Ascidian Eggs Research Articles

MRNA Localization to the Endoplasmic Reticulum in Plant Endosperm Cells.

Subcellular mRNA localization is an evolutionarily conserved mechanism to spatially and temporally drive local translation and, in turn, protein targeting. Hence, this mechanism achieves precise control of gene expression and establishes functional and structural networks during cell growth and development as well as during stimuli response. Since its discovery in ascidian eggs, mRNA localization has been extensively studied in animal and yeast cells. Although our knowledge of subcellular mRNA localization in plant cells lags considerably behind other biological systems, mRNA localization to the endoplasmic reticulum (ER) has also been well established since its discovery in cereal endosperm cells in the early 1990s. Storage protein mRNA targeting to distinct subdomains of the ER determines efficient accumulation of the corresponding proteins in different endosomal storage sites and, in turn, underlies storage organelle biogenesis in cereal grains. The targeting process requires the presence of RNA localization elements, also called zipcodes, and specific RNA-binding proteins that recognize and bind these zipcodes and recruit other factors to mediate active transport. Here, we review the current knowledge of the mechanisms and functions of mRNA localization to the ER in plant cells and address directions for future research.

Dynamic changes in the association between maternal mRNAs and endoplasmic reticulum during ascidian early embryogenesis

Axis formation is one of the most important events occurring at the beginning of animal development. In the ascidian egg, the antero-posterior axis is established at this time owing to a dynamic cytoplasmic movement called cytoplasmic and cortical reorganisation. During this movement, mitochondria, endoplasmic reticulum (ER), and maternal mRNAs (postplasmic/PEM RNAs) are translocated to the future posterior side. Although accumulating evidence indicates the crucial roles played by the asymmetrical localisation of these organelles and the translational regulation of postplasmic/PEM RNAs, the organisation of ER has not been described in sufficient detail to date owing to technical difficulties. In this study, we developed three different multiple staining protocols for visualising the ER in combination with mitochondria, microtubules, or mRNAs in whole-mount specimens. We defined the internally expanded “dense ER” using these protocols and described cisterna-like structures of the dense ER using focused ion beam-scanning electron microscopy. Most importantly, we described the dynamic changes in the colocalisation of postplasmic/PEM mRNAs and dense ER; for example, macho-1 mRNA was detached and excluded from the dense ER during the second phase of ooplasmic movements. These detailed descriptions of the association between maternal mRNA and ER can provide clues for understanding the translational regulation mechanisms underlying axis determination during ascidian early embryogenesis.

Phylogenetic comparison of egg transparency in ascidians by hyperspectral imaging

The transparency of animals is an important biological feature. Ascidian eggs have various degrees of transparency, but this characteristic has not yet been measured quantitatively and comprehensively. In this study, we established a method for evaluating the transparency of eggs to first characterize the transparency of ascidian eggs across different species and to infer a phylogenetic relationship among multiple taxa in the class Ascidiacea. We measured the transmittance of 199 eggs from 21 individuals using a hyperspectral camera. The spectrum of the visual range of wavelengths (400–760 nm) varied among individuals and we calculated each average transmittance of the visual range as bio-transparency. When combined with phylogenetic analysis based on the nuclear 18S rRNA and the mitochondrial cytochrome c oxidase subunit I gene sequences, the bio-transparencies of 13 species were derived from four different families: Ascidiidae, Cionidae, Pyuridae, and Styelidae. The bio-transparency varied 10–90% and likely evolved independently in each family. Ascidiella aspersa showed extremely high (88.0 ± 1.6%) bio-transparency in eggs that was maintained in the “invisible” larva. In addition, it was indicated that species of the Ascidiidae family may have a phylogenetic constraint of egg transparency.

Non-centrosomal microtubule structures regulated by egg activation signaling contribute to cytoplasmic and cortical reorganization in the ascidian egg

In the first ascidian cell cycle, cytoplasmic and cortical reorganization is required for distributing maternal factors to their appropriate positions, resulting in the formation of the embryonic axis. This cytoplasmic reorganization is considered to depend on the cortical microfilament network in the first phase and on the sperm astral microtubule (MT) in the second phase. Recently, we described three novel MT structures: a deeply extended MT meshwork (DEM) in the entire subcortical region of the unfertilized egg, transiently accumulated MT fragments (TAF) in the vegetal pole, and a cortical MT array in the posterior vegetal cortex (CAMP). Particularly, our previous study showed CAMP to contribute to the movement of myoplasm. In addition, it is very similar to the parallel MT array, which appears during cortical rotation in Xenopus eggs. However, how these MT structures are organized is still unclear. Here, we investigated the relationship between the egg activation pathway and MT structures during the first ascidian cell cycle. First, we carefully analyzed cell cycle progression through meiosis I and II and the first mitosis, and successfully established a standard time table of cell cycle events. Using this time table as a reference, we precisely described the behavior of novel MT structures and revealed that it was closely correlated with cell cycle events. Moreover, pharmacological experiments supported the relationship between these MT structures and the signal transduction mechanisms that begin after fertilization, including Ca2+ signaling, MPF signaling, and MEK/MAPK signaling. Especially, CAMP formation was directed by activities of cyclin-dependent kinases. As these MT structures are conserved, at least, within chordate group, we emphasize the importance of understanding the controlling mechanisms of MT dynamics, which is important for embryonic axis determination in the ascidian egg.

Microtubule array observed in the posterior-vegetal cortex during cytoplasmic and cortical reorganization of the ascidian egg.

Body axis formation during embryogenesis results from asymmetric localization of maternal factors in the egg. Shortly before the first cleavage in ascidian eggs, cell polarity along the anteroposterior (A-P) axis is established and the cytoplasmic domain (myoplasm) relocates from the vegetal to the posterior region in a microtubule-dependent manner. Through immunostaining, tubulin accumulation during this reorganization is observable on the myoplasm cortex. However, more detailed morphological features of microtubules remain relatively unknown. In this study, we invented a new reagent that improves the immunostaining of cortical microtubules and successfully visualized a parallel array of thick microtubules. During reorganization, they covered nearly the entire myoplasm cortical region, beneath the posterior-vegetal cortex. We designated this microtubule array as CAMP (cortical array of microtubules in posterior vegetal region). During the late phase of reorganization, CAMP shrank and the myoplasm formed a crescent-like cytoplasmic domain. When the CAMP formation was inhibited by sodium azide, myoplasmic reorganization and A-P axis formation were both abolished, suggesting that CAMP is important for these two processes.

Disinfection of fertilized eggs of the edible ascidian Halocynthia roretzi for prevention of soft tunic syndrome.

Azumiobodo hoyamushi, the causative agent of soft tunic syndrome, was likely introduced to farming sites of the edible ascidian Halocynthia roretzi via ascidian spat. The source of infection is thought to be cysts of A. hoyamushi that reside in the substrates on which the ascidian spat are attached, but not the spat themselves. Thus, there is a need to develop methods to prevent contamination of the substrates with A. hoyamushi during seed production of the ascidian. We evaluated the protozoacidal effects of sodium hypochlorite and povidone-iodine against the flagellate and temporary cyst forms of A. hoyamushi. Additionally, we evaluated the effects of these disinfectants on the development of fertilized ascidian eggs. The flagellate form of A. hoyamushi was completely inactivated by povidone-iodine (5 ppm, 1 min) and sodium hypochlorite (1 ppm, 1 min). The temporary cysts of A. hoyamushi were completely inactivated by both disinfectants (5 ppm, 1 min). Disinfection with 50 ppm povidone-iodine for 15 min or 5 ppm sodium hypochlorite for 15 min had no effect on ascidian embryogenesis. Thus, horizontal transmission of A. hoyamushi via the substrates can be efficiently prevented by disinfecting ascidian eggs or tools used for spawning with povidone-iodine baths ranging from 5 ppm for 1 min to 50 ppm for 15 min without any side effects.

Mitochondrial inhibitor sodium azide inhibits the reorganization of mitochondria‐rich cytoplasm and the establishment of the anteroposterior axis in ascidian embryo

In ascidian eggs, cytoplasmic and cortical reorganization, previously called ooplasmic segregation, occurs in two phases during the first cell cycle. In the second phase of reorganization, the mitochondria-rich cytoplasm (myoplasm) moves to the future posterior side, concurrent with sperm aster migration along the egg cortex. Although this reorganization is the critical step for establishing the anteroposterior axis, its molecular mechanism is not fully understood. In this study, we showed that low concentrations of the mitochondrial inhibitor sodium azide (NaN3 ), which showed the low toxicity in sperm, inhibited the second phase of reorganization without the microtubule depolymerization. In the NaN3 -treated embryo, the sperm aster was not attracted to the cortex and altered its migration pathway; therefore, the myoplasm remained at the vegetal pole. Consequently, the anteroposterior axis was not established. Another mitochondrial inhibitor, oligomycin, did not affect these processes. These results suggest that NaN3 inhibits unknown molecules that are important for the second phase of reorganization. Identifying the target molecule of NaN3 will lead to a molecular understanding of cytoplasmic and cortical reorganization.

Release from meiotic arrest in ascidian eggs requires the activity of two phosphatases but not CaMKII

The fertilising sperm triggers a transient Ca2+ increase that releases eggs from cell cycle arrest in the vast majority of animal eggs. In vertebrate eggs, Erp1, an APC/Ccdc20 inhibitor, links release from metaphase II arrest with the Ca2+ transient and its degradation is triggered by the Ca2+-induced activation of CaMKII. By contrast, many invertebrate groups have mature eggs that arrest at metaphase I, and these species do not possess the CaMKII target Erp1 in their genomes. As a consequence, it is unknown exactly how cell cycle arrest at metaphase I is achieved and how the fertilisation Ca2+ transient overcomes the arrest in the vast majority of animal species. Using live-cell imaging with a novel cyclin reporter to study cell cycle arrest and its release in urochordate ascidians, the closest living invertebrate group to the vertebrates, we have identified a new signalling pathway for cell cycle resumption in which CaMKII plays no part. Instead, we find that the Ca2+-activated phosphatase calcineurin (CN) is required for egg activation. Moreover, we demonstrate that parthenogenetic activation of metaphase I-arrested eggs by MEK inhibition, independent of a Ca2+ increase, requires the activity of a second egg phosphatase: PP2A. Furthermore, PP2A activity, together with CN, is required for normal egg activation during fertilisation. As ascidians are a sister group of the vertebrates, we discuss these findings in relation to cell cycle arrest and egg activation in chordates.

Release from meiotic arrest in ascidian eggs requires the activity of two phosphatases but not CaMKII

The fertilising sperm triggers a transient Ca(2+) increase that releases eggs from cell cycle arrest in the vast majority of animal eggs. In vertebrate eggs, Erp1, an APC/C(cdc20) inhibitor, links release from metaphase II arrest with the Ca(2+) transient and its degradation is triggered by the Ca(2+)-induced activation of CaMKII. By contrast, many invertebrate groups have mature eggs that arrest at metaphase I, and these species do not possess the CaMKII target Erp1 in their genomes. As a consequence, it is unknown exactly how cell cycle arrest at metaphase I is achieved and how the fertilisation Ca(2+) transient overcomes the arrest in the vast majority of animal species. Using live-cell imaging with a novel cyclin reporter to study cell cycle arrest and its release in urochordate ascidians, the closest living invertebrate group to the vertebrates, we have identified a new signalling pathway for cell cycle resumption in which CaMKII plays no part. Instead, we find that the Ca(2+)-activated phosphatase calcineurin (CN) is required for egg activation. Moreover, we demonstrate that parthenogenetic activation of metaphase I-arrested eggs by MEK inhibition, independent of a Ca(2+) increase, requires the activity of a second egg phosphatase: PP2A. Furthermore, PP2A activity, together with CN, is required for normal egg activation during fertilisation. As ascidians are a sister group of the vertebrates, we discuss these findings in relation to cell cycle arrest and egg activation in chordates.

