from in vivo experiments revealed that 20E at 0.2 μg/g body weight significantly stimulated the expression of SpEcR and vitellogenin (SpVg) in female crabs during the early vitellogenic stage but not during the previtellogenic stage. This was confirmed by results from in vitro experiments which indicated that SpEcR and SpVg expression levels were significantly upregulated in early vitellogenic ovarian explants incubated with 5.0 μM 20E at 3 and 6 h but not in previtellogenic ovarian explants. Finally, results from in vitro gene silencing experiments indicated that the expression of SpEcR and SpVg in the ovary was significantly inhibited by SpEcR dsRNA. All these results together indicated that in S. paramamosain, 20E, and SpEcR, located in the follicular cells, play important roles in the promotion of ovarian development via regulating the expression of SpVg.

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Ecdysone receptor in the mud crab Scylla paramamosain: a possible role in promoting ovarian development

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