Two forms of vitellogenin (Vg), Vg A and Vg B, were identified in serum from estrogen-treated barfin flounder (Verasper moseri). Structural changes of lipovitellins (Lvs) derived from the two Vgs were examined during vitellogenesis and oocyte maturation. Two Lvs, vLv A and vLv B, were identified electrophoretically and immunologically in postvitellogenic oocytes. Each appeared to be composed of distinct heavy chains (vLvH A, M(r) 107,000, and vLvH B, M(r) 94,000) and light chains (vLvL A, M(r) 30,000, and vLvL B, M(r) 28,000) when analyzed by SDS-PAGE. Results from N-terminal amino acid sequencing and Western blotting using antisera to vLvH A and vLvH B verified that there are two Vg polypeptides in serum from estrogen-treated fish, Vg A (M(r) 168,000) and Vg B (M(r) 175,000), which give rise to vLvH A-vLvL A and vLvH B-vLvL B, respectively. N-terminal sequencing revealed two sequences for both phosvitin and beta'-component, supporting the concept of duality for all three classes of Vg-derived yolk proteins. During oocyte maturation, native dimeric vLv B was dissociated into a native M(r) 170,000 monomer (oLv B). Meanwhile, vLv A was extensively cleaved including complete degradation of vLvH A into free amino acids. We propose that the quantitative ratio of vLv A to vLv B in postvitellogenic oocytes regulates the buoyancy of the spawned pelagic eggs by controlling availability of free amino acids which function as osmotic effectors during oocyte hydration. The vLv A/vLv B ratio likely also controls the proportional availability of different types of nutrients, free amino acids versus Lv, for use during embryonic development.

Three complete cDNAs encoding different forms of vitellogenin (Vtg) were isolated from a white perch (Morone americana) liver cDNA library and characterized with respect to immunobiochemical and functional features of the three Vtgs and their product yolk proteins (YPs) in this species and in the congeneric striped bass (Morone saxatilis). The two longest cDNAs encoded Vtgs with a complete suite of yolk protein domains that, based on comparisons with vtg sequences from other species, were categorized as VtgAa and VtgAb using the current nomenclature for multiple teleost Vtgs. The shorter cDNA encoded a Vtg that lacked a phosvitin domain, had a shortened C-terminus, and was categorized as VtgC. Mapping of peptide sequences from the purified Vtgs and their derived YPs to Vtg sequences deduced from the cDNAs definitively identified the white perch VtgAa, VtgAb, and VtgC proteins. Detailed comparisons of the primary structures of each Vtg with partial or complete sequences of Morone yolk proteins or of Vtgs from other fishes revealed conserved and variant structural elements of teleost Vtgs with functional significance, including, as examples, signal peptide cleavage sites, dimerization sites, cathepsin D protease recognition sites, and receptor-binding domains. These comparisons also yielded an interim revision of the classification scheme for multiple teleost Vtgs.

Full-length cDNAs encoding three forms of vitellogenin (Vg) were obtained from a liver cDNA library of estrogen-treated red seabream, Pagrus major. Two of the three Vg sequences had high homology with type-A and -B Vgs (VgA and VgB) of other teleosts. The third red seabream Vg was classified as a type-C or phosvitinless (Pvl) Vg due to its lack of a phosvitin (Pv) domain. Two Vg preparations (610 and 340 kDa) from blood serum of estradiol-treated fish were biochemically characterized. Analyses of precursor-product relationships by examination of N-terminal amino acid sequences verified cleavage of the 610 kDa Vg into a 540 kDa lipovitellin (Lv) and a 32 kDa beta'-component. Each of these yolk preparations comprising both VgA- and VgB-derived polypeptides. The 340 kDa Vg, which was immunologically verified to be a PvlVg, was accumulated by vitellogenic oocytes with no alterations to its native molecular mass. During oocyte maturation, the VgA- and VgB-derived yolk proteins were differentially processed, presumably to generate a pool of free amino acids for oocyte hydration or for allocation of specific types of nutrients, amino acids, and proteins, to the developing embryo. Conversely, the 340 kDa Vg-derived yolk protein is unlikely to contribute to oocyte hydration or diffusible nutrients since the molecule underwent only minor proteolytic nicking during oogenesis. The present study elucidates for the first time specific functions of three different forms of Vg and their product yolk proteins in a higher taxonomic group of marine teleosts that spawn pelagic eggs.

