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Mouse Oocytes (mouse + oocyte)

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Life Sciences	95%

Selected Abstracts

Mouse oocytes derived from fetal germ cells are competent to support the development of embryos by in vitro fertilization

MOLECULAR REPRODUCTION & DEVELOPMENT, Issue 11 2008
Wei Shen
Figure 1B in the paper above appeared to be an enlargement of a portion of Figure 1A, while the legend indicated otherwise. This error had been made because the authors labeled the sections by date, but not by source, and because experimental and control sections were similar. The authors have now confirmed their overall results and have submitted new figures for Figure 1 and Figure 4A presented below. The original article to which this erratum refers, was published in Molecular Reproduction and Development 2006;73(10):1312,1317. Mol. Reprod. Dev. 75: 1688,1689, 2008. © 2008 Wiley-Liss, Inc. [source]

Robust computer-controlled system for intracytoplasmic sperm injection and subsequent cell electro-activation

THE INTERNATIONAL JOURNAL OF MEDICAL ROBOTICS AND COMPUTER ASSISTED SURGERY, Issue 1 2009
K. K. Tan
Abstract Introduction Intracytoplasmic sperm injection (ICSI) and the subsequent cell electro-activation process is a relatively new enhanced procedure to address male factor infertility. The current method involves the engagement of experienced embryologists for such a purpose. More advanced methodologies, which use high precision instrumentation tools, will speed up the whole procedure. Methods In this paper, the development of a computer-controlled system for ICSI and the subsequent cell electro-activation process is presented. The system is integrated to a microinjection workstation and piezo-actuator to perform the ICSI procedure, with vision capability to automatically position the components precisely. A micro-pump assembly is utilized for automatic medium refreshment and a heater plate assembly provides temperature control during the cell electro-activation process. The overall system is comprehensive, comprising modular functional components integrated within a hardware architecture. Results Experimental results on mice oocytes verified the effectiveness of the developed system over the current method. Conclusions Further improvements on the instrumentation tools will improve the robustness and overall performance of the developed system. Copyright © 2009 John Wiley & Sons, Ltd. [source]

Oosp1 encodes a novel mouse oocyte-secreted protein

GENESIS: THE JOURNAL OF GENETICS AND DEVELOPMENT, Issue 3 2001
Changning Yan
Abstract Summary: Oocyte-somatic cell communication is necessary for normal ovarian function. However, the identities of the majority of oocyte-secreted proteins remain unknown. A novel cDNA encoding mouse oocyte- secreted protein 1 (OOSP1) was identified using a modified subtractive hybridization screen. The Oosp1 cDNA encodes a 202-amino acid protein that contains a 21-amino acid signal peptide sequence, 5 putative N-linked glycosylation consensus sequences, and 6 cysteines that are predicted to form 3 disulfide bonds. OOSP1 shares amino acid identity with placental-specific protein 1 (PLAC1), a secreted protein expressed in the placenta and the ectoplacental cone. The Oosp1 mRNA is approximately 1.0 kb and is present at high levels in the oocytes of adult ovaries and at lower levels in the spleen. The mouse Oosp1 gene is 5 exons, spans greater than 16.4 kb, and localizes to chromosome 19 at a position that shares synteny with human chromosome 11q12,11q13. The identification of OOSP1 as a new oocyte-secreted protein permits future in vitro and in vivo functional analyses to define its role in ovarian folliculogenesis. genesis 31:105,110, 2001. © 2001 Wiley-Liss, Inc. [source]

Role of AMPK throughout meiotic maturation in the mouse oocyte: Evidence for promotion of polar body formation and suppression of premature activation

MOLECULAR REPRODUCTION & DEVELOPMENT, Issue 10 2010
Stephen M. Downs
Abstract This study was conducted to assess the role of AMPK in regulating meiosis in mouse oocytes from the germinal vesicle stage to metaphase II. Exposure of mouse cumulus cell-enclosed oocytes (CEO) and denuded oocytes (DO) during spontaneous maturation in vitro to AMPK-activating agents resulted in augmentation of the rate and frequency of polar body formation. Inhibitors of AMPK had an opposite, inhibitory effect. In addition, the AMPK inhibitor, compound C (Cmpd C) increased the frequency of oocyte activation. The stimulatory action of the AMPK-activating agent, AICAR, and the inhibitory action of Cmpd C were diminished if exposure was delayed, indicating an early action of AMPK on polar body formation. The frequency of spontaneous and Cmpd C-induced activation in CEO was reduced as the period of hormonal priming was increased, and AMPK stimulation eliminated the activation response. Immunostaining of oocytes with antibody to active AMPK revealed an association of active kinase with chromatin, spindle poles, and midbody during maturation. Immunolocalization of the ,1 catalytic subunit of AMPK showed an association with condensed chromatin and the meiotic spindle but not in the spindle poles or midbody; ,2 stained only diffusely throughout the oocyte. These data suggest that AMPK is involved in a regulatory capacity throughout maturation and helps promote the completion of meiosis while suppressing premature activation. Mol. Reprod. Dev. 77:888,899, 2010. © 2010 Wiley-Liss, Inc. [source]

Curcumin disrupts meiotic and mitotic divisions via spindle impairment and inhibition of CDK1 activity

CELL PROLIFERATION, Issue 4 2010
A. Bielak-Zmijewska
Objectives:, Curcumin, a natural compound, is a potent anti-cancer agent, which inhibits cell division and/or induces cell death. It is believed that normal cells are less sensitive to curcumin than malignant cells; however, the mechanism(s) responsible for curcumin's effect on normal cells are poorly understood. The aim of this study was to verify the hypothesis that curcumin affects normal cell division by influencing microtubule stability, using mouse oocyte and early embryo model systems. Materials and methods:, Maturating mouse oocytes and two-cell embryos were treated with different concentrations of curcumin (10,50 ,m), and meiotic resumption and mitotic cleavage were analysed. Spindle and chromatin structure were visualized using confocal microscopy. In addition, acetylation and in vitro polymerization of tubulin, in the presence of curcumin, were investigated and the damage to double-stranded DNA was studied using ,H2A.X. CDK1 activity was measured. Results and conclusions:, We have shown for the first time, that curcumin, in a dose-dependent manner, delays and partially inhibits meiotic resumption of oocytes and inhibits meiotic and mitotic divisions by causing disruption of spindle structure and does not induce DNA damage. Our analysis indicated that curcumin affects CDK1 kinase activity but does not directly affect microtubule polymerization and tubulin acetylation. As our study showed that curcumin impairs generative and somatic cell division, its future clinical use or of its derivatives with improved bioavailability after oral administration, should take into consideration the possibility of extensive side-effects on normal cells. [source]

Cytoplasmic localization of oocyte-specific variant of porcine DNA methyltransferase-1 during early development

DEVELOPMENTAL DYNAMICS, Issue 7 2009
Young Sun Jeong
Abstract DNA methyltransferase-1 (Dnmt1) is involved in the maintenance of genomic methylation patterns. Rather than full-length Dnmt1, mouse oocytes have a truncated variant called Dnmt1o. Immunofluorescence data showed that Dnmt1o localized to the cytoplasm, but this has not been confirmed using more direct methods. The cytoplasmic localization of Dnmt1o has been assigned to the main cause of global DNA demethylation in early mouse embryos. We studied localization of Dnmt1o in mouse and pig embryos. We identified pig Dnmt1o protein and its transcript with unique 5,-end sequence. Physically separating mouse and pig 2-cell embryos into their nuclear and cytoplasmic components demonstrated that Dnmt1o of both species localized to the cytoplasm. Cloned pig embryos had Dnmt1o as the main form, with no indication of somatic Dnmt1. These findings indicate that Dnmt1o is cytoplasmic during early development; its presence in both pig and mouse embryos further suggests that Dnmt1o is conserved in mammals. Developmental Dynamics 238:1666,1673, 2009. © 2009 Wiley-Liss, Inc. [source]

De Novo DNA methylation independent establishment of maternal imprint on X chromosome in mouse oocytes

GENESIS: THE JOURNAL OF GENETICS AND DEVELOPMENT, Issue 12 2008
Hatsune Chiba
Mouse blastocyst stage embryo stained for histone H3 lysine-27 trimethylation (red) and DNA (blue). H3K27me3 marks the inactive X chromosome. The study by Chiba et al. in this issue suggests that de novo DNA methyltransferases are dispensable for setting the imprint on the maternally-derived X chromsome in growing oocytes. See Chiba et al. in this issue. [source]

Role of AMPK throughout meiotic maturation in the mouse oocyte: Evidence for promotion of polar body formation and suppression of premature activation

Fatty acid oxidation and meiotic resumption in mouse oocytes

MOLECULAR REPRODUCTION & DEVELOPMENT, Issue 9 2009
Stephen M. Downs
We have examined the potential role of fatty acid oxidation (FAO) in AMP-activated protein kinase (AMPK)-induced meiotic maturation. Etomoxir and malonyl CoA, two inhibitors of carnitine palmitoyl transferase-1 (CPT1), and thus FAO, blocked meiotic induction in dbcAMP-arrested cumulus cell-enclosed oocytes (CEO) and denuded oocytes (DO) by the AMPK activator, AICAR. C75, an activator of CPT1 and FAO, stimulated meiotic resumption in CEO and DO. This effect was insensitive to the AMPK inhibitor, compound C, indicating an action downstream of AMPK. Palmitic acid or carnitine also promoted meiotic resumption in DO in the presence of AICAR. Since C75 also suppresses the activity of fatty acid synthase (FAS), we tested another FAS inhibitor, cerulenin. Cerulenin stimulated maturation in arrested oocytes, but to a lesser extent, exhibited significantly slower kinetics and was effective in CEO but not DO. Moreover, etomoxir completely blocked C75-induced maturation but was ineffective in cerulenin-treated oocytes, suggesting that the meiosis-inducing action of C75 is through activation of FAO within the oocyte, while that of cerulenin is independent of FAO and acts within the cumulus cells. Finally, we determined that long chain, but not short chain, fatty acyl carnitine derivatives were stimulatory to oocyte maturation. Palmitoyl carnitine stimulated maturation in both CEO and DO, with rapid kinetics in DO; this effect was blocked by mercaptoacetate, a downstream inhibitor of FAO. These results indicate that activation of AMPK stimulates meiotic resumption in mouse oocytes by eliminating a block to FAO. Mol. Reprod. Dev. 76: 844,853, 2009. © 2009 Wiley-Liss, Inc. [source]

Changes in global histone acetylation pattern in somatic cell nuclei after their transfer into oocytes at different stages of maturation

MOLECULAR REPRODUCTION & DEVELOPMENT, Issue 3 2008
Helena Fulka
Abstract In our study, we have examined the pattern of global histone modification changes in somatic cell nuclei after their transfer into mouse oocytes at different stages of maturation or after their parthenogenetic activation. While germinal vesicle (GV) staged immature oocytes are strongly labeled with anti-acetylated histone H3 and H4 antibodies, the signal is absent in both metaphase I and metaphase II oocytes (MI, MII). In contrast, the oocytes of all maturation stages show a presence of trimethylated H3/K4 in their chromatin. When somatic cells were fused to intact or enucleated GV oocytes, both the GV and the somatic cell nucleus showed a very strong signal for all the antibodies used. On the other hand, when somatic cells nuclei that are AcH3 and AcH4 positive before fusion are introduced into either intact or enucleated MI or MII oocytes, their acetylation signal decreased rapidly and was totally absent after a prolonged culture. This was not the case when anti-trimethyl H3/K4 antibody was used. The somatic cell chromatin showed only a slight decrease in the intensity of labeling after its transfer into MI or MII oocytes. This decrease was, however, evident only after a prolonged culