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Open AccessArticle

When Isolated at Full Receptivity, in Vitro Fertilized Wheat (Triticum aestivum, L.) Egg Cells Reveal [Ca²⁺]_cyt Oscillation of Intracellular Origin

by Zsolt Pónya, Ilaria Corsi, Richárd Hoffmann, Melinda Kovács, Anikó Dobosy, Attila Zoltán Kovács, Mauro Cresti and Beáta Barnabás

Int. J. Mol. Sci. 2014, 15(12), 23766-23791; https://doi.org/10.3390/ijms151223766 - 19 Dec 2014

Cited by 4 | Viewed by 6814

Abstract

During in vitro fertilization of wheat (Triticum aestivum, L.) in egg cells isolated at various developmental stages, changes in cytosolic free calcium ([Ca²⁺]_cyt) were observed. The dynamics of [Ca²⁺]_cyt elevation varied, reflecting the difference [...] Read more.

During in vitro fertilization of wheat (Triticum aestivum, L.) in egg cells isolated at various developmental stages, changes in cytosolic free calcium ([Ca²⁺]_cyt) were observed. The dynamics of [Ca²⁺]_cyt elevation varied, reflecting the difference in the developmental stage of the eggs used. [Ca²⁺]_cyt oscillation was exclusively observed in fertile, mature egg cells fused with the sperm cell. To determine how [Ca²⁺]_cyt oscillation in mature egg cells is generated, egg cells were incubated in thapsigargin, which proved to be a specific inhibitor of the endoplasmic reticulum (ER) Ca²⁺-ATPase in wheat egg cells. In unfertilized egg cells, the addition of thapsigargin caused an abrupt transient increase in [Ca²⁺]_cyt in the absence of extracellular Ca²⁺, suggesting that an influx pathway for Ca²⁺ is activated by thapsigargin. The [Ca²⁺]_cyt oscillation seemed to require the filling of an intracellular calcium store for the onset of which, calcium influx through the plasma membrane appeared essential. This was demonstrated by omitting extracellular calcium from (or adding GdCl₃ to) the fusion medium, which prevented [Ca²⁺]_cyt oscillation in mature egg cells fused with the sperm. Combined, these data permit the hypothesis that the first sperm-induced transient increase in [Ca²⁺]_cyt depletes an intracellular Ca²⁺ store, triggering an increase in plasma membrane Ca²⁺ permeability, and this enhanced Ca²⁺ influx results in [Ca²⁺]_cyt oscillation. Full article

(This article belongs to the Special Issue Cellular and Molecular Mechanisms of Sperm-Egg Interaction)

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4208 KiB

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Sperm and Spermatids Contain Different Proteins and Bind Distinct Egg Factors

by Marta Teperek, Kei Miyamoto, Angela Simeone, Renata Feret, Michael J. Deery, John B. Gurdon and Jerome Jullien

Int. J. Mol. Sci. 2014, 15(9), 16719-16740; https://doi.org/10.3390/ijms150916719 - 19 Sep 2014

Cited by 3 | Viewed by 8766

Abstract

Spermatozoa are more efficient at supporting normal embryonic development than spermatids, their immature, immediate precursors. This suggests that the sperm acquires the ability to support embryonic development during spermiogenesis (spermatid to sperm maturation). Here, using Xenopus laevis as a model organism, we performed [...] Read more.

