The zona pellucida surrounding the mammalian oocyte contains a major glycoprotein species, ZP3, that serves as a cell- and species-specific receptor for spermatozoa. In this study we have determined the primary amino acid structure of marmoset ZP3 (marZP3) and examined the expression of marZP3 mRNA within the ovary. The marZP3 gene possesses an open reading frame of 1272 nucleotides which is expressed specifically by the oocyte and encodes a polypeptide chain of 424 amino acids that exhibits 91% homology with the human ZP3 sequence. The disparity between these molecules was confined to a short domain spanning residues 322–352; otherwise the molecules were very similar, showing conservation of many structural features including the N–linked glycosylation sites, location and number of cysteine and proline residues and hydrophobicity profile. The results of this study have important implications for the use of the marmoset monkey as an animal model for the development of contraceptive vaccines targeting ZP3.

Staurosporine, an inhibitor of protein kinase activity, causes premature intercellular flattening of blastomeres but does not induce their premature polarisation. The flattening induced is calcium dependent, is reversed transiently at mitosis and requires the continuing presence of the drug. Staurosporine also blocks the decompacting effect of phorbol ester on 8-cell embryos.

Spermatozoa from the mollusc Ostrea edulis are capable of fusing to and entering de-chorionated ascidian eggs. During interaction they generate activation currents, comparable to the fertilisation currents induced by homologous spermatozoa. Activation currents are inward at − 80 mV, with a mean initial slope of 111 ± 124 pA/s for Ciona intestinalis eggs and 47 ± 25 pA/s for Phallusia mammillata eggs, while the mean peak currents are 2782 ± 1132 pA and 1523 ± 1668 pA, respectively. The fertilisation and activation currents reverse at a holding potential of 0 mV to + 20 mV, suggesting that oyster sperm and ascidian sperm gate the same channel precursor, a non-specific, large conductance channel described previously (Dale & DeFelice, 1984). In contrast to homologous fertilisation, the activation current is not followed by a polarised contraction of the egg surface, nor other signs of egg activation. Staining eggs with Hoechst 33342 after insemination shows the female nucleus and a single oyster sperm nucleus at the antipode. This suggests a specialised predetermined site at the vegetal pole for sperm entry. Homologous and heterologous spermatozoa delivered, in a large pipette, to localised areas of the egg surface generate fast inward currents of 200–2000 pA, but do not induce contraction of the egg surface. This shows that although channel precursors are located globally over the egg surface, channel activation does not necessarily trigger the contraction wave. Subsequent induction of both a fertilisation current and a contraction by homologous sperm added to the bath, implies a regionalised activation site with an accumulation of channel precursors and a ‘pacemaker’ for the initiation of the contraction wave.

In the starfish, spermatozoa undergo the acrosome reaction upon encountering the jelly coat of eggs. A highly sulphated glycoprotein in the jelly coat is called acrosome-reaction-inducing substance (ARIS) because it is the key signal molecule to trigger the acrosome reaction. The activity of ARIS is mainly attributed to its sulphate and saccharide residues. The extremely large molecular size and speciesspecific action of ARIS suggest the presence of a specific ARIS receptor on the sperm surface, but no experimental evidence for the receptor has been presented. We therefore measured specific binding of ARIS and its pronase digest (P-ARIS), which retains the full activity of ARIS, to homologous spermatozoa by using fluorescien-isothiocyanate-labelled ARIS and125 I-labelled P-ARIS, respectively. The spermatozoa had the ability to bind ARIS, as well as P-ARIS, specifically. The binding was species-specific, and mostly localised to the head region of spermatozoa. Scatchard plot analysis indicated the presence of one class of ARIS receptor on the surface of acrosome-intact speramatozoa. Furthermore, the specific binding of P-ARIS to the anterior region of sperm heads was microscopically confirmed by using P-ARIS conjugated to polystyrene latex beads with intense fluorescence. It is concluded that starfish spermatozoa have a specific receptor for ARIS on the surface of the anterior region of heads.

Intracellular membranes isolated from fully grown immature oocytes, mature oocytes (eggs) and activated eggs of Xenopus laevis were fractionated through a discontinuous sucrose density gradient into light, intermediate and heavy fractions. Electron microscopy showed that the light and intermediate fractions consisted mainly of smooth membranes, while the heavy fraction consisted mainly of rough membranes and mitochondria. Variations in the proteins associate with samples taken at different stages were observed by SDS-PAGE. The following differences were consistently observed: a 200 kDa protein was present only in the intermediate fraction of activated eggs, 29 and 44 kDa proteins were present only in the intermediate fractions of immature oocytes and activated eggs, and 120 and 145 kDa proteins were present only in the heavy fractions of mature oocytes and activated eggs. Examination of Western blots showed that cyclins A and B2 did not associate with membrane fractions at any stage of meiosis. Instead, cyclin A was present in the cytosols of mature oocytes and cyclin B2 was present in the cytosols of immature and mature oocytes. c-mos protein was detected in the cytosols and occasionally in the light fractions of mature oocytes and activated eggs. While α- and β-tubulins were detected in the light and intermediate fractions at all the stages of meiosis examined, only β-tubulin was present in the heavy fraction. βtubulin present in the heavy fraction was detected only at interphase, i.e. in immature oocutes and activated eggs, and not in mature oocytes. Using immunogold labelling we confirmed these results and found evidence to suggest that β-tubulin associates with the rough endoplasmic reticulum of interphase cells by a linking protein.