Identification of Novel Isoforms of Vitellogenin Expressed in Ascidian Eggs

Vitellogenin is a precursor of yolk protein that is necessary for embryonic development. This protein is a large multi-domain protein consisting of a signal peptide, a heavy-chain lipovitellin, a phosvitin, a light-chain lipovitellin, a von Willebrand factor type D domain (vWF-D), and a C-terminal coding region (CT), which are processed to respective domains after uptake into oocytes. It is currently believed that only lipovitellin and phosvitin domains are necessary for nutrient supply to oocytes. Thus, molecular species of vitellogenin lacking these domains are not known. Here, we show that two novel isoforms of vitellogenin, both of which possess vWF-D and CT domains but not a lipovitellin or phosvitin domain, are expressed in the gonad of the ascidian Halocynthia roretzi. In situ hybridization revealed that mRNAs of these proteins are specifically expressed in oocytes and test cells, accessory cells in the perivitelline space of ascidian eggs. Immunohistochemistry showed that these proteins are localized around the surface of test cells in immature oocytes. Immunoelectron microscopy revealed that vitellogenin associates with vesicles located beneath the vitelline coat (VC) before fertilization but that it dissociates from the VC after fertilization. These results, together with our previous results showing that vWF-D and CT domains are capable of binding to the two sperm proteases HrProacrosin and HrSpermosin, led us to propose that novel isoforms of vitellogenin, which are expressed in oocytes and test cells and released to the perivitelline space during oocyte maturation, may participate in gamete interaction upon fertilization.

Purification of mitochondrial proteins HSP60 and ATP synthase from ascidian eggs: implications for antibody specificity.

Use of antibodies is a cornerstone of biological studies and it is important to identify the recognized protein with certainty. Generally an antibody is considered specific if it labels a single band of the expected size in the tissue of interest, or has a strong affinity for the antigen produced in a heterologous system. The identity of the antibody target protein is rarely confirmed by purification and sequencing, however in many cases this may be necessary. In this study we sought to characterize the myoplasm, a mitochondria-rich domain present in eggs and segregated into tadpole muscle cells of ascidians (urochordates). The targeted proteins of two antibodies that label the myoplasm were purified using both classic immunoaffinity methods and a novel protein purification scheme based on sequential ion exchange chromatography followed by two-dimensional gel electrophoresis. Surprisingly, mass spectrometry sequencing revealed that in both cases the proteins recognized are unrelated to the original antigens. NN18, a monoclonal antibody which was raised against porcine spinal cord and recognizes the NF-M neurofilament subunit in vertebrates, in fact labels mitochondrial ATP synthase in the ascidian embryo. PMF-C13, an antibody we raised to and purified against PmMRF, which is the MyoD homolog of the ascidian Phallusia mammillata, in fact recognizes mitochondrial HSP60. High resolution immunolabeling on whole embryos and isolated cortices demonstrates localization to the inner mitochondrial membrane for both ATP synthase and HSP60. We discuss the general implications of our results for antibody specificity and the verification methods which can be used to determine unequivocally an antibody's target.