Spermatozoa are more efficient at supporting normal embryonic development than spermatids, their immature, immediate precursors. This suggests that the sperm acquires the ability to support embryonic development during spermiogenesis (spermatid to sperm maturation). Here, using Xenopus laevis as a model organism, we performed 2-D Fluorescence Difference Gel Electrophoresis (2D-DIGE) and mass spectrometry analysis of differentially expressed proteins between sperm and spermatids in order to identify factors that could be responsible for the efficiency of the sperm to support embryonic development. Furthermore, benefiting from the availability of egg extracts in Xenopus, we also tested whether the chromatin of sperm could attract different egg factors compared to the chromatin of spermatids. Our analysis identified: (1) several proteins which were present exclusively in sperm; but not in spermatid nuclei and (2) numerous egg proteins binding to the sperm (but not to the spermatid chromatin) after incubation in egg extracts. Amongst these factors we identified many chromatin-associated proteins and transcriptional repressors. Presence of transcriptional repressors binding specifically to sperm chromatin could suggest its preparation for the early embryonic cell cycles, during which no transcription is observed and suggests that sperm chromatin has a unique protein composition, which facilitates the recruitment of egg chromatin remodelling factors. It is therefore likely that the acquisition of these sperm-specific factors during spermiogenesis makes the sperm chromatin suitable to interact with the maternal factors and, as a consequence, to support efficient embryonic development. Full article

(This article belongs to the Special Issue Cellular and Molecular Mechanisms of Sperm-Egg Interaction)

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Open AccessArticle

Sperm Proteases that May Be Involved in the Initiation of Sperm Motility in the Newt, Cynops pyrrhogaster

by Misato Yokoe, Makoto Sano, Honami Shibata, Daisuke Shibata, Eriko Takayama-Watanabe, Kazuo Inaba and Akihiko Watanabe

Int. J. Mol. Sci. 2014, 15(9), 15210-15224; https://doi.org/10.3390/ijms150915210 - 28 Aug 2014

Cited by 7 | Viewed by 7316

Abstract

A protease of sperm in the newt Cynops pyrrhogaster that is released after the acrosome reaction (AR) is proposed to lyse the sheet structure on the outer surface of egg jelly and release sperm motility-initiating substance (SMIS). Here, we found that protease activity [...] Read more.

A protease of sperm in the newt Cynops pyrrhogaster that is released after the acrosome reaction (AR) is proposed to lyse the sheet structure on the outer surface of egg jelly and release sperm motility-initiating substance (SMIS). Here, we found that protease activity in the sperm head was potent to widely digest substrates beneath the sperm. The protease activity measured by fluorescein thiocarbamoyl-casein digestion was detected in the supernatant of the sperm after the AR and the activity was inhibited by 4-(2-aminoethyl) benzenesulfonyl fluoride (AEBSF), an inhibitor for serine or cysteine protease, suggesting the release of serine and/or cysteine proteases by AR. In an in silico analysis of the testes, acrosins and 20S proteasome were identified as possible candidates of the acrosomal proteases. We also detected another AEBSF-sensitive protease activity on the sperm surface. Fluorescence staining with AlexaFluor 488-labeled AEBSF revealed a cysteine protease in the principal piece; it is localized in the joint region between the axial rod and undulating membrane, which includes an axoneme and produces powerful undulation of the membrane for forward sperm motility. These results indicate that AEBSF-sensitive proteases in the acrosome and principal piece may participate in the initiation of sperm motility on the surface of egg jelly. Full article

(This article belongs to the Special Issue Cellular and Molecular Mechanisms of Sperm-Egg Interaction)

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Open AccessArticle

Distinct Roles of Soluble and Transmembrane Adenylyl Cyclases in the Regulation of Flagellar Motility in Ciona Sperm

by Kogiku Shiba and Kazuo Inaba

Int. J. Mol. Sci. 2014, 15(8), 13192-13208; https://doi.org/10.3390/ijms150813192 - 28 Jul 2014

Cited by 20 | Viewed by 6610

Abstract

Adenylyl cyclase (AC) is a key enzyme that synthesizes cyclic AMP (cAMP) at the onset of the signaling pathway to activate sperm motility. Here, we showed that both transmembrane AC (tmAC) and soluble AC (sAC) are distinctly involved in the regulation of sperm [...] Read more.

