Evidences for Red Pigment Concentrating Hormone (RPCH) and Beta-Pigment Dispersing Hormone (ß-PDH) Inducing Oocyte Meiotic Maturation in the Chinese Mitten Crab, Eriocheir sinensis.

Red pigment concentrating hormone (RPCH) and pigment dispersing hormone (PDH) are crustacean neuropeptides involved in broad physiological processes including body color changes, circadian rhythm, and ovarian growth. In this study, the full-length cDNA of RPCH and PDH were identified from the brain of the Chinese mitten crab Eriocheir sinensis. The deduced RPCH and PDH mature peptides shared identical sequence to the adipokinetic hormone/RPCH peptides family and the ß-PDH isoforms and were designated as Es-RPCH and Es-ß-PDH, respectively. Es-RPCH and Es-ß-PDH transcripts were distributed in the brain and eyestalks. The positive signals of Es-RPCH and Es-ß-PDH were localized in the neuronal clusters 6, 8, 9, 10, and 17 of the brain as revealed by in situ hybridization. The expression level of Es-RPCH and Es-ß-PDH mRNA in nervous tissues were all significantly increased at vitellogenic stage, and then decreased at the final meiotic maturation stage. The administrated with synthesized Es-RPCH peptide results in germinal vesicles shift toward the plasma membrane in vitellogenic oocyte, and significant decrease of the gonad-somatic index (GSI) and mean oocyte diameter as well as the expression of vitellogenin mRNA at 30 days post injection in vivo. Similar results were also found when injection of the Es-ß-PDH peptide. In vitro culture demonstrated that Es-RPCH and Es-ß-PDH induced germinal vesicle breakdown of the late vitellogenic oocytes. Comparative ovarian transcriptome analysis indicated that some reproduction/meiosis-related genes such as cdc2 kinase, cyclin B, 5-HT-R and retinoid-X receptor were significantly upregulated in response to Es-RPCH and Es-ß-PDH treatments. Taken together, these results provided the evidence for the inductive effect of Es-RPCH and Es-ß-PDH on the oocyte meiotic maturation in E. sinensis.

Transcriptional regulation of ferritin mRNA levels by iron in the freshwater giant prawn, Macrobrachium rosenbergii.

Ferritin, a major iron storage protein of most living organisms play a crucial role in iron metabolism. A cDNA encoding a heavy-chain homologue of ferritin was isolated and sequenced in the freshwater giant prawn, Macrobrachium rosenbergii. The cloned cDNA was 1012 bp long containing an iron-responsive element (IRE) sequence in the 5'-untranslated region and a complete open-reading frame encoding for a 172 amino acid residues protein with a conserved domain for the ferroxidase center characteristic for heavy chains of vertebrate ferritin. Prawn ferritin transcripts are expressed in muscle, heart, hepatopancreas, stomach, hemocytes, ovary and testis. Quantitative real-time PCR revealed that the abundance of ferritin transcript was highest in the hepatopancreas, followed by the testis. The expression of the ferritin transcript in the muscle significantly increased 6-fold at 3 h after injection of iron. In the ovary, a high expression of ferritin transcript was first detected with nearly a 4-fold increase after 3 h post-injection that remained elevated for 48 h. Heart ferritin mRNA expression increased up to 8-fold at 24 h. No significant difference was found in the hepatopancreas, hemocytes and testis. These results strongly suggested that the expression of prawn ferritin is regulated by iron at the transcriptional level as found in insects, and iron increased prawn ferritin transcripts in a tissue-specific manner.