Calcium signaling differentiation during Xenopus oocyte maturation

Ca(2+) is the universal signal for egg activation at fertilization in all sexually reproducing species. The Ca(2+) signal at fertilization is necessary for egg activation and exhibits specialized spatial and temporal dynamics. Eggs acquire the ability to produce the fertilization-specific Ca(2+) signal during oocyte maturation. However, the mechanisms regulating Ca(2+) signaling differentiation during oocyte maturation remain largely unknown. At fertilization, Xenopus eggs produce a cytoplasmic Ca(2+) (Ca(2+)(cyt)) rise that lasts for several minutes, and is required for egg activation. Here, we show that during oocyte maturation Ca(2+) transport effectors are tightly modulated. The plasma membrane Ca(2+) ATPase (PMCA) is completely internalized during maturation, and is therefore unable to extrude Ca(2+) out of the cell. Furthermore, IP(3)-dependent Ca(2+) release is required for the sustained Ca(2+)(cyt) rise in eggs, showing that Ca(2+) that is pumped into the ER leaks back out through IP(3) receptors. This apparent futile cycle allows eggs to maintain elevated cytoplasmic Ca(2+) despite the limited available Ca(2+) in intracellular stores. Therefore, Ca(2+) signaling differentiates in a highly orchestrated fashion during Xenopus oocyte maturation endowing the egg with the capacity to produce a sustained Ca(2+)(cyt) transient at fertilization, which defines the egg's competence to activate and initiate embryonic development.     

Modeling Ca2+ signaling differentiation during oocyte maturation

Ca2+ is a fundamental intracellular signal that mediates a variety of disparate physiological functions often in the same cell. Ca2+ signals span a wide range of spatial and temporal scales, which endow them with the specificity required to induce defined cellular functions. Furthermore, Ca2+ signaling is highly plastic as it is modulated dynamically during normal physiological development and under pathological conditions. However, the molecular mechanisms underlying Ca2+ signaling differentiation during cellular development remain poorly understood. Oocyte maturation in preparation for fertilization provides an exceptionally well-suited model to elucidate Ca2+ signaling regulation during cellular development. This is because a Ca2+ signal with specialized spatial and temporal dynamics is universally essential for egg activation at fertilization. Here we use mathematical modeling to define the critical determinants of Ca2+ signaling differentiation during oocyte maturation. We show that increasing IP3 receptor (IP3R) affinity replicates both elementary and global Ca2+ dynamics observed experimentally following oocyte maturation. Furthermore, our model reveals that because of the Ca2+ dependency of both SERCA and the IP3R, increased IP3R affinity shifts the system's equilibrium to a new steady state of high cytosolic Ca2+, which is essential for fertilization. Therefore our model provides unique insights into how relatively small alterations of the basic molecular mechanisms of Ca2+ signaling components can lead to dramatic alterations in the spatio-temporal properties of Ca2+ dynamics.     

Increased sensitivity and clustering of elementary Ca2+ release events during oocyte maturation

The universal signal for egg activation at fertilization is a rise in cytoplasmic Ca(2+) with defined spatial and temporal kinetics. Mammalian and amphibian eggs acquire the ability to produce such Ca(2+) signals during a maturation period that precedes fertilization and encompasses resumption of meiosis and progression to metaphase II. In Xenopus, immature oocytes produce fast, saltatory Ca(2+) waves that can be oscillatory in nature in response to IP(3). In contrast, mature eggs produce a single continuous, sweeping Ca(2+) wave in response to IP(3) or sperm fusion. The mechanisms mediating the differentiation of Ca(2+) signaling during oocyte maturation are not well understood. Here, I characterized elementary Ca(2+) release events (Ca(2+) puffs) in oocytes and eggs and show that the sensitivity of IP(3)-dependent Ca(2+) release is greatly enhanced during oocyte maturation. Furthermore, Ca(2+) puffs in eggs have a larger spatial fingerprint, yet are short lived compared to oocyte puffs. Most interestingly, Ca(2+) puffs cluster during oocyte maturation resulting in a continuum of Ca(2+) release sites over space in eggs. These changes in the spatial distribution of elementary Ca(2+) release events during oocyte maturation explain the continuous nature and slower speed of the fertilization Ca(2+) wave.     