Cytosolic subunits of ATP synthase are localized to the cortical endoplasmic reticulum‐rich domain of the ascidian egg myoplasm

Previously, we revealed that p58, one of the ascidian maternal factors, is identical to the alpha-subunit of F1-ATP synthase (ATPα), a protein complex of the inner mitochondrial membrane. In the current study, we used immunological probes for ascidian mitochondria components to show that the ascidian ATPα is ectopically localized to the cytosol. Virtually all mitochondrial components were localized to the mitochondria-rich myoplasm. However, in detail, ATP synthase subunits and the matrix proteins showed different localization patterns. At least at the crescent stage, transmission electron microscopy (TEM) distinguished the mitochondria-less, endoplasmic reticulum (ER)-rich cortical region and the mitochondria-rich internal region. ATPα was enriched in the cortical region and MnSOD was limited to the internal region. Using subcellular fractionation, although all of the mitochondria components were highly enriched in the mitochondria-enriched fraction, a considerable amount of ATPα and F1-ATP synthase beta-subunit (ATPβ) were recovered in the insoluble cytoplasmic fraction. Even under these conditions, F1-ATP synthase gamma-subunit (ATPγ) and F0-ATP synthase subunit b (ATPb) were not recovered in the insoluble cytoplasmic fraction. This result strongly supports the exomitochondrial localization of both ATPα and ATPβ. In addition, the detergent extraction of eggs supports the idea that these cytosolic ATP synthase subunits are associated with the egg cytoskeleton. These results suggest that the subunits of ATP synthase might play dual roles at different subcellular compartments during early development.

Urochordate Ascidians Possess a Single Isoform of Aurora Kinase That Localizes to the Midbody via TPX2 in Eggs and Cleavage Stage Embryos

Aurora kinases are key proteins found throughout the eukaryotes that control mitotic progression. Vertebrate Aurora-A and B kinases are thought to have evolved from a single Aurora-kinase isoform closest to that found in present day urochordates. In urochordate ascidians Aurora binds both TPX2 (a vertebrate AURKA partner) and INCENP (a vertebrate AURKB partner) and localizes to centrosomes and spindle microtubules as well as chromosomes and midbody during both meiosis and mitosis. Ascidian Aurora also displays this localization pattern during mitosis in echinoderms, strengthening the idea that non-vertebrate deuterostomes such as the urochordates and echinoderms possess a single form of Aurora kinase that has properties of vertebrate Aurora-kinase A and B. In the ascidian, TPX2 localizes to the centrosome and the spindle poles also as in vertebrates. However, we were surprised to find that TPX2 also localized strongly to the midbody in ascidian eggs and embryos. We thus examined more closely Aurora localization to the midbody by creating two separate point mutations of ascidian Aurora predicted to perturb binding to TPX2. Both forms of mutated Aurora behaved as predicted: neither localized to spindle poles where TPX2 is enriched. Interestingly, neither form of mutated Aurora localized to the midbody where TPX2 is also enriched, suggesting that ascidian Aurora midbody localization required TPX2 binding in ascidians. Functional analysis revealed that inhibition of Aurora kinase with a pharmacological inhibitor or with a dominant negative kinase dead form of Aurora caused cytokinesis failure and perturbed midbody formation during polar body extrusion. Our data support the view that vertebrate Aurora-A and B kinases evolved from a single non-vertebrate deuterostome ancestor. Moreover, since TPX2 localizes to the midbody in ascidian eggs and cleavage stage embryos it may be worthwhile re-assessing whether Aurora A kinase or TPX2 localize to the midbody in eggs and cleavage stage embryos.