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Vitellogenin (Vg), a major estrogen-inducible yolk precursor protein, has become an important biomarker for assessing the estrogenic potency of chemicals and the exposure of animals to estrogenic contaminants present in aquatic environments. These contaminants, which can disrupt functioning of the vertebrate neuroendocrine system, are

The production of fertile eggs with the capacity to develop into larvae and subsequently into marketable fish is centrally important to the aquaculture industry. This entails not only the programmed production of large numbers of eggs, but also high quality eggs with the potential to support normal development and high survival of offspring to juvenile and later stages of development and growth. Numerous studies highlight the maternal contributions to the development of embryos, including transcripts that regulate cell division and determine oocyte polarity, pattern development during early and late embryonic stages and the transition from maternal to zygotic gene expression and translation. Since most fish embryos develop independently within an enclosed egg envelope, they rely on compounds deposited within the oocytes during their various stages of development. In addition to regulatory nucleic acids (maternal DNA and RNA), these include proteins and other compounds that contribute to the structure and function of the egg envelope and the bulk molecular cargo that will be used as a source of cellular energy and structural components for formation of embryos and larvae. These latter components notably include yolk lipids and proteins deposited during oocyte growth and water acquired at the same time and during cytoplasmic maturation. In this review we highlight recent advances made in revealing the transcripts deposited within the oocyte that contribute to the structural and morphological development of the embryo, and to the regulation of gene expression and translation during oocyte development. Significant advances have been made in revealing the molecular mechanisms of lipid accumulation and metabolism within the oocyte, the intricacies of yolk protein formation via endocytosis of multiple yolk precursor proteins by multiple oocyte receptors, and the complex machinery supporting massive accumulation of water by maturing oocytes of many species. Additionally, many advances have been made in our understanding of the endocrine regulation of all of these processes during oogenesis. We provide here an overview of recent advances in our knowledge on these various aspects of oogenesis and identify several gaps in our knowledge for future studies.

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; 3U.S. Environmental Protection Agency, Gulf Breeze, Florida Vitellogenin (VTG), the serum phospholipoglycoprotein precursor to egg yolk, is potentially an ideal biomarker for environmental estrogens. This study was undertaken to develop antibodies against conserved regions on the VTG molecule that could form the basis for establishing bioassays to detect estrogen exposure in any oviparous vertebrate. We developed monoclonal antibodies (mAbs) generated against purified rainbow trout (Oncorhynchus mykiss) VTG and selected for the property of specifically recognizing VTG purified from two phylogenetically distant vertebrates, trout and striped bass (Morone saxatilis). Results of enzyme-linked immunosorbent assay and Western blotting indicated that these mAbs specifically recognize purified VTG and VTG or other estrogen-inducible proteins in plasma or serum from representative species of four vertebrate classes (fish, amphibians, reptiles, and birds). All of the mAbs generated were IgM class. A polyclonal antiserum was raised against a synthetic consensus peptide representing the conserved N-terminal amino acid sequence of VTG. The results of Western blotting indicate that this antiserum specifically recognizes VTG in plasma or serum from teleost fish of diverse families. It was used to detect VrG in Western blots of serum from brown bullhead (Ameiurus nebulosus) with cancer (hepatocellular and cholangio-carcinoma) collected from a contaminated industrial site outside of their normal vitellogenic season. Our results indicate that it is feasible to generate antibodies capable of recognizing VTG without regard to species and that development of a universal VTG assay is an achievable goal. -Environ Health Perspect 1 03(Suppl 7): 9-15 (1995)