Internalization of plasma membrane Ca2+-ATPase during Xenopus oocyte maturation

A transient increase in intracellular Ca²⁺ is the universal signal for egg activation at fertilization. Eggs acquire the ability to mount the specialized fertilization-specific Ca²⁺ signal during oocyte maturation. The first Ca²⁺ transient following sperm entry in vertebrate eggs has a slow rising phase followed by a sustained plateau. The molecular determinants of the sustained plateau are poorly understood. We have recently shown that a critical determinant of Ca²⁺ signaling differentiation during oocyte maturation is internalization of the plasma membrane calcium ATPase (PMCA). PMCA internalization is representative of endocytosis of several integral membrane proteins during oocyte maturation, a requisite process for early embryogenesis. Here we investigate the mechanisms regulating PMCA internalization. To track PMCA trafficking in live cells we cloned a full-length cDNA of Xenopus PMCA1, and show that GFP-tagged PMCA traffics in a similar fashion to endogenous PMCA. Functional data show that MPF activation during oocyte maturation is required for full PMCA internalization. Pharmacological and co-localization studies argue that PMCA is internalized through a lipid raft endocytic pathway. Deletion analysis reveal a requirement for the N-terminal cytoplasmic domain for efficient internalization. Together these studies define the mechanistic requirements for PMCA internalization during oocyte maturation.     

Modeling Ca2+ signaling differentiation during oocyte maturation

Ca²⁺ is a fundamental intracellular signal that mediates a variety of disparate physiological functions often in the same cell. Ca²⁺ signals span a wide range of spatial and temporal scales, which endow them with the specificity required to induce defined cellular functions. Furthermore, Ca²⁺ signaling is highly plastic as it is modulated dynamically during normal physiological development and under pathological conditions. However, the molecular mechanisms underlying Ca²⁺ signaling differentiation during cellular development remain poorly understood. Oocyte maturation in preparation for fertilization provides an exceptionally well-suited model to elucidate Ca²⁺ signaling regulation during cellular development. This is because a Ca²⁺ signal with specialized spatial and temporal dynamics is universally essential for egg activation at fertilization. Here we use mathematical modeling to define the critical determinants of Ca²⁺ signaling differentiation during oocyte maturation. We show that increasing IP₃ receptor (IP₃R) affinity replicates both elementary and global Ca²⁺ dynamics observed experimentally following oocyte maturation. Furthermore, our model reveals that because of the Ca²⁺ dependency of both SERCA and the IP₃R, increased IP₃R affinity shifts the system's equilibrium to a new steady state of high cytosolic Ca²⁺, which is essential for fertilization. Therefore our model provides unique insights into how relatively small alterations of the basic molecular mechanisms of Ca²⁺ signaling components can lead to dramatic alterations in the spatio-temporal properties of Ca²⁺ dynamics.     

Ca2+ homeostasis regulates Xenopus oocyte maturation

In contrast to the well-defined role of Ca2+ signals during mitosis, the contribution of Ca2+ signaling to meiosis progression is controversial, despite several decades of investigating the role of Ca2+ and its effectors in vertebrate oocyte maturation. We have previously shown that during Xenopus oocyte maturation, Ca2+ signals are dispensable for entry into meiosis and for germinal vesicle breakdown. However, normal Ca2+ homeostasis is essential for completion of meiosis I and extrusion of the first polar body. In this study, we test the contribution of several downstream effectors in mediating the Ca2+ effects during oocyte maturation. We show that calmodulin and calcium-calmodulin-dependent protein kinase II (CAMK2) are not critical downstream Ca2+ effectors during meiotic maturation. In contrast, accumulation of Aurora kinase A (AURKA) protein is disrupted in cells deprived of Ca2+ signals. Since AURKA is required for bipolar spindle formation, failure to accumulate AURKA may contribute to the defective spindle phenotype following Ca2+ deprivation. These findings argue that Ca2+ homeostasis is important in establishing the oocyte's competence to undergo maturation in preparation for fertilization and embryonic development.     

