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The sea urchin Arbacia lixula coexist with Paracentrotus lividus in the Mediterranean, but the two sea urchin species are quite different from each other. Concerning the female gamete, A. lixula eggs are much darker than those of P. lividus due to the characteristic pigmentation. Upon insemination, the fertilization envelope formed by A. lixula eggs is remarkably thinner than that of P. livius eggs, which implies that the cortical organization of the eggs in the two species may be quite different. In this communication, we examined the phenotypic plasticity of A. lixula eggs in the changing osmolality. The plasma membrane, cortical actin cytoskeleton and vesicles are extensively altered in the eggs exposed to 40% seawater for 15 min. When fertilized, the Ca 2+ response in these eggs was significantly compromised and the sperm often failed to enter the eggs. Remarkably, the pattern of the Ca 2+ response was restored when these eggs were transferring back to the natural seawater before fertilization, while the actin cytoskeleton partially reverted to the original state. Nonetheless, these eggs restored in seawater failed to regain the innate sperm receptivity that allows only one sperm to enter in natural seawater. Thus, the ability to guide monospermic fertilization is lost by water entry into the eggs, and the eggs incorporated either multiple or no sperm. On the other hand, eggs briefly exposed to hypertonic seawater exhibited no evident morphological anomaly. Nonetheless, the monospermic eggs that experienced a brief exposure (15 min) to hypertonic seawater prior to fertilization in natural seawater displayed a subtly altered sperm-induced Ca 2+ response and morpho-functional anomaly around the pluteus stage. Our results suggest that A. lixula eggs attain only a limited extent of cytological plasticity, and that the osmolality shock affects the physical nature of the egg surface which in turn affects the developmental programming., (Copyright © 2021 Elsevier B.V. All rights reserved.)

The onset of fertilization in echinoderms is characterized by instantaneous increase of Ca 2+ in the egg cortex, which is called 'cortical flash', and the subsequent Ca 2+ wave. While the cortical flash is due to the ion influx through L-type Ca 2+ channels in starfish eggs, its amplitude was shown to be affected by the integrity of the egg cortex. Here, we investigated the contribution of cortical granules (CG) and yolk granules (YG) to the sperm-induced Ca 2+ signals in sea urchin eggs. To this end, prior to fertilization, Paracentrotus lividus eggs were treated with agents that disrupt or relocate CG beneath the plasma membrane: namely, glycyl-L-phenylalanine 2-naphthylamide (GPN), procaine, urethane, and NH 4 Cl. All these pretreatments consistently suppressed the cortical flash in the fertilized eggs, and accelerated the decay kinetics of the subsiding Ca 2+ wave in most cases. By contrast, centrifugation of the eggs, which stratifies organelles but not the CG, did not exhibit such changes except that the CF was much enhanced in the centrifugal pole where YG are localized. Surprisingly, we noted that pretreatment of the eggs with these CG-disrupting agents or with the inhibitors of L-type Ca 2+ channels all drastically reduced the density of the microvilli and their individual shapes on the egg surface. Taken together, our results suggest that the integrity of the egg cortex ensures successful generation of the Ca 2+ responses at fertilization, and that modulation of microvilli shape and density may serve as a mechanism of controlling ion flux across the plasma membrane., Competing Interests: Competing Interests: The authors have declared that no competing interest exists.

Starfish and sea urchin are excellent models to study the mechanisms that regulate oocyte maturation and egg activation. Hormonal stimulation of starfish oocytes and their following interaction with spermatozoa induce rapid changes of F-actin and Ca 2+ increases which are prerequisites for normal fertilization and development. Fully grown oocytes isolated from the gonads of starfish contain a large nucleus (∼60-70 μm) (termed germinal vesicle, GV), which is arrested at the first prophase of meiosis. If inseminated, these immature oocytes are penetrated by additional spermatozoa. However, starfish oocytes naturally shed into the sea have already initiated the (meiotic) maturation and are normally fertilized between GV breakdown and the extrusion of the first polar body. This is considered the optimum period to ensure monospermic instead of polyspermic fertilization. By contrast, sea urchin eggs are fertilized only after being fully matured, i.e., at the end of the two meiotic divisions. Here, we provide a comparative review of the role of the actin cytoskeleton in oocyte maturation and fertilization in starfish and sea urchin. It has become increasingly evident that the exquisite regulation of the cortical F-actin is involved in nearly all aspects of the molecular events taking place during the progression of meiotic maturation and fertilization., (Copyright © 2018 Elsevier Inc. All rights reserved.)

Background/aims: Eggs of all animal species display intense cytoplasmic Ca2+ increases at fertilization. Previously, we reported that unfertilized eggs of Astropecten aranciacus exposed to an actin drug latrunculin A (LAT-A) exhibit similar Ca2+ waves and cortical flashes after 5-10 min time lag. Here, we have explored the molecular mechanisms underlying this unique phenomenon., Methods: Starfish eggs were pretreated with various agents such as other actin drugs or inhibitors of phospholipase C (PLC), and the changes of the intracellular Ca2+ levels were monitored by use of Calcium Green in the presence or absence of LAT-A. The concomitant changes of the actin cytoskeleton were visualized with fluorescent F-actin probes in confocal microscopy., Results: We have shown that the LAT-A-induced Ca2+ increases are related to the disassembly of actin flaments: i) not only LAT-A but also other agents depolymerizing F-actin (i.e. cytochalasin B and mycalolide B) induced similar Ca2+ increases, albeit with slightly lower efficiency; ii) drugs stabilizing F-actin (i.e. phalloidin and jasplakinolide) either blocked or significantly delayed the LAT-A-induced Ca2+ increases. Further studies utilizing pharmacological inhibitors of PLC (U-73122 and neomycin), dominant negative mutant of PLC-ɣ, specific sequestration of PIP2 (RFP-PH), InsP3 uncaging, and quantitation of endogenous InsP3 all indicated that LAT-A induces Ca2+ increases by stimulating PLC rather than sensitizing InsP3 receptors. In support of the idea, it bears emphasis that LAT-A timely increased intracellular contents of InsP3 with concomitant decrease of PIP2 levels in the plasma membrane., Conclusion: Taken together, our results suggest that suboolemmal actin filaments may serve as a scaffold for cell signaling and modulate the activity of the key enzyme involved in intracellular Ca2+ signaling., (© 2018 The Author(s). Published by S. Karger AG, Basel.)

It has been known that the intracellular Ca(2+) level transiently rises at the specific stages of mitosis such as the moment of nuclear envelope breakdown and at the metaphase-anaphase transition. Comparable intracellular Ca(2+) increases may also take place during meiosis, as was intermittently reported in mouse, Xenopus, and starfish oocytes. In a majority of starfish species, the maturing oocytes display an intracellular Ca(2+) increase within few minutes after the addition of the maturation hormone, 1-methyladenine (1-MA). Although starfish oocytes at meiosis also manifest a Ca(2+) increase at the time of polar body extrusion, a similar Ca(2+) increase has never been observed during the envelope breakdown of the nucleus (germinal vesicle, GV). Here, we report, for the first time, the existence of an additional Ca(2+) response in the maturing oocytes of Asterina pectinifera at the time of GV breakdown. In contrast to the immediate early Ca(2+) response to 1-MA, which is independent of external Ca(2+) and takes a form of intracellular Ca(2+) wave traveling three times as fast as that in the fertilized eggs, this late stage Ca(2+) response comprised a train of numerous spikes representing Ca(2+) influx. These Ca(2+) spikes coinciding with GV breakdown were mostly eliminated when the GV was removed from the oocytes prior to the addition of 1-MA, suggesting that the Ca(2+) spikes are rather a consequence of the GV breakdown. In support of the idea that these Ca(2+) spikes play a physiological role, the oocytes matured in calcium-free seawater had a higher rate of cleavage failure 2h after the fertilization in natural seawater. Specific inhibitors of L-type Ca(2+) channels, verapamil and diltiazem, severely suppressed the amplitude of the individual Ca(2+) spikes, but not their frequencies. On the other hand, latrunculin-A (LAT-A), which promotes net depolymerization of the actin cytoskeleton, had a dual effect on this late Ca(2+) response. When added immediately after the hormone-dependent period, LAT-A inhibited the occurrence (frequency) of the spikes in a dose-dependent manner, but the amplitude of the prevailing Ca(2+) spikes itself was rather significantly increased. These results suggest that the cortical actin cytoskeleton and some nuclear factors may play a role in regulating ion channel activities during this stage of meiotic progression., (Copyright © 2015 Elsevier Ltd. All rights reserved.)

