Modifications of carbohydrate residues and ZP2 and ZP3 glycoproteins in the mouse zona pellucida after fertilization

Oligosaccharide side chains of zona pellucida (ZP) glycoproteins play a key role in the sperm-egg interaction phenomena during fertilization. In the present study, modifications of the ZP glycoproteins during the fertilization process in the mouse were studied by the lectin-gold technique and immunocytochemistry in conjunction with quantitative analysis. Binding of PNA, RCA I, DSA, LFA, MAA, AAA, and anti-ZP2 and anti-ZP3 antibodies was observed throughout the ZP of both unfertilized and fertilized eggs. However, HPA and BSAIB4 labeling was found only in the inner region of the ZP. After neuraminidase treatment (Neu), HPA showed an affinity for the entire ZP. Labeling by LFA, WGA, MAA, PNA, BSAIB4, and AAA decreased in the ZP of fertilized eggs; however, there was an increase in the binding of RCA I. HPA and Neu-HPA increased only in the inner region of the ZP. Immunoreactivity to antibodies against ZP2 and ZP3 also decreased after fertilization. The present results demonstrate that 1) terminal carbohydrate residues contained in the ZP glycoproteins are modified after fertilization and 2) inner and outer regions of the ZP contain different oligosaccharide side chains.     

Isolation, physicochemical properties, and the macromolecular composition of the vitelline and fertilization envelopes from Xenopus laevis eggs

As a step toward defining in molecular terms the sperm-triggered block to polyspermy reaction established by the egg at fertilization, vitelline (VE) and fertilization (FE) envelopes were isolated from eggs of the Sounth African clawed toad Xenopus laevis and some of their physicochemical properties determined. Envelopes were isolated after lysis of the fertilized or unfertilized eggs by sieving techniques; isolated envelopes retained their in situ morphology as determined by electron microscopy. The isolated envelopes had different solubility properties and, in general, VE was more readily dissolved by aqueous solvents than FE, although both could be completely dissolved by detergents or chaotropic agents. Changes in envelope solubility correlated with the progression of the cortical reaction implicating a role for cortical granule material in modifying the solubility properties of the envelope. The VE and FE were composed of protein and carbohydrate with no lipid components detected. As determined by immunodiffusion experiments, the FE contained the same antigens as the VE plus components derived from the cortical granules and the innermost jelly layer, J. The macromolecular composition of the envelopes was determined by sodium dodecyl sulfate gel electrophoresis. The VE contained at least 11 glycoproteins with molecular weights ranging from 125 000 to less than 16 000 with two components (40 000 and 33 000) accounting for almost two-thirds of the total stainable material. The FE contained ten glycoproteins that had the same molecular weights as those in the VE. One glycoprotein component underwent a reduction in molecular weight from 77 000 to 67 500 when the VE was converted to the FE. This molecular weight change was interpreted as the probable result of limited proteolysis. In addition, the FE gel electrophoresis patterns contained macromolecular components derived from the cortical granules and jelly layer, J, consistent with the immunodiffusion experiments. These components were absent when the FE was prepared in the absence of Ca2+, suggesting a role for Ca2+ in binding the VE, cortical granules, and J components together. We concluded that the conversion of the glycoproteinaceous VE to FE at fertilization is caused by interaction of the VE with components from the cortical granules and jelly layer J. These interactions are of both a chemical and physical nature.     

Cytochalasin B-induced triploidy in mouse oocytes fertilized in vitro

Mouse eggs and spermatozoa were treated in various ways with 5 or 10 mug cytochalasin B/ml. The fertilization rate in vitro was reduced by treatment with the drug but 80-90% of the eggs fertilized were triploid. Many of the experimental eggs were penetrated by one or more spermatozoa but remained unfertilized (75% compared with 9% in control eggs). It is suggested that cytochalasin B weakens the zona reaction and interferes with fusion of gametes but does not prevent the block to polyspermy.     

The 350-kDa sea urchin egg receptor for sperm is localized in the vitelline layer

Previous studies have established by several methods that the 350-kDa egg receptor for sperm is localized on the plasma membrane-vitelline layer complex of the egg of the sea urchin Strongylocentrotus purpuratus. In addition, it has been found that molecules which are cross-reactive with anti-receptor antibody are present in the cortical granules located at the inner face of the plasma membrane. The objective of this study was to define more precisely the locale of the cell surface receptor. We have found that following fertilization, the immunoreactive receptor initially found on the egg surface moved to the fertilization envelope (FE) and then disappeared in parallel with the loss of sperm binding activity. A cross-linked, high-molecular-weight derivative of soybean trypsin inhibitor (hMW-SBTI) which was unable to pass through the elevating FE blocked the loss of both immunoreactivity and the sperm binding activity of the FE, but did not inhibit the vitelline delaminase activity that has been implicated in FE formation. Western blot analysis following SDS-PAGE of the proteins of the FE isolated in the presence of hMW-SBTI and benzamidine revealed the presence of the 350/320-kDa proteins which cross-reacted with anti-receptor antibody. Experiments in which molecules on the surface of unfertilized eggs were labeled with biotin and traced after FE formation revealed that a significant portion of the 350/320-kDa glycoproteins that were incorporated into the FE originated from the cell surface, rather than from the cortical granules. These findings provide strong evidence that in unfertilized eggs the egg receptor for sperm exists as part of the protein complex known as the vitelline layer which serves as a precursor of the FE. Evidence is presented indicating that some of the receptor in the vitelline layer is cryptic and a possible function for this cryptic form of the receptor is proposed.     

The molecular genetics of the zona pellucida: mouse mutations and infertility

The zona pellucida is an extracellular matrix surrounding growing oocytes, ovulated eggs and the preimplantation embryo. After mediating the relatively species-specific events of fertilization, the zona pellucida provides a post-fertilization block to polyspermy and protects the growing embryo as it passes down the oviduct. The genes that encode the three zona pellucida proteins (ZP1, ZP2, ZP3) have been characterized in mouse and human. The ability to genetically manipulate the zona pellucida genes in mouse models has enhanced our knowledge of zona pellucida structure and function in vivo and may translate into a better understanding of human fertility.     

Xenopus laevis sperm receptor gp69/64 glycoprotein is a homolog of the mammalian sperm receptor ZP2

Little is known about sperm-binding proteins in the egg envelope of nonmammalian vertebrate species. We report here the molecular cloning and characterization of a recently identified sperm receptor (gp69/64) in the Xenopus laevis egg vitelline envelope. Our data indicate that the gp69 and gp64 glycoproteins are two glycoforms of the receptor and have the same number of N-linked oligosaccharide chains but differ in the extent of O-glycosylation. The amino acid sequence of the receptor is closely related to that of the mouse zona pellucida protein ZP2. Most of the sequence conservation, including a ZP domain, a potential furin cleavage site, and a putative transmembrane domain are located in the C-terminal half of the receptor. Proteolytic cleavage of the gp69/64 protein by a cortical granule protease during fertilization removes 27 amino acid residues from the N terminus of gp69/64 and results in loss of sperm binding to the activated eggs. Similarly, we find that treatment of eggs with type I collagenase removes 31 residues from the N terminus of gp69/64 and has the same effect on sperm binding. The isolated and purified N terminus-truncated receptor protein is inactive as an inhibitor of sperm-egg binding. Earlier studies on the effect of Pronase digestion on receptor activity suggest that this N-terminal peptide may contain an O-linked glycan that is involved in the binding process. Based on these results and the findings on the primary structure of the receptor, a pathway for the maturation and secretion of gp69/64, as well as its inactivation following fertilization, is proposed.     

Relative changes in F-actin during the first cell cycle: evidence for two distinct pools of F-actin in the sea urchin egg

The cortical actin cytoskeleton undergoes dramatic rearrangements during fertilization of sea urchin eggs. To characterize these changes further, we quantified the relative changes in filamentous actin (F-actin) during fertilization and the first cell cycle in both intact eggs and in isolated cortices by quantitative fluorescence microscopy. The level of F-actin in the intact egg decreased after fertilization and continued to decrease throughout the first cell cycle. By 60 min after fertilization, the level of F-actin had decreased to 50% of the unfertilized sea urchin egg. By cytokinesis, the level of F-actin had decreased to 30% of the unfertilized egg. After completion of cell division, individual blastomeres had 10% of the F-actin in the unfertilized egg. In contrast, there was an increase in cortical F-actin to 370% of the level in the unfertilized egg after fertilization. This increase corresponded to the formation of microvilli. There was little change in the level of cortical F-actin during the first cell cycle. We draw parallels to other systems that increase the amount of F-actin in the Triton-insoluble cytoskeleton by recruiting actin from a Triton-soluble pool of F-actin.     

Meiotic cells monitor the status of the interhomolog recombination complex

Cortical granule exocytosis is important for the block to polyspermy at fertilization in the eggs of most vertebrates and many invertebrates. Cortical granules are poised at the cell surface and exocytose in response to sperm stimulation. Following exocytosis, the cortical granule contents modify the extracellular environment of the egg, the major result of which is to block additional sperm binding. Here we show that proteins homologous to members of the SNARE hypothesis-a molecular model designed to explain the trafficking, docking, and exocytosis of vesicles in the secretory compartment-are present in eggs at the right time and place to be involved in the regulation of cortical granule exocytosis. Using polymerase chain reaction (PCR) screens we have found homologues of synaptobrevin/VAMP, syntaxin, synaptotagmin, and rab3. Antibodies generated to fusion proteins or to synthetic peptides encoded by the cloned cDNAs were used in an immunofluorescence assay to show that each of the cognate proteins are present in the cortex of the egg. A synaptobrevin/VAMP homologue appears to be specifically associated with the membrane of cortical granules before fertilization and, following cortical granule exocytosis, is incorporated into the plasma membrane of the zygote. A rab3 homologue is also associated with cortical granules specifically but, following fertilization, the protein reassociates with different, yet undefined, vesicles throughout the cytoplasm of the zygote. Homologues of synaptotagmin and syntaxin are also present at the egg cortex but, in contrast to rab3 and VAMP, appear to be associated with the plasma membrane. Following fertilization, syntaxin and tagmin remain associated with the plasma membrane and are more readily immunolabeled, presumably due to an increased accessibility of the antibodies to the target protein domains. We also show by immunoblotting experiments that the cognate proteins are of the sizes predicted for these homologues. These results suggest that at least some steps in the biology of cortical granules may be mediated by SNARE homologues, and this finding, along with the unique biology of cortical granules, should facilitate examination of specific events of the fertilization reaction and the mechanism of stimulus-dependent exocytosis.     

Human ZP3 restores fertility in Zp3 null mice without affecting order-specific sperm binding

The mammalian zona pellucida surrounding ovulated eggs mediates sperm binding at fertilization, provides a postfertilization block to polyspermy, and facilitates passage of pre-implantation embryos down the oviduct. Although the three zona proteins (ZP1, ZP2, ZP3) are well conserved, mammalian fertilization is relatively specific and human sperm do not bind to the mouse zona pellucida. There are considerable in vitro data that ZP3 acts as a primary sperm adhesion molecule in mice and, by analogy, a similar role has been postulated for human ZP3. Genetically altered mice lacking ZP3 (Zp3(tm/tm)) do not form a zona pellucida and are infertile. To rescue this phenotype, transgenic mice expressing human ZP3 (67% identical to mouse ZP3) were produced and bred with Zp3(tm/tm) null mice. The resultant human ZP3 rescue females had chimeric zonae pellucidae composed of mouse ZP1, mouse ZP2 and human ZP3. Human ZP3 expressed in mouse oocytes had an apparent mass (64 kDa) indistinguishable from native human ZP3 and distinct from mouse ZP3 (83 kDa). Despite the presence of human ZP3, human sperm did not bind to the chimeric zona pellucida, and notwithstanding the absence of mouse ZP3, mouse sperm bound to ovulated eggs in vitro and fertility was restored in vivo. These data have implications regarding the molecular basis of mouse and human sperm binding to their respective zonae pellucidae.     

Heterogeneity of the zona pellucida carbohydrate distribution in human oocytes failing to fertilize in vitro

The mammalian zona pellucida contains several glycoproteins whose oligosaccharide moieties are known to play a key role in the interaction with spermatozoa. Since zona pellucida defects may represent one of the most likely causes of failed fertilization in human in-vitro reproduction, we have studied the carbohydrate composition and distribution over the human zona pellucida by means of lectins. Donated, not inseminated cumulus-oocyte complexes, from cohorts with high fertilization rates, and fertilization-failed oocytes from cohorts inseminated with proven fertile donor semen, were analysed using 11 fluorescein-labelled lectins, on deplasticized semi-thin epoxy sections. Results showed that wheat germ agglutinin (WGA), Maclura pomifera (MPA) and Pisum sativum (PSA) bound to the extracellular matrix bordering the zona pellucida-corona radiata interface of cumulus-oocytes complexes, while the zona pellucida was labelled by WGA, Concanavalin A (ConA) and PSA. WGA labelling and correlative electron microscopy on the cumulus-oocyte complexes demonstrated that this lectin is a useful tool to trace the cortical granule distribution in the human oocyte. Surprisingly, in the failed-fertilized oocytes the zona pellucida was also labelled by MPA and showed three different patterns: (i) labelling of the zona pellucida outer surface; (ii) uniform labelling; (iii) labelling of an outer zona pellucida layer with variable thickness. Comparative analysis of WGA and MPA labelling on single failed-fertilized oocytes demonstrated that MPA zona pellucida patterns are not related to the cortical reaction. The nature and meaning of the MPA pattern of failed-fertilized oocytes were discussed in the light of zona pellucida defects impairing sperm receptivity.     

Evidence for a polyspermy block at the level of sperm-egg plasma membrane fusion in Urechis caupo

The results of sperm binding experiments reveal no change in the sperm binding properties of the egg surface coat at fertilization of Urechis caupo eggs. When fertilized eggs are reinseminated, sperm continue to attach to the egg surface coat. The acrosomal tubules of supernumerary sperm are observed in the perivitelline space closely apposed to the egg membrane. Thus, the polyspermy block in Urechis eggs involves neither alteration of sperm binding sites nor inhibition of the acrosome reaction. Our results suggest that the block is at the level of sperm-egg membrane fusion.     

Sperm and its soluble extract cause transient increases in intracellular calcium concentration and in membrane potential of sea urchin zygotes

Fertilization is known to initiate a transient increase of intracellular calcium concentration and an accompanying change of membrane potential in sea urchin eggs. If the fertilization membrane and hyaline layer are removed, sperm can again enter fertilized eggs (refertilization). We have found that Ca2+ and voltage transients were repeatedly induced in fertilized eggs during refertilization. Similar changes were also obtained by external application of a soluble extract of sperm to fertilized eggs. This sperm extract caused no changes in unfertilized eggs. The active factor in the sperm extract survives heating (100 degrees C, 10 min) and incubation with pronase. Its molecular weight is less than 1300.     

Binding of antibiotics to glycoproteins of the vitelline and fertilization envelopes of cherry salmon eggs

The binding of antibiotics (gentamicin, oleandomycin and chloramphenicol) to vitelline and fertilization envelopes and their extracts was investigated by immunohistochemical and immunocytochemical techniques and immunoblot analysis using mature and artificially activated eggs of the fish Oncorhynchus masou. Binding of antibiotics was detected in the vitelline and fertilization envelope outermost layers, the fertilization envelope inner surface and cortical alveolus exudates, with differences in immunoreactive intensity and deposition. The fertilization envelope outermost layer had the capacity to bind much greater amounts of the antibiotics than the vitelline envelope outermost layer. The greater capacity was caused by the deposition of cortical alveolus exudates, which were known to be responsible for functional roles of protection against bacteria, fungi and noxious materials. Treatment of the vitelline and fertilization envelopes with neuraminidase markedly reduced the binding of gentamicin and chloramphenicol but slightly increased that of oleandomycin; binding of the latter to the vitelline and fertilization envelope outermost layers was considerably reduced after treatment with alpha-fucosidase. Treatment of the two envelopes with alpha-mannosidase, beta-galactosidase or beta-D-glucosaminidase did not cause any alteration in immunoreactive intensity or number of immunoreactive deposits. Immunoblot analysis of the vitelline or fertilization envelope extracts indicated that many of the antibiotic-binding substances were glycoproteins, and several major bands were bound by all three antibiotics. These results suggest that the vitelline or fertilization envelopes may have the ability to protect the egg itself, or the embryo, respectively, by trapping antibiotics, and the trapping may be related to the presence of carbohydrate moieties, such as sialyl or fucosyl residues.     

Pilocarpine-induced changes in the saccharide composition of the tectorial membrane and interdental cells of the organ of Corti: a study with gold-labeled lectins

The glycoconjugates in the cytoplasm of inner ear interdental cells and those constituting the limbal tectorial membrane were identified by a post-embedding cytochemical method using low-temperature embedding in Lowicryl K4M and labeling with biotinylated lectins, goat anti-biotin antibody, rabbit anti-goat antibody, and gold-labeled protein A in control animals, and after the systemic injection of pilocarpine. The lectins used were ConA, PHA-E, PSA, RCA, SBA, Succ-WGA, UEA, and WGA. In control animals, a semiquantitive analysis of gold particles showed that Succ-WGA produced the strongest labeling on the tectorial membrane, followed by SBA, ConA, WGA, RCA, PHA-E, and PSA. The lowest values were obtained with UEA. The cytoplasm of the interdental cells was also labeled with all the lectins, but the number of particles/microns2 was lower than on the tectorial membrane. The concentration of gold particles on the limbal tectorial membrane in pilocarpine-treated animals was higher than in control animals for some lectins (RCA, PSA, UEA) but lower for others (WGA, SBA, PHA-E, Succ-WGA). The changes in the labeling pattern of the cytoplasm of the interdental cells paralleled those in the tectorial membrane. These results demonstrate that the saccharide composition of the limbal tectorial membrane can be modified by systemic injection of pilocarpine. This action may take place through a change in either the secretion rate or the amount of some glycoconjugates by the interdental cells.     

Survival, metabolic activity and conjugative interactions of indigenous and introduced streptomycete strains in soil microcosms

Exocytosis of cortical granules in mouse eggs is required to produce the zona pellucida block to polyspermy. In this study, we examined the role of microfilaments and microtubules in the regulation of cortical granule movement toward the cortex during oocyte maturation and anchoring of cortical granules in the cortex. Fluorescently labeled cortical granules, microfilaments, and microtubules were visualized using laser-scanning confocal microscopy. It was observed that cortical granules migrate to the periphery of the oocyte during oocyte maturation. This movement is blocked by the treatment of oocytes with cytochalasin D, an inhibitor of microfilament polymerization, but not with nocodazole or colchicine, inhibitors of microtubule polymerization. Cortical granules, once anchored at the cortex, remained in the cortex following treatment of metaphase II-arrested eggs with each of these inhibitors; i.e., there was neither inward movement nor precocious exocytosis. Finally, the single cortical granule-free domain that normally becomes localized over the metaphase II spindle was not observed when the chromosomes become scattered following microtubule disruption with nocodazole or colchicine. In these instances a cortical granule-free domain was observed over each individual chromosome, suggesting that the chromosome or chromosome-associated material, and not the spindle, dictates the localization of the cortical granule-free domain.     

Determination of DNA damage in F344 rats induced by geometric isomers of tamoxifen and analogues

The transformation of Trypanosoma cruzi epimastigotes to mammal-infective metacyclic trypomastigotes (metacyclogenesis) can be performed in vitro under chemically defined conditions (TAU 3AAG medium). During this process, changes in the nature of cell surface sugar composition and sugar distribution was evaluated using FITC and gold-labeled lectins and observed by flow cytometry and transmission electron microscopy. The pattern of labeling with the lectins from Triticum vulgaris (WGA), Arachis hypogaea (PNA), Limax flavus (LFA), Canavalia ensiformis (Con-A), and Ricinus communis (RCA-I) significantly changed during the metacyclogenic process. The results obtained are discussed in relation to the role played by T. cruzi cell surface carbohydrate residues on the process of parasite-host cell interaction.     

Electroencephalographic changes in experimental autoimmune myasthenia gravis

A procedure for the determination of activity and linkage specificity of sialidases is described. The sialoglycoprotein fetuin is coated onto a microtiter plate and incubated with sialidases from different sources. Enzymatic activities and linkage specificities are then determined by a sandwich method which measured the binding of different lectins to fetuin. The lectins used were peanut agglutinin (PNA) from Arachis hypogaea, which binds specifically the galactose beta-1-3-N-acetylgalactosamine structures that are unmasked following sialidase treatment of fetuin, the lectins from Sambucus nigra (SNA) and Maackia amurensis (MAA) that are specific for alpha-2-6 and alpha-2-3 bound sialic acids, respectively, and the slug agglutinin from Limax flavus (LFA) that is specific for N-acetyl and N-glycolyl neuraminic acids. Increased PNA and decreased LFA, SNA, and MAA lectin binding correlated with sialidase-induced desialylation of the substrate. In this report, the assay was used to determine the activities and specificities of influenza, Vibrio cholerae, and Arthrobacter ureafaciens sialidases.     