MAP kinases regulate unfertilized egg apoptosis and fertilization suppresses death via Ca2+ signaling

The default fate for eggs from many species is death by apoptosis and thus, successful fertilization depends upon suppression of the maternal death program. Little is known about the molecular triggers which activate this process or how the fertilization signal suppresses the default maternal apoptotic pathway. The MAP kinase (MAPK) family member, ERK, plays a universal and critical role in several stages of oocyte meiotic maturation, and fertilization results in ERK inactivation. In somatic cells, ERK and other MAPK family members, p38 and JNK, provide opposing signals to regulate apoptosis, however, it is not known whether MAPKs play a regulatory role in egg apoptosis, nor whether suppression of apoptosis by fertilization is mediated by MAPK activity. Here we demonstrate that MAPKs are involved in starfish egg apoptosis and we investigate the relationship between the fertilization induced signaling pathway and MAPK activation. ERK is active in post-meiotic eggs just until apoptosis onset and then p38, JNK and a third kinase are activated, and remain active through execution. Sequential activation of ERK and p38 is necessary for apoptosis, and newly synthesized proteins are required both upstream of ERK and downstream of p38 for activation of the full apoptotic program. Fertilization causes a dramatic rise in intracellular Ca2+, and we report that Ca2+ provides a necessary and sufficient pro-survival signal. The Ca2+ pathway following fertilization of both young and aged eggs causes ERK to be rapidly inactivated, but fertilization cannot rescue aged eggs from death, indicating that ERK inactivation is not sufficient to suppress apoptosis.     

Developmental control of oocyte maturation and egg activation in metazoan models

Production of functional eggs requires meiosis to be coordinated with developmental signals. Oocytes arrest in prophase I to permit oocyte differentiation, and in most animals, a second meiotic arrest links completion of meiosis to fertilization. Comparison of oocyte maturation and egg activation between mammals, Caenorhabditis elegans, and Drosophila reveal conserved signaling pathways and regulatory mechanisms as well as unique adaptations for reproductive strategies. Recent studies in mammals and C. elegans show the role of signaling between surrounding somatic cells and the oocyte in maintaining the prophase I arrest and controlling maturation. Proteins that regulate levels of active Cdk1/cyclin B during prophase I arrest have been identified in Drosophila. Protein kinases play crucial roles in the transition from meiosis in the oocyte to mitotic embryonic divisions in C. elegans and Drosophila. Here we will contrast the regulation of key meiotic events in oocytes.     

Induction of maturation-promoting factor during Xenopus oocyte maturation uncouples Ca(2+) store depletion from store-operated Ca(2+) entry.

During oocyte maturation, eggs acquire the ability to generate specialized Ca(2+) signals in response to sperm entry. Such Ca(2+) signals are crucial for egg activation and the initiation of embryonic development. We examined the regulation during Xenopus oocyte maturation of store-operated Ca(2+) entry (SOCE), an important Ca(2+) influx pathway in oocytes and other nonexcitable cells. We have previously shown that SOCE inactivates during Xenopus oocyte meiosis. SOCE inactivation may be important in preventing premature egg activation. In this study, we investigated the correlation between SOCE inactivation and the Mos-mitogen-activated protein kinase (MAPK)-maturation-promoting factor (MPF) kinase cascade, which drives Xenopus oocyte maturation. SOCE inactivation at germinal vesicle breakdown coincides with an increase in the levels of MAPK and MPF. By differentially inducing Mos, MAPK, and MPF, we demonstrate that the activation of MPF is necessary for SOCE inactivation during oocyte maturation. In contrast, sustained high levels of Mos kinase and the MAPK cascade have no effect on SOCE activation. We further show that preactivated SOCE is not inactivated by MPF, suggesting that MPF does not block Ca(2+) influx through SOCE channels, but rather inhibits coupling between store depletion and SOCE activation.     

钙信号在卵母细胞激活中的作用

钙信号是胞内主要的第二信使之一,发挥广泛的作用如细胞分裂、细胞凋亡等,对细胞的生命活动起着非常重要的作用。在精子和卵母细胞中,钙信号对精子获能、顶体反应、卵母细胞成熟、受精及卵裂等一系列复杂的过程有非常重要的影响。现就Ca2 在卵母细胞中的释放机制、信号转导途径、调控功能作一综述。     

Identification of a starfish egg PLC-gamma that regulates Ca2+ release at fertilization

At fertilization, eggs undergo a cytoplasmic free Ca2+ rise, which is necessary for stimulating embryogenesis. In starfish eggs, studies using inhibitors designed against vertebrate proteins have shown that this Ca2+ rise requires an egg Src family kinase (SFK) that directly or indirectly activates phospholipase C-gamma (PLC-gamma) to produce IP3, which triggers Ca2+ release from the egg's endoplasmic reticulum (ER) [reviewed in Semin. Cell Dev. Biol. 12 (2001) 45]. To examine in more detail the endogenous factors in starfish eggs that are required for Ca2+ release at fertilization, an oocyte cDNA encoding PLC-gamma was isolated from the starfish Asterina miniata. This cDNA, designated AmPLC-gamma, encodes a protein with 49% identity to mammalian PLC-gamma1. A 58-kDa Src family kinase interacted with recombinant AmPLC-gamma Src homology 2 (SH2) domains in a specific, fertilization-responsive manner. Immunoprecipitations of sea urchin egg PLC-gamma using an affinity-purified antibody directed against AmPLC-gamma revealed fertilization-dependent phosphorylation of PLC-gamma. Injecting starfish eggs with the tandem SH2 domains of AmPLC-gamma (which inhibits PLC-gamma activation) specifically inhibited Ca2+ release at fertilization. These results indicate that an endogenous starfish egg PLC-gamma interacts with an egg SFK and mediates Ca2+ release at fertilization via a PLC-gamma SH2 domain-mediated mechanism.     

Calcium and other ion dynamics during gamete maturation and fertilization

Ion currents and cytosolic free calcium ([Ca(2+)](i)) elevations are crucial events in triggering the complex machinery involved in both gamete maturation and fertilization. Oocyte maturation is triggered by hormone signaling which causes ion currents and [Ca(2+)](i) increase. Extracellular calcium seems to be required for meiosis progression since: (i) calcium depletion in the maturation medium severely affects oocyte developmental competence; (ii) the activity of plasma membrane L-type Ca(2+) currents decreases during maturation; (iii) the exposure to verapamil, a specific Ca(2+) channel blocker, decreases in vitro maturation efficiency. In spermatozoa, maturation initiates inside the epididymis and ends in the female genital tract. During their journey through the female reproductive tract, sperm undergo a dramatic selection and capacitation achieving fertilization competence. Adhesion to the tubal epithelium extends sperm life through depression of [Ca(2+)](i) until capacitation signals trigger an [Ca(2+)](i) elevation followed by sperm release. At fertilization, egg-sperm interaction evokes well-described transient and almost simultaneous events: i.e., fertilization current, a change in resting potential, and an increase in free [Ca(2+)](i) concentration. These events, termed oocyte activation, are the direct consequence of sperm interaction via either activation of a receptor or entry of a sperm factor. The latter hypothesis has been recently supported by the discovery of PCLzeta, a sperm-specific isozyme triggering a dramatic [Ca(2+)](i) increase via inositol 1,4,5-trisphosphate (IP(3)) production. The course of ion currents and [Ca(2+)](i) transients during maturation and fertilization plays a pivotal role in correct embryo development.     

