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To elucidate the molecular mechanisms underling hydration during oocyte maturation, we characterized the structure of Japanese eel (Anguilla japonica) novel-water selective aquaporin 1 (AQP1b) that thought to be involved in oocyte hydration. The aqp1b cDNA encodes a 263 amino acid protein that includes the six potential transmembrane domains and two Asn-Pro-Ala motifs. Reverse transcription-polymerase chain reaction showed transcription of Japanese eel aqp1b in ovary and testis but not in the other tissues. In situ hybridization studies with the eel aqp1b cRNA probe revealed intense eel aqp1b signal in the oocytes at the perinucleolus stage and the signals became faint during the process of oocyte development. Light microscopic immunocytochemical analysis of ovary revealed that the Japanese eel AQP1b was expressed in the cytoplasm around the yolk globules which were located in the peripheral region of oocytes during the primary yolk globule stage; thereafter, the immunoreactivity was observed throughout the cytoplasm of oocyte as vitellogenesis progressed. The immunoreactivity became localized around the large membrane-limited yolk masses which were formed by the fusion of yolk globules during the oocyte maturation phase. These results together indicate that AQP1b, which is synthesized in the oocyte during the process of oocyte growth, is essential for mediating water uptake into eel oocytes. [ABSTRACT FROM AUTHOR]

Three female specific serum proteins were detected immunologically in the sera of grey mullet ( Mugil cephalus) which were named vitellogenin A (VgA), VgB, and VgC, based upon their distinct antigenicity against specific antisera raised against three types of mullet lipovitellins (Lvs). These Vgs were subsequently purified from the serum of estradiol-treated mullet by combining several types of chromatography columns (anion exchanger, hydroxylapatite, immunoadsorbent column, and gel filtration). Purified native VgA, VgB, and VgC exhibited molecular masses of 570, 580, and 335 kDa, respectively. Following, SDS-PAGE, the estimated mass of polypeptide bands evident for VgA and VgB were ∼179 and ∼175 kDa, respectively; VgC appeared to be ∼132 kDa. The two larger Vgs (VgA and VgB) appeared to be phosphorylated, suggesting that these Vgs contain a highly phosphorylated, serine-rich phosvitin (Pv) domain. Furthermore, two discrete Vg-type specific antisera, anti-VgA and anti-VgB, were developed and each generated two precipitin lines against ovary extracts in immunoelectrophoresis, indicating that these Vgs contain additional antigenic yolk protein domains: Lv and β′-component. The small Vg (VgC) appeared to lack a Pv domain because of its low serine content (5.35%) and failure to show positive results in phospho-staining experiments. In conjunction with N-terminal amino acid sequencing analyses of the purified Vgs, our present results have conclusively identified the purified Vg products in grey mullet as typical A-type (VgA), B-type (VgB), and C-type (VgC) Vgs. [ABSTRACT FROM AUTHOR]

To elucidate the utilization of the major yolk nutrient stocks in eggs and larvae of walleye pollock Theragra chalcogramma, the contents of free amino acids (FAA), the major yolk protein (180 kDa lipovitellin originated from vitellogenin B in ovulated eggs: oLv B), and lipids were measured. Most eggs hatched 18 days after fertilization at 5°C, and all larvae absorbed almost all their yolk mass by 28 days. The total FAA content showed no change during the first 6 days, and then decreased to 28% of the initial level by 18 days. The oLv B contents, measured by an enzyme-linked immunosorbent assay using a specific antiserum against oLv B, gradually decreased from 6 to 18 days, followed by a rapid decline. The content of phospholipids (PL) and triacylglycerols (TG) showed no marked change until hatching, and then decreased until disappearance of yolk sac. From these results, it is proposed that there are two main periods for nutrient utilization in embryos and larvae of walleye pollock. In the first period, FAA was mainly utilized until 18 days after fertilization. Active utilization of oLv B and lipids (PL and TG) instead of FAA occurred during the second period from 18 to 28 days. [ABSTRACT FROM AUTHOR]

We investigated the effects of endocrine-disrupting chemicals (EDCs) on the expression of ubiquitin C-terminal hydrolase (UCH) mRNA in the testis and brain of the Japanese common goby, Acanthogobius flavimanus. The cDNA sequence of goby UCH contained an open reading frame encoding 220 amino acid residues (Mr=24,223) with 51.3% overall sequence identity with human and mouse UCHL1. A competitive PCR assay was used to quantify the levels of UCH mRNA in the testis and brain of male gobies after exposure to bisphenol A, nonylphenol, or estradiol-17β for 3 weeks. Exposure to estradiol-17β at a nominal concentration of 100ng/L induced significant increase in UCH mRNA levels in both testis and brain (P<0.05), whereas exposure to nonylphenol induced a significant decrease in UCH mRNA levels in the testis (P<0.01). These results suggest that EDCs can either positively or negatively regulate UCH mRNA levels. [Copyright &y& Elsevier]

Abstract: To elucidate the utilization of major yolk nutrients in eggs and yolk-sac larvae of Japanese eel Anguilla japonica, contents of free amino acids (FAA), 380 kDa lipovitellin (oLv; the major yolk protein in ovulated eggs), and lipids were measured. All larvae hatched by the 2nd day after fertilization at a water temperature of 23 °C, and the hatched larvae absorbed almost all of the yolk mass by the 10th day. The FAA composition in ovulated eggs is unique in that glutamine (Gln) is markedly high (32% of total FAA) in comparison to other pelagic eggs of marine teleosts suggesting the importance of Gln for early development of the Japanese eel. Total FAA content decreased to 45% of the initial level by the 4th day after fertilization. Gln content showed a rapid decrease to 5% of the initial level within 2 days after fertilization. oLv content, measured by enzyme-linked immunosorbent assay using an antiserum against lipovitellin, decreased until the 4th day and was, thereafter, undetected. Total content of triacylglycerol (TG) was stable until the 2nd day, and then decreased until the 8th day. On the other hand, the content of phospholipids (PL) gradually decreased until the end of yolk sac absorption. From these data, we divided the pattern of nutrient utilization in embryos and larvae of Japanese eel into two phases: first, utilization of FAA (especially Gln) and oLv from the 1st day, followed by TG utilization from the 2nd to 4th day by which time more than 30% of the FAA and the oLv stock have been utilized. In addition, PL was utilized as a subsidiary energy source throughout development. Almost all the yolk nutrient were absorbed by the 8th day after fertilization, and it is considered that the larvae need to start feeding before this period. [Copyright &y& Elsevier]