The cytoskeletal organisation of the isolated embryo sac and egg cells of Plumbago zeylanica was examined before, during and after pollen tube penetration into the embryo sac to determine the potential involvement of microtubules and actin filaments in fertilisation. Material was singly and triply stained using Hoechst 33258 to localise DNA, fluorescein isothiocyanate (FITC)-labelled anti- α-tubulin to detect microtubules and rhodamine-phalloidin to visualise F-actin. Microtubules in the unfertilised egg cell are longitudinally aligned in the micropylar and mid-lateral areas, aggregating into bundles near the filiform apparatus. In the perinuclear cytoplasm of the egg cell, microtubules become more or less randomly aligned. F-actin bundles form a longitudinally aligned mesh in the chalazal cytoplasm of the egg cell. In the central cell, microtubules and F-actin are distributed along transvacuolar strands and are also evident in the perinuclear region and at the periphery of the cell. During pollen tube penetration, sparse microtubule bundles near the pathway of the pollen tube may form an apparent microtubular ‘conduit’ surrounding the male gametes at the delivery site. Actin aggregates become organised near the pathway of the pollen tube and at the delivery site of the sperm cells. Subsequently, actin aggregates form a ‘corona’ structure in the intercellular region between the egg and central cell where gametic fusion occurs. The corona may have a role in maintaining the close proximity of the egg and central cell and helping the two sperm cells move and bind to their target cells. The cytoskeleton may also be involved in causing the two nuclei of the egg and central cell to approach one another at the site of gametic fusion and transporting the two sperm nuclei into alignment with their respective female nucleus. The cytoskeleton is reorganised during early embryogenesis.

Evidence of sexual dimorphism before fetal gonadal differentiation in mammals has been accumulating, suggesting that male embryos develop faster than female ones. The current investigation was performed to evaluate whether the development rate of precompacted human embryos is controlled by sex chromosomes. Sex was determined by polymerase chain reaction and fluorescence in situ hybridisation in 172 arrested embryos derived from in vitro fertilisation. The sex ratio (1.02:0.98) did not differ significantly from 1:1. Although more males appeared to have greater fragmentation, the difference between the sex ratios of highly fragmented and normal embryos (1.08:0.92) was not significant. Arrested female embryos had a tendency to exhibit more than five nuclei and less than 10% fragmentation, but the trend was not statistically significant. The current results suggest that the first developmental block in human embryos occurs prior to and shortly after genomic activation and is not determined by the presence of the Y chromosome.

The plasma membrane (oolemma) of the hamster egg retains the ability to fuse with spermatozoa even after exhaustive treatment with proteases and glycosidases. In contrast, when mouse oolemma is treated with proteases, the ability of eggs to fuse with spermatozoa is reduced. In the present study, similar treatments effective in reducing fusibility in the mouse were reexamined in the hamster. Of the several enzymes and treatments tested, only trypsin in Ca2+-free medium significantly reduced the hamster oolemma's ability to fuse with spermatozoa. This is suggestive of a cadherin-like system of binding and fusion. When hamster oolemmae were treated with the same protease regimen that reduced fusibility of mouse oolemma for mouse spermatozoa, heterologous fusion of hamster oolemmae with mouse spermatozoa was reduced, without affecting the fusion of these oolemmae with hamster spermatozoa. These data suggest that a protease-sensitive oolemma molecule is of critical importance for mouse sperm-oolemma fusion but not for hamster sperm-oolemma fusion.

Several ultrastructural changes occur during dehydration and subsequent rehydration of Arabidopsis thaliana pollen. The cytoplasmic channels, present in the outer part of the intine of the mature, dehydrating pollen grain, degenerate and develop into electron-dense inclusions. At the same time a large quantity of electron-dense material is deposited in the cavities of the exine. A large number of vesicles is produced in the vegetative cell, and they become predominantly located in the peripheral region near the intine. Starch of amyloplasts is consumed and the lipid bodies which originally surround the sperm cells become randomly distributed. In addition, the individual lipid bodies become enveloped by single rough endoplasmic reticulum cisterns.