NAADP and InsP3 play distinct roles at fertilization in starfish oocytes

NAADP participates in the response of starfish oocytes to sperm by triggering the fertilization potential (FP) through the activation of a Ca2+ current which depolarizes the membrane to the threshold of activation of the voltage-gated Ca2+ channels. The aim of this study was to investigate whether this Ca2+ influx is linked to the onset of the concomitant InsP3-mediated Ca2+ wave by simultaneously employing Ca2+ imaging and single-electrode intracellular recording techniques. In control oocytes, the sperm-induced membrane depolarization always preceded by a few seconds the onset of the Ca2+ wave. Strikingly, the self-desensitization of NAADP receptors either abolished the Ca2+ response or resulted in abnormal oocyte activation, i.e., the membrane depolarization followed the Ca2+ wave and the oocyte was polyspermic. The inhibition of InsP3 signaling only impaired the propagation of the Ca2+ wave and shortened the FP. The duration of FP was also reduced in low-Na+ sea water. Finally, uncaged InsP3 produced a Ca2+ increase, which depolarized the membrane upon the activation of a Ca2+-sensitive cation current. These results support the hypothesis that Ca2+ entry during the NAADP-triggered FP is required for the onset of the Ca2+ wave at fertilization. The InsP3-mediated Ca2+ wave, in turn, may interact with the NAADP-evoked depolarization by activating a Ca2+-dependent Na+ entry.     

The MEK inhibitor, U0126, alters fertilization-induced [Ca2+]i oscillation parameters and secretion: differential effects associated with in vivo and in vitro meiotic maturation

Although mitogen-activated protein kinase (MAPK) is a well-known cell cycle regulator, emerging studies have also implicated its activity in the regulation of intracellular calcium concentration ([Ca2+](i)) and secretion. Those studies raise the hypothesis that MAPK activity during oocyte maturation and early fertilization is required for normal egg Ca2+ oscillations and cortical granule (CG) secretion. We extend the findings of [Lee, B., Vermassen, E., Yoon, S.-Y., Vanderheyden, V., Ito, J., Alfandari, D., De Smedt, H., Parys, J.B., Fissore, R.A., 2006. Phosphorylation of IP(3)R1 and the regulation of [Ca2+](i) responses at fertilization: a role for the MAP kinase pathway. Development 133, 4355-4365] by demonstrating acute effects on Ca2+ oscillation frequency, amplitude, and duration in fertilized mouse eggs matured in vitro with the MAPK inhibitor, U0126. Frequency was increased, whereas amplitude and duration were greatly decreased. These effects were significantly reduced in eggs matured in vivo and fertilized in the presence of the inhibitor. Ionomycin studies indicated that intracellular Ca2+ stores were differentially affected in eggs matured in vitro with U0126. Consistent with these effects on [Ca2+](i) elevation, fertilization-induced CG exocytosis and metaphase II exit were also reduced in in vitro-matured eggs with U0126, but not in those similarly treated after in vivo maturation. These results indicate that MAPK targets Ca2+ regulatory proteins during both maturation and fertilization, as well as provide a new hypothesis for MAPK function, which is to indirectly regulate events of early development by controlling Ca2+ oscillation parameters.     

Eph and NMDA receptors control Ca2+/calmodulin-dependent protein kinase II activation during C. elegans oocyte meiotic maturation

Fertilization in the female reproductive tract depends on intercellular signaling mechanisms that coordinate sperm presence with oocyte meiotic progression. To achieve this coordination in Caenorhabditis elegans, sperm release an extracellular signal, the major sperm protein (MSP), to induce oocyte meiotic maturation and ovulation. MSP binds to multiple receptors, including the VAB-1 Eph receptor protein-tyrosine kinase on oocyte and ovarian sheath cell surfaces. Canonical VAB-1 ligands called ephrins negatively regulate oocyte maturation and MPK-1 mitogen-activated protein kinase (MAPK) activation. Here, we show that MSP and VAB-1 regulate the signaling properties of two Ca2+ channels that are encoded by the NMR-1 N-methyl D-aspartate type glutamate receptor subunit and ITR-1 inositol 1,4,5-triphosphate receptor. Ephrin/VAB-1 signaling acts upstream of ITR-1 to inhibit meiotic resumption, while NMR-1 prevents signaling by the UNC-43 Ca2+/calmodulin-dependent protein kinase II (CaMKII). MSP binding to VAB-1 stimulates NMR-1-dependent UNC-43 activation, and UNC-43 acts redundantly in oocytes to promote oocyte maturation and MAPK activation. Our results support a model in which VAB-1 switches from a negative regulator into a redundant positive regulator of oocyte maturation upon binding to MSP. NMR-1 mediates this switch by controlling UNC-43 CaMKII activation at the oocyte cortex.     

Parthenogenetic activation of porcine oocytes after nuclear transfer

Mature porcine oocytes are arrested at metaphase II of meiosis. At fertilization, like all mammalian oocytes they exhibit a low frequency Ca(2+) oscillation lasting several hours. This oscillation is thought to be the signal that triggers resumption of meiosis and activates the developmental program of the oocyte. The signal transduction mechanism of the sperm-induced Ca(2+) signal is not known in detail, and attempts to generate the oscillation artificially have met with little success. Nevertheless, artificial activation of the oocyte is a crucial step during nuclear transfer. Methods are available to induce a transient elevation in the intracellular free Ca(2+) concentration to surpass the meiotic arrest and induce development of the constructed embryo. Further studies concentrating on the mechanism of Ca(2+) signaling during fertilization will help to improve the efficiency of the procedures used for parthenogenetic activation of the oocyte.     

Phosphorylation of IP3R1 and the regulation of [Ca2+]i responses at fertilization: a role for the MAP kinase pathway.

A sperm-induced intracellular Ca2+ signal ([Ca2+]i) underlies the initiation of embryo development in most species studied to date. The inositol 1,4,5 trisphosphate receptor type 1 (IP3R1) in mammals, or its homologue in other species, is thought to mediate the majority of this Ca2+ release. IP3R1-mediated Ca2+ release is regulated during oocyte maturation such that it reaches maximal effectiveness at the time of fertilization, which, in mammalian eggs, occurs at the metaphase stage of the second meiosis (MII). Consistent with this, the [Ca2+]i oscillations associated with fertilization in these species occur most prominently during the MII stage. In this study, we have examined the molecular underpinnings of IP3R1 function in eggs. Using mouse and Xenopus eggs, we show that IP3R1 is phosphorylated during both maturation and the first cell cycle at a MPM2-detectable epitope(s), which is known to be a target of kinases controlling the cell cycle. In vitro phosphorylation studies reveal that MAPK/ERK2, one of the M-phase kinases, phosphorylates IP3R1 at at least one highly conserved site, and that its mutation abrogates IP3R1 phosphorylation in this domain. Our studies also found that activation of the MAPK/ERK pathway is required for the IP3R1 MPM2 reactivity observed in mouse eggs, and that eggs deprived of the MAPK/ERK pathway during maturation fail to mount normal [Ca2+]i oscillations in response to agonists and show compromised IP3R1 function. These findings identify IP3R1 phosphorylation by M-phase kinases as a regulatory mechanism of IP3R1 function in eggs that serves to optimize [Ca2+]i release at fertilization.     