Role of Mos/MEK/ERK cascade and Cdk1 in Ca2+ oscillations in fertilized ascidian eggs

Intracellular calcium ion concentration ([Ca2+]i) transients are observed in the fertilized eggs of all species investigated so far, and are critical for initiating several events related to egg activation and cell cycle control. Here, we investigated the role of the Mos/MEK/ERK cascade and Cdk1 on Ca2+ oscillations in fertilized ascidian eggs. The egg of the ascidian Phallusia nigra shows [Ca2+]i oscillations after fertilization: Ca2+ waves immediately following fertilization (phase I), and [Ca2+]i oscillations between the first and second polar body extrusions (phase II). Our results show that in P. nigra eggs, ERK activity peaked just before the extrusion of the first polar body, and decreased gradually, eventually disappearing at the extrusion of the second polar body. Cyclin-dependent protein kinase 1(Cdk1) activity decreased to undetectable levels immediately after fertilization, and then periodically increased according to the meiotic and mitotic cell cycle. When the unfertilized eggs were incubated with U0126, an inhibitor of MEK, before insemination, ERK was immediately inactivated, and the phase II [Ca2+]i oscillations disappeared. Alternatively, when the constitutively active Mos protein (GST-Mos) was injected into the unfertilized eggs, ERK activity was preserved for at least 120min after fertilization, and the phase II [Ca2+]i oscillations lasted for more than120min after the second polar body extrusion. These results suggest that ERK activity is necessary for maintaining [Ca2+]i oscillations. GST-ΔN85-cyclin, which maintains Cdk1 activity, caused ERK activity in the eggs to persist for over 120min after fertilization, and prolonged [Ca2+]i oscillations. Moreover, the effects of GST-ΔN85-cyclin on the egg were abrogated by the application of U0126. Thus, Cdk1-mediated [Ca2+]i oscillations seem to require ERK activity. However, GST-Mos triggered [Ca2+]i oscillations after the second polar body extrusion, whereas GST-ΔN85-cyclin did not, although it prolongs the duration of [Ca2+]i oscillations. Interestingly, GST-ΔN85-cyclin increased the frequency of [Ca2+]i transients in the Mos-induced [Ca2+]i oscillations after the extrusion of the second polar body. Thus, Cdk1 could maintain, but not activate, ERK and [Ca2+]i oscillations. ERK activity and [Ca2+]i oscillations seem to form a negative feedback loop which may be responsible for maintaining the meiotic period.

Cytoplasmic localization and reorganization in ascidian eggs: role of postplasmic/PEM RNAs in axis formation and fate determination

Localization of maternal molecules in eggs and embryos and cytoplasmic movements to relocalize them are fundamental for the orderly cellular and genetic processes during early embryogenesis. Ascidian embryos have been known as 'mosaic eggs' because of their autonomous differentiation abilities based on localized cell fate determinants. This review gives a historical overview of the concept of cytoplasmic localization, and then explains the key features such as ooplasmic movements and cell lineages that are essential to grasp the process of ascidian development mediated by localized determinant activities. These activities are partly executed by localized molecules named postplasmic/PEM RNAs, originating from approximately 50 genes, of which the muscle determinant, macho-1, is an example. The cortical domain containing these RNAs is relocalized to the posterior-vegetal region of the egg by cytoskeletal movements after fertilization, and plays crucial roles in axis formation and cell fate determination. The cortical domain contains endoplasmic reticulum and characteristic granules, and gives rise to a subcellular structure called the centrosome-attracting body (CAB), in which postplasmic/PEM RNAs are highly concentrated. The CAB is responsible for a series of unequal partitionings of the posterior-vegetal cytoplasmic domain and the postplasmic/PEM RNAs at the posterior pole during cleavage. Some components of this domain, which is rich in granules, are eventually inherited by prospective germline cells with particular postplasmic/PEM RNAs such as vasa. The postplasmic/PEM RNAs are classified into two groups according to their final cellular destinations and localization pathways. Localization of these RNAs is regulated by specific nucleotide sequences in the 3' untranslated regions (3'UTRs).