Adenylyl cyclase (AC) is a key enzyme that synthesizes cyclic AMP (cAMP) at the onset of the signaling pathway to activate sperm motility. Here, we showed that both transmembrane AC (tmAC) and soluble AC (sAC) are distinctly involved in the regulation of sperm motility in the ascidian Ciona intestinalis. A tmAC inhibitor blocked both cAMP synthesis and the activation of sperm motility induced by the egg factor sperm activating and attracting factor (SAAF), as well as those induced by theophylline, an inhibitor of phoshodiesterase. It also significantly inhibited cAMP-dependent phosphorylation of a set of proteins at motility activation. On the other hand, a sAC inhibitor does not affect on SAAF-induced transient increase of cAMP, motility activation or protein phosphorylation, but it reduced swimming velocity to half in theophylline-induced sperm. A sAC inhibitor KH-7 induced circular swimming trajectory with smaller diameter and significantly suppressed chemotaxis of sperm to SAAF. These results suggest that tmAC is involved in the basic mechanism for motility activation through cAMP-dependent protein phosphorylation, whereas sAC plays distinct roles in increase of flagellar beat frequency and in the Ca²⁺-dependent chemotactic movement of sperm. Full article

(This article belongs to the Special Issue Cellular and Molecular Mechanisms of Sperm-Egg Interaction)

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Review

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Open AccessReview

Transmembrane Signal Transduction in Oocyte Maturation and Fertilization: Focusing on Xenopus laevis as a Model Animal

by Ken-ichi Sato

Int. J. Mol. Sci. 2015, 16(1), 114-134; https://doi.org/10.3390/ijms16010114 - 23 Dec 2014

Cited by 11 | Viewed by 10273

Abstract

Fertilization is a cell biological phenomenon of crucial importance for the birth of new life in a variety of multicellular and sexual reproduction species such as algae, animal and plants. Fertilization involves a sequence of events, in which the female gamete “egg” and [...] Read more.

Fertilization is a cell biological phenomenon of crucial importance for the birth of new life in a variety of multicellular and sexual reproduction species such as algae, animal and plants. Fertilization involves a sequence of events, in which the female gamete “egg” and the male gamete “spermatozoon (sperm)” develop, acquire their functions, meet and fuse with each other, to initiate embryonic and zygotic development. Here, it will be briefly reviewed how oocyte cytoplasmic components are orchestrated to undergo hormone-induced oocyte maturation and sperm-induced activation of development. I then review how sperm-egg membrane interaction/fusion and activation of development in the fertilized egg are accomplished and regulated through egg coat- or egg plasma membrane-associated components, highlighting recent findings and future directions in the studies using Xenopus laevis as a model experimental animal. Full article

(This article belongs to the Special Issue Cellular and Molecular Mechanisms of Sperm-Egg Interaction)

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Open AccessReview

Calcium Signaling and Meiotic Exit at Fertilization in Xenopus Egg

by Alexander A. Tokmakov, Vasily E. Stefanov, Tetsushi Iwasaki, Ken-Ichi Sato and Yasuo Fukami

Int. J. Mol. Sci. 2014, 15(10), 18659-18676; https://doi.org/10.3390/ijms151018659 - 15 Oct 2014

Cited by 17 | Viewed by 8679

Abstract

Calcium is a universal messenger that mediates egg activation at fertilization in all sexually reproducing species studied. However, signaling pathways leading to calcium generation and the mechanisms of calcium-induced exit from meiotic arrest vary substantially among species. Here, we review the pathways of [...] Read more.

Calcium is a universal messenger that mediates egg activation at fertilization in all sexually reproducing species studied. However, signaling pathways leading to calcium generation and the mechanisms of calcium-induced exit from meiotic arrest vary substantially among species. Here, we review the pathways of calcium signaling and the mechanisms of meiotic exit at fertilization in the eggs of the established developmental model, African clawed frog, Xenopus laevis. We also discuss calcium involvement in the early fertilization-induced events in Xenopus egg, such as membrane depolarization, the increase in intracellular pH, cortical granule exocytosis, cortical contraction, contraction wave, cortical rotation, reformation of the nuclear envelope, sperm chromatin decondensation and sister chromatid segregation. Full article

(This article belongs to the Special Issue Cellular and Molecular Mechanisms of Sperm-Egg Interaction)

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Open AccessReview

Molecular and Cellular Mechanisms of Sperm-Oocyte Interactions Opinions Relative to in Vitro Fertilization (IVF)

by George Anifandis, Christina Messini, Konstantinos Dafopoulos, Sotiris Sotiriou and Ioannis Messinis

Int. J. Mol. Sci. 2014, 15(7), 12972-12997; https://doi.org/10.3390/ijms150712972 - 22 Jul 2014

Cited by 49 | Viewed by 14537

Abstract

One of the biggest prerequisites for pregnancy is the fertilization step, where a human haploid spermatozoon interacts and penetrates one haploid oocyte in order to produce the diploid zygote. Although fertilization is defined by the presence of two pronuclei and the extraction of [...] Read more.

One of the biggest prerequisites for pregnancy is the fertilization step, where a human haploid spermatozoon interacts and penetrates one haploid oocyte in order to produce the diploid zygote. Although fertilization is defined by the presence of two pronuclei and the extraction of the second polar body the process itself requires preparation of both gametes for fertilization to take place at a specific time. These preparations include a number of consecutive biochemical and molecular events with the help of specific molecules and with the consequential interaction between the two gametes. These events take place at three different levels and in a precise order, where the moving spermatozoon penetrates (a) the outer vestments of the oocyte, known as the cumulus cell layer; (b) the zona pellucida (ZP); where exocytosis of the acrosome contents take place and (c) direct interaction of the spermatozoon with the plasma membrane of the oocyte, which involves a firm adhesion of the head of the spermatozoon with the oocyte plasma membrane that culminates with the fusion of both sperm and oocyte membranes (Part I). After the above interactions, a cascade of molecular signal transductions is initiated which results in oocyte activation. Soon after the entry of the first spermatozoon into the oocyte and oocyte activation, the oocyte’s coat (the ZP) and the oocyte’s plasma membrane seem to change quickly in order to initiate a fast block to a second spermatozoon (Part II). Sometimes, two spermatozoa fuse with one oocyte, an incidence of 1%–2%, resulting in polyploid fetuses that account for up to 10%–20% of spontaneously aborted human conceptuses. The present review aims to focus on the first part of the human sperm and oocyte interactions, emphasizing the latest molecular and cellular mechanisms controlling this process. Full article

(This article belongs to the Special Issue Cellular and Molecular Mechanisms of Sperm-Egg Interaction)

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Open AccessReview

Sperm-Egg Fusion: A Molecular Enigma of Mammalian Reproduction

by Karolina Klinovska, Natasa Sebkova and Katerina Dvorakova-Hortova

Int. J. Mol. Sci. 2014, 15(6), 10652-10668; https://doi.org/10.3390/ijms150610652 - 13 Jun 2014

Cited by 51 | Viewed by 13369

Abstract

The mechanism of gamete fusion remains largely unknown on a molecular level despite its indisputable significance. Only a few of the molecules required for membrane interaction are known, among them IZUMO1, which is present on sperm, tetraspanin CD9, which is present on the [...] Read more.

The mechanism of gamete fusion remains largely unknown on a molecular level despite its indisputable significance. Only a few of the molecules required for membrane interaction are known, among them IZUMO1, which is present on sperm, tetraspanin CD9, which is present on the egg, and the newly found oolema protein named Juno. A concept of a large multiprotein complex on both membranes forming fusion machinery has recently emerged. The Juno and IZUMO1, up to present, is the only known extracellular receptor pair in the process of fertilization, thus, facilitating the essential binding of gametes. However, neither IZUMO1 nor Juno appears to be the fusogenic protein. At the same time, the tetraspanin is expected to play a role in organizing the egg membrane order and to interact laterally with other factors. This review summarizes, to present, the known molecules involved in the process of sperm-egg fusion. The complexity and expected redundancy of the involved factors makes the process an intricate and still poorly understood mechanism, which is difficult to comprehend in its full distinction. Full article

(This article belongs to the Special Issue Cellular and Molecular Mechanisms of Sperm-Egg Interaction)

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