Regulation of the polyspermy block in the mouse egg: maturation-dependent differences in cortical granule exocytosis and zona pellucida modifications induced by inositol 1,4,5-trisphosphate and an activator of protein kinase C

Germinal vesicle (GV)-intact fully grown mouse oocytes do not undergo cortical granule (CG) exocytosis in response to A23187 treatment, whereas metaphase II (MII)-arrested eggs do. This differential response may reflect the development of the ability of the egg to undergo CG exocytosis, which is responsible for the biochemical modification of the glycoprotein ZP2 in the zona pellucida. Accordingly, we compared in these two stages the ability of 12-O-tetradecanoyl phorbol 13-acetate (TPA) or inositol 1,4,5-trisphosphate (IP3) to promote CG exocytosis and/or the ZP2 to ZP2f conversion; these agents are known to stimulate early events of mouse egg activation. TPA (10 ng/ml) treatment for 60 and 120 min resulted in a 25% and 52% CG loss in GV-intact oocytes and a 38% and 76% loss in MII eggs, respectively; fertilization resulted in a CG loss of approximately 70-80%. Although a similar extent of ZP2 to ZP2f conversion was observed in oocytes and eggs after a 120-min TPA treatment (approximately 70-80%), a greater extent of conversion was observed in oocytes after a 60-min treatment (80% for oocytes, 50% for eggs). Microinjection of IP3 (final concentration 1 microM) into MII eggs resulted in an extent of ZP2 conversion similar to that observed following fertilization, whereas little conversion occurred in GV-intact oocytes similarly injected. These results indicate that a protein kinase C sensitivity develops prior to meiotic maturation, whereas responsiveness to IP3 develops after maturation has resumed. We propose that the regulatory mechanism involving an IP3-mediated calcium release is deficient in GV-stage oocytes.     

Surface alterations of the bovine oocyte and its investments during and after maturation and fertilization in vitro

Surface characteristics of the bovine oocyte and its investments before, during, and after maturation, and fertilization in vitro were evaluated by scanning electron microscopy (SEM). Oocyte diameters were also measured during SEM analysis of the oocyte. The cumulus cells manifested a compact structure with minimal intercellular spaces among them in the immature oocytes. These became fully expanded with increased intercellular spaces after maturation in vitro, but contracted again after fertilization. The zona pellucida (ZP) showed a fibrous, open mesh-like structure in the maturing and matured oocytes. The size and number of meshes on the ZP decreased dramatically after fertilization. The vitelline surface of immature oocytes was characterized by distribution of tongue-shaped protrusions (TSPs) varying in density. After 10 and 22 hr of maturation incubation, oocyte surface microvilli (MV) increased to become the predominant surface structure, and TSPs decreased substantially. The vitelline surface of fertilized oocytes (at 6 and 20 hr) was similar to that of the matured oocytes, but unfertilized oocytes had less dense MV than did fertilized oocytes (at 20 hr). The diameter of the oocytes decreased from 99 to 80 microns during maturation and increased to 106 microns after insemination (P < 0.05). Membrane maturation was characterized by surface changes from a TSP-predominant pattern to a MV-predominant pattern. Thus, the bovine oocyte maturation process was found to involve the expansion of cumulus cells and the maturation of the ZP, which changes dramatically upon fertilization. Also, volumetric changes occurred in ooplasm processed for SEM following oocyte maturation and insemination.     

On the occurence of isopeptide bonds in the chorion of the rainbow trout (Salmo gairdneri Rich.) (author's transl)

Chorions of unfertilized and fertilized eggs of the rainbow trout were isolated from tissue and yolk components and investigated for isopeptide bonds. The complete hydrolysis of the alpha-amide bonds was obtained by a system of four proteases. A chromatographic ion exchange system was used to separate Nepsilon-(beta-aspartyl)lysine as well as Nepsilon-(gamma-glutamyl)lysine from single amino acids. Chorions of unfertilized eggs contain neither Asp Lys nor Glu Lys bonds. Only chorions of fertilized eggs contain Glu Lys isopeptide. It is probably the high content of this isopeptide that is responsible for the greater mechanical stability of the fertilized teleostean egg chorion.