The maternal muscle determinant in the ascidian egg

Muscle formation in ascidian embryos has been investigated for more than a century as a representative example of cell fate specification by localized maternal factors within the egg cytoplasm. Observations of colored cytoplasm in combination with micromanipulation techniques have suggested the presence of a muscle-forming factor. The molecular basis has been elucidated with the discovery of macho-1. macho-1 mRNA is already present in the unfertilized egg, and translocates to the posterior region of the egg during ooplasmic movements. It encodes a zinc-finger transcription factor that positively regulates the expression of target genes. macho-1-binding cis-elements have been identified in muscle-specific zygotic genes. Maternally localized macho-1 appears to have originated in the ascidian lineage, but it activates a muscle-forming developmental program that is shared by the vertebrates. macho-1 is also involved in establishment of the anterior-posterior axis as a competence factor in mesenchyme induction in the posterior region. It is suggested that translation of the macho-1 protein is initiated at the eight-cell stage, and that the protein is inherited by all descendant blastomeres of the posterior-vegetal region. The macho-1 activities in nonmuscle descendants are suppressed or modified by cell interactions during the cleavage stages. In addition to the primary muscle specified by maternal macho-1, ascidian embryos develop secondary muscle, whose fate is determined by cell interactions. Dozens of maternal mRNAs show similar localization to macho-1, and these are known as postplasmic/PEM RNAs, being also involved in various posterior-specific developmental events. Evolutionary aspects relevant to macho-1 and tail muscle formation are also discussed in this article.

Valosin‐containing protein/p97 interacts with sperm‐activating and sperm‐attracting factor (SAAF) in the ascidian egg and modulates sperm‐attracting activity

Sperm chemotaxis toward an egg is observed in many animals, and the control of sperm-attracting activity is thought to play an important role in ensuring fertilization. However, the mechanism underlying the release of a sperm attractant from an egg is still obscure. In this study, we examined the systems involved in the release of sperm-activating and sperm-attracting factor (SAAF), which is the sperm attractant of the ascidian Ciona intestinalis. Here, we show that the egg acquires sperm-attracting activity after germinal vesicle breakdown. Further, since the cytoplasmic extracts of immature oocytes exhibit no sperm-attracting activity, the SAAF in oocytes may be activated after germinal vesicle breakdown. We found 13 SAAF-binding proteins in an egg plasma membrane extract and identified five proteins by proteomic analysis: valosin-containing protein (VCP)/p97, proteasome alpha 2 subunit, MGC97756 protein, proteasome subunit Y, and beta-tubulin. In particular, the interaction between VCP/p97 and SAAF was confirmed by a pull-down assay. VCP/p97 is initially localized in the germinal vesicle, and during oocyte maturation, it shifts to the endoplasmic reticulum in the cortical regions. Thus, VCP/p97 is a potential modulator of SAAF release from the egg.

Valosin Containing Protein VCP/p97 Binds to the Sperm Attractant SAAF and May Control Its Release from the Ascidian Egg.

Valosin Containing Protein VCP/p97 Binds to the Sperm Attractant SAAF and May Control Its Release from the Ascidian Egg.

The ascidian egg envelope in fertilization: structural and molecular features

In this report, unpublished and recent findings concerning the structure and function of the ascidian egg coat are compiled in context with fertilization. In the initial stage of ascidian fertilization, sperm interact with a complex egg investment that consists of a layer of follicle cells attached to an acellular vitelline coat. Increasing evidence exists that ascidian sperm are activated at their encounter with the follicle cells. The molecular basis of sperm-follicle cell interactions is discussed in context with sperm binding, membrane proteins and sperm bound glycosidase. The model that suggests a block to polyspermy established by glycosidase released from the follicle cells on fertilization is evaluated and compared with assured facts. Although a number of questions remain to be answered, our recent findings that a cloned beta-hexosaminidase from P. mammillata binds exclusively to the follicle cells of unfertilized but not fertilized eggs, indicates that the follicle cells participate in the block to polyspermy. A dual function, mediating sperm activation and a block to polyspermy attributes to the ascidian follicle cells a key position in fertilization.

Toward a new paradigm for studying ascidian developmental dynamics

Toward a new paradigm for studying ascidian developmental dynamics