A functional genomic and proteomic perspective of sea urchin calcium signaling and egg activation

The sea urchin egg has a rich history of contributions to our understanding of fundamental questions of egg activation at fertilization. Within seconds of sperm-egg interaction, calcium is released from the egg endoplasmic reticulum, launching the zygote into the mitotic cell cycle and the developmental program. The sequence of the Strongylocentrotus purpuratus genome offers unique opportunities to apply functional genomic and proteomic approaches to investigate the repertoire and regulation of Ca(2+) signaling and homeostasis modules present in the egg and zygote. The sea urchin "calcium toolkit" as predicted by the genome is described. Emphasis is on the Ca(2+) signaling modules operating during egg activation, but the Ca(2+) signaling repertoire has ramifications for later developmental events and adult physiology as well. Presented here are the mechanisms that control the initial release of Ca(2+) at fertilization and additional signaling components predicted by the genome and found to be expressed and operating in eggs at fertilization. The initial release of Ca(2+) serves to coordinate egg activation, which is largely a phenomenon of post-translational modifications, especially dynamic protein phosphorylation. Functional proteomics can now be used to identify the phosphoproteome in general and specific kinase targets in particular. This approach is described along with findings to date. Key outstanding questions regarding the activation of the developmental program are framed in the context of what has been learned from the genome and how this knowledge can be applied to functional studies.     

Inositol trisphosphate, inositol phospholipid metabolism, and germinal vesicle breakdown in surf clam oocytes

Others have reported that microinjection of inositol 1,4,5-trisphosphate (InsP3) releases stored intracellular Ca2+ and causes fertilization envelope elevation, part of the activation process normally initiated by fertilization in deuterostome eggs. In the protostome, Spisula solidissima, germinal vesicle breakdown (GVBD) is the first visible response of the egg to fertilization. To test the effects of InsP3 on egg activation in this organism, we microinjected the compound into oocytes. Microinjection of 0.4-7.0 x 10(-21) moles of InsP3 (equivalent to 5-80 pM if distributed throughout the cell) elicited GVBD in a dose-dependent manner, demonstrating that increased oocyte InsP3 can mimic part of the activation process in this protostome. Synthesis of InsP3 occurs in vivo when phosphatidylinositol 4,5-bisphosphate (PtdInsP2) is hydrolyzed by phospholipase C. To determine whether stimulus-induced synthesis of InsP3 occurs after fertilization of Spisula oocytes, we labeled oocyte lipids with [32P]orthophosphate and measured the radioactivity in phospholipids after insemination. Fertilization resulted in a rapid, transient loss of radioactivity from PtdInsP2. Because the radioactivity in phosphatidylinositol 4-phosphate and other phospholipids did not change, the loss of radioactivity from PtdInsP2 is most likely due to its hydrolysis, yielding InsP3 and diacylglycerol. The latter compound activates protein kinase C which has also been shown to be involved in regulating Spisula oocyte GVBD. Since both of these compounds appear to be early products of fertilization, they could coordinately activate Ca2+- and protein kinase C-dependent processes involved in Spisula oocyte GVBD. These data indicate that egg activation in this protostome includes pathways similar to those found in deuterostome eggs and in other eukaryotic cells.     

Translation of incenp during oocyte maturation is required for embryonic development in Xenopus laevis

The chromosome passenger complex (CPC) consists of Aurora-B kinase and several other subunits. One of these, incenp, binds Aurora-B and regulates its kinase activity. During Xenopus oocyte maturation, incenp accumulates through translation, contributing to aurora-b activation. A previous study has demonstrated that inhibition of incenp translation during oocyte maturation diminishes aurora-b activation but does not interfere with oocyte maturation, characterized by normal maturation-specific cyclin-b phosphorylation, degradation, and resynthesis. Here we have extended these findings, showing that inhibition of incenp translation during oocyte maturation did not interfere with meiosis I or II, as indicated by the normal emission of the first polar body and metaphase II arrest, followed by the successful emission of the second polar body upon parthenogenetic egg activation. Most importantly, however, when transferred to host frogs and subsequently ovulated, the incenp-deficient eggs were fertilized but failed to undergo mitotic cleavage. Thus, translation of incenp during oocyte maturation appears to be part of oocyte cytoplasmic maturation, preparing the egg for the rapid mitosis following fertilization.