Sea urchin embryos initiate cell specifications at the 16-cell stage by forming the mesomeres, macromeres and micromeres according to the relative position of the cells in the animal-vegetal axis. The most vegetal cells, micromeres, autonomously differentiate into skeletons and induce the neighbouring macromere cells to become mesoendoderm in the β-catenin-dependent Wnt8 signalling pathway. Although the underlying molecular mechanism for this progression is largely unknown, we have previously reported that the initial events might be triggered by the Ca2+ influxes through the egg-originated L-type Ca2+ channels distributed asymmetrically along the animal-vegetal axis and through the stretch-dependent Ca2+channels expressed specifically in the micromere at the 4th cleavage. In this communication, we have examined whether one of the earliest Ca2+ targets, protein kinase C (PKC), plays a role in cell specification upstream of β-catenin. To this end, we surveyed the expression pattern of β-catenin in early embryos in the presence or absence of the specific peptide inhibitor of Hemicentrotus pulcherrimus PKC (HpPKC-I). Unlike previous knowledge, we have found that the initial nuclear entrance of β-catenin does not take place in the micromeres, but in the macromeres at the 16-cell stage. Using the HpPKC-I, we have demonstrated further that PKC not only determines cell-specific nucleation of β-catenin, but also regulates a variety of cell specification events in the early sea urchin embryos by modulating the cell adhesion structures, actin dynamics, intracellular Ca2+ signalling, and the expression of key transcription factors.

The interaction of the spermatozoon with the egg at fertilization remains one of the most fascinating mysteries of life. Much of our scientific knowledge on fertilization comes from studies on sea urchin and starfish, which provide plenty of gametes. Large and transparent, these eggs have served as excellent model systems for studying egg activation and embryo development in seawater, a plain natural medium. Starfish oocytes allow the study of the cortical, cytoplasmic and nuclear changes during the meiotic maturation process, which can also be triggered in vitro by hormonal stimulation. These morphological and biochemical changes ensure successful fertilization of the eggs at the first metaphase. On the other hand, sea urchin eggs are fertilized after the completion of meiosis, and are particularly suitable for the study of sperm-egg interaction, early events of egg activation, and embryonic development, as a large number of mature eggs can be fertilized synchronously. Starfish and sea urchin eggs undergo abrupt changes in the cytoskeleton and ion fluxes in response to the fertilizing spermatozoon. The plasma membrane and cortex of an egg thus represent "excitable media" that quickly respond to the stimulus with the Ca(2+) swings and structural changes. In this article, we review some of the key findings on the rapid dynamic rearrangements of the actin cytoskeleton in the oocyte/egg cortex upon hormonal or sperm stimulation and their roles in the modulation of the Ca(2+) signals and in the control of monospermic fertilization., (Copyright © 2015 Elsevier Inc. All rights reserved.)

Starfish oocytes undergo massive intracellular Ca2+ signalling during meiotic maturation and fertilization. Although the igniting stimulus of Ca2+ mobilization may differ in different cell contexts, its final leverage is usually the Ca2+-releasing second messengers such as InsP3, cADPr and NAADP. The general scheme of intracellular Ca2+ release is that the corresponding receptors for these molecules serve as ion channels to release free Ca2+ from its internal stores such as the lumen of the endoplasmic reticulum. However, a growing body of evidence has suggested that intracellular Ca2+ release can be strongly modulated by the actin cytoskeleton. Although it is known that Ca2+ contributes to remodelling of the actin cytoskeleton, whether the actin cytoskeleton modulates Ca2+ signalling in return has not been much explored. An emerging candidate to answer to this reciprocal causality of Ca2+ and the actin cytoskeleton may be actin-binding proteins. In this review, we discuss how the actin cytoskeleton may fit into the known mechanisms of intracellular Ca2+ release, and propose two models to explain the experimental data.

The explosive increase in Ca2+ that occurs in the cytosol at fertilization is brought about by the activation of Ca2+-release channels in the intracellular stores. Inositol 1,4,5-trisphosphate (InsP3) is traditionally considered to be the messenger that initiates the increase and spreading of the activating Ca2+ wave. In line with this hypothesis, recent evidence suggests that the penetrating sperm delivers into mammalian eggs a novel isoform of phospholipase C (PLC), which promotes the formation of InsP3. By contrast, data from echinoderms studies indicate that the newly discovered second messenger nicotinic adenine dinucleotide phosphate (NAADP) promotes an initial, localized increase in Ca2+, which is then followed by the InsP3-mediated globalization of the Ca2+ wave. The mechanism by which the interacting sperm triggers the production of NAADP and subsequently that of InsP3 remains obscure.

The micromeres, the first cells to be specified in sea urchin embryos, are generated by unequal cleavage at the fourth cell division. The micromeres differentiate autonomously to form spicules and dispatch signals to induce endomesoderm in the neighbouring macromeres cells in the embryo. Using a calcium indicator Fura-2/AM and a mixture of dextran conjugated Oregon green-BAPTA 488 and Rhodamine red, the intracellular calcium ion concentration ([Ca2+]i) was studied in embryos at the 16-cell stage. [Ca2+]i was characteristically elevated in the micromeres during furrowing at the 4th cleavage. Subsequently, Ca2+ oscillated for about 10 min in the micromeres, resulting in episodic high levels of [Ca2+]i. High [Ca2+]i regions were associated with regional localizations of the endoplasmic reticulum (ER), though not with ER accumulated at the vegetal pole of the micromeres during the 4th division. Pharmacological studies, using a blocker of IP3-mediated Ca2+ release (Xestospongin), a store-operated Ca2+ entry inhibitor (2 aminoethoxydiphenyl borate (2-APB)) and an inhibitor of stretch-dependent ion channels (gadolinium), suggest that the high [Ca2+]i and oscillations in the micromeres are triggered by calcium influx caused by the activation of stretch-dependent calcium channels, followed by the release of calcium ions from the endoplasmic reticulum. On the basis of these new findings, a possible mechanism for autonomous formation of the micromeres is discussed.

The resumption of the meiotic cycle (maturation) induced by 1-methyladenine in prophase-arrested starfish oocytes is indicated by the breakdown of the germinal vesicle and is characterized by the increased sensitivity of the Ca2+ stores to inositol 1,4,5-trisphosphate (InsP3) to InsP3 starting at the animal hemisphere (where the germinal vesicle was originally located) and propagating along the animal/vegetal axis of the oocyte. This initiates Ca2+ signals around the germinal vesicle before nuclear envelope breakdown. Previous studies have suggested that the final activation of the maturation-promoting factor (MPF), a cyclin-dependent kinase, which is the major element controlling the entry of eukaryotic cells into the M phase, occurs in the nucleus. MPF is then exported to the cytoplasm where its activity is autocatalytically amplified following a similar animal/vegetal spatial pattern. We have investigated whether activated MPF was involved in the increased sensitivity of the Ca2+ response to InsP3. We have found that the development of increased sensitivity of the Ca2+ stores to InsP3 receptors together with the Ca2+ signals in the perinuclear region was blocked in oocytes treated with the specific MPF inhibitor roscovitine. That the nuclear MPF activation is indeed required for changes of the InsP3 receptors sensitivity was shown by enucleating or by dissecting oocytes into vegetal and animal hemispheres prior to the addition of 1-MA. MPF activity 50 min after 1-methyladenine addition was much lower in the enucleated oocytes and in the vegetal hemisphere, which did not contain the germinal vesicle, as compared with the animal hemisphere, which did contain it. The Ca2+ increase induced by InsP3 under these experimental conditions correlated with the changes in actin cytoskeleton induced by MPF.

Cyclic ADP-ribose (cADPr) is a second messenger that regulates intracellular free [Ca2+] ([Ca2+](i)) in a variety of cell types, including immature oocytes from the starfish Astropecten auranciacus. In this study, we employed confocal laser scanning microscopy and voltage clamp techniques to investigate the source of the cADPr-elicited Ca2+ wave originating from the cortical Ca2+ patches we have described previously. The Ca2+ swing was accompanied by a membrane current with a reversal potential of approximately +20 mV. Decreasing external Na+ almost abolished the current without affecting the Ca2+ response. Removal of extracellular Ca2+ altered neither the Ca2+ transient nor the ionic current, nor did the holding potential exert any effect on the Ca2+ wave. Both the Ca2+ response and the membrane current were abolished when BAPTA, ruthenium red or 8-NH(2)-cADPr were preinjected into the oocytes, while perfusion with ADPr did not elicit any [Ca2+](i) increase or ionic current. However, elevating [Ca2+](i) by uncaging Ca2+ from nitrophenyl- (NP-EGTA) or by photoliberating inositol 1,4,5-trisphosphate (InsP(3)) induced an ionic current with biophysical properties similar to that elicited by cADPr. These results suggest that cADPr activates a Ca2+ wave by releasing Ca2+ from intracellular ryanodine receptors and that the rise in [Ca2+](i) triggers a non-selective monovalent cation current that does not seem to contribute to the global Ca2+ elevation.

We found that in starfish oocytes two different enzymes, phosphatidylserine synthase-1 (PSS1) and -2 (PSS2), which synthesize phosphatidylserine by a base-exchange reaction, are present. We studied phosphatidylserine synthesis in immature oocytes which still contain the nucleus (germinal vesicles) and in mature cells, in which the re-initiation of the meiotic cycle induced by the hormone 1-methyladenine led to structural changes in the endoplasmic reticulum, to the disappearance of the nuclear envelope and to the intermixing of the nucleoplasm with the cytoplasm. It was found that the levels of PSS1 and PSS2 transcripts were higher in immature and mature oocytes, respectively. The level of the expressed PSS2 protein, higher than that of PSS1, was not influenced by the maturation process, whereas the level of PSS1 protein was higher in immature than in mature oocytes. Serine incorporation into phosphatidylserine was enhanced in immature oocytes. The depletion of calcium stores by thapsigargin resulted in 50% lowering of phosphatidylserine synthesis. We suggest that changes in phosphatidylserine synthesis may be affected by the release of calcium stored in the nuclear envelope and in the endoplasmic reticulum, the membranes that undergo disintegration and fragmentation during meiosis. The reason for the greater synthesis of PS may be the higher level of expression of PSS1 in immature oocytes.

The effects of actin cytoskeleton disruption by cytochalasin D and latrunculin A on Ca2+ signals evoked by ADP, UTP or thapsigargin were investigated in glioma C6 cells. Despite the profound alterations of the actin cytoskeleton architecture and cell morphology, ADP and UTP still produced cytosolic calcium elevation in this cell line. However, calcium mobilization from internal stores and Ca2+ influx through store-operated Ca2+ channels induced by ADP and UTP were strongly reduced. Cytochalasin D and latrunculin A also diminished extracellular Ca2+ influx in unstimulated glioma C6 cells previously incubated in Ca2+ free buffer. In contrast, the disruption of the actin cytoskeleton had no effect on thapsigargin-induced Ca2+ influx in this cell line. Both agonist- and thapsigargin-generated Ca2+ entry was significantly decreased by the blocker of store-operated Ca2+ channels, 2-aminoethoxydiphenylborate. The data reveal that two agonists and thapsigargin activate store-operated Ca2+ channels but the mechanism of activation seems to be different. While the agonists evoke a store-mediated Ca2+ entry that is dependent on the actin cytoskeleton, thapsigargin apparently activates an additional mechanism, which is independent of the disruption of the cytoskeleton.

Cortical granules (secretory vesicles located under the cortex of mature oocytes) release their contents to the medium at fertilization. Their exocytosis modifies the extracellular environment, blocking the penetration of additional sperm. The granules translocate to the surface during the maturation process, and it has been suggested that they move to the cortex via cytoskeletal elements. In this paper we show that the increase in intracellular Ca2+, which the maturing hormone 1-methyladenine (1-MA) induces in starfish through the activation of inositol 1,4, 5-trisphosphate (InsP3) receptors, triggers changes in filamentous actin, which then direct the correct movement and reorientation of the cortical granules and the elevation of the fertilization envelope., (Copyright 1999 Academic Press.)

The dynamics of the cytoplasmic and nuclear Ca2+ pools in starfish oocytes arrested at the prophase of the first meiotic division or after induction of meiosis by 1-methyladenine (1-MA) have been studied by confocal microscopy. A 70 kDa fluorescent Ca2+ indicator has been injected in either the cytoplasm or the nucleus, and shown to remain restricted to the compartment of injection. 1-MA induced a first Ca2+ transient in the cytosol, followed by a nuclear transient, and eventually by a second cytosolic transient. The latter failed to occur if the nuclear peak was suppressed. This required the nuclear injection of antagonists of the inositol 1,4,5-trisphosphate (InsP3) and cyclic-ADPribose (cADPr) Ca2+ channels, showing that both channel types were active in the inner envelope membrane. The nuclear injection of the Ca2+ channel antagonists affected the process of meiosis reinitiation: in about one third of the injected oocytes no breakdown of the nuclear envelope (GVBD) was observed. In the others, even if GVBD eventually occurred, the intermixing of the nucleoplasm and cytoplasm was inhibited., (Copyright 1998 Academic Press.)

The regulation of cell cycle progression is a complex process which involves kinase cascades, protease action, production of second messengers and other operations. Increasing evidence now compellingly suggests that changes in the intracellular Ca2+ concentration may also have a crucial role. Ca2+ transients occur at the awakening from quiescence, at the G/S transition, during S-phase, and at the exit from mitosis. They may lead to the activation of Ca2+ binding proteins like S-100, but the key decoder of the Ca2+ signals in the cycle is calmodulin. Activation of calmodulin leads to the stimulation of protein kinases, i.e., CaM-kinase II, and of the CaM-dependent protein phosphatase calcineurin. Ample evidence now indicates the G/S transition, the progression from G2 to M, and the metaphase/anaphase transition as specific points of intervention of CaM-kinase II. Another attractive possibility for the role of Ca2+ in the cycle is through the activation of the Ca(2+)-dependent protease calpain: other proteases (e.g., the proteasome) have been suggested to be responsible for the degradation of some of cyclins, which is essential to the progression of the cycle. One of the cyclins, however, (D1) is instead degraded by calpain, which has been shown to promote both mitosis and meiosis when injected into somatic cells or oocytes.

Proteolysis is a key event in the control of the cell cycle. Most of the proteins which are degraded at specific cycle points, e.g. cyclins A, B, and E, are substrates of the ubiquitin/proteasome pathway. The Ca2+ dependent neutral protease calpain also cleaves cell cycle proteins, among them cyclin D1 and the c-mos proto-oncogene product which is a component of the CSF. The proteasome itself, however, may be under Ca2+ control through the binding of Ca2+ to its 29 kDa regulatory subunit. Calpain undergoes relocation among cell compartments during the various steps of the mitotic and meitotic cycles. It promotes the initiation and the progression of mitosis when injected into the perinuclear space of synchronized PtK1 cells, and the resumption of meiosis when directly injected into the nuclei of prophase-arrested starfish oocytes. Apart from the proteins mentioned above, most of the substrates of calpain which become cleaved during mitosis and meiosis are still unknown. Microtubule-associated proteins are likely candidates.

Regulation of Ca2+ in the nucleus is a debated issue, essentially due to the presence in the envelope of the pores, which are large enough to permit the passive traffic of small molecules like Ca2+. Work with a number of cell systems has shown that Ca2+ diffuses freely in and out of the nucleus, whereas other studies have suggested instead that the nuclear envelope could become an efficient Ca2+ filter: electrophysiological work has shown that it could become impermeable to ions, and persistent nucleus cytoplasmic Ca2+ gradients have been documented in various cell types. The problem of the control of nuclear Ca2+ thus is still open: mechanisms for gating of the pores, based on the state of depletion of the cell Ca2+ stores, have been proposed. Irrespective of the mechanisms for possible pore gating, a final picture on the traffic of Ca2+ in and out of the nucleus must also include the Ca2+ pump as well as the InsP3 and cyclic ADP ribose-modulated Ca2+ channels in the envelope. The channels can be activated by their ligands from inside the nucleus, producing Ca2+ transients in the nucleoplasm; the machinery for producing InsP3 has been documented in the envelope. Most Ca2+-sensitive nuclear functions are jointly modulated by Ca2+ and calmodulin: calmodulin-dependent kinases and the calmodulin-dependent phosphatase calcineurin have been documented in the nucleus. An interesting case for the modulation of intranuclear processes by calmodulin-dependent kinases is that of immediate early genes, i.e., CREB. Other Ca2+-modulated nuclear processes are calmodulin independent: chief among them is the intranucleosomal cleavage of chromatin and the fragmentation of nuclear proteins during apoptosis.

The treatment of prophase-arrested starfish oocytes with the hormone 1-methyladenine (1-MA) induces the elevation of Ca2+ in both the cytoplasm and the germinal vesicle (nucleus), and is followed by the resumption of meiosis. The injection of the modulators of the intracellular Ca2+ channels inositol 1,4,5-trisphosphate (InsP3) or cyclic adenosine diphosphate ribose (cADPr) into the germinal vesicle promoted meiosis resumption in the absence of 1-MA in about 50% of the oocytes. Caged InsP3 or caged cADPr were injected into the nuclei of oocytes together with the Ca2+ indicator calcium green dextran; their photoreleasing elicited nuclear calcium spikes which, in the case of cADPr, had repetitive behaviour. The spikes were abolished by the nuclear injection of antagonists or antibodies to the InsP3 or cADPr-sensitive Ca2+ channels. cADPr-modulated channels were localized on the membranes of the nuclear envelope using specific antibodies conjugated with IgG-gold complexes.

Here we report that an increase in nuclear Ca2+ is hormonally regulated and is required for reinitiation of meiosis in starfish oocytes. We have shown in Asterina pectinifera that a Ca2+ transient and a baseline increase of Ca2+ occur both in the cytoplasm and in the nucleus. These increases are in response to 1-methyladenine (1-MA), the hormone responsible for the induction of meiotic maturation. The Ca2+ transients are not necessary for the reinitiation of meiosis. However, blocking the baseline increase in nuclear Ca2+ blocks both the nuclear envelope breakdown (NEBD) and the continuation of meiosis.

Starfish oocytes undergo massive intracellular Ca2+ signalling during meiotic maturation and fertilization. Although the igniting stimulus of Ca2+ mobilization may differ in different cell contexts, its final leverage is usually the Ca2+-releasing second messengers such as InsP3, cADPr and NAADP. The general scheme of intracellular Ca2+ release is that the corresponding receptors for these molecules serve as ion channels to release free Ca2+ from its internal stores such as the lumen of the endoplasmic reticulum. However, a growing body of evidence has suggested that intracellular Ca2+ release can be strongly modulated by the actin cytoskeleton. Although it is known that Ca2+ contributes to remodelling of the actin cytoskeleton, whether the actin cytoskeleton modulates Ca2+ signalling in return has not been much explored. An emerging candidate to answer to this reciprocal causality of Ca2+ and the actin cytoskeleton may be actin-binding proteins. In this review, we discuss how the actin cytoskeleton may fit into the known mechanisms of intracellular Ca2+ release, and propose two models to explain the experimental data. [ABSTRACT FROM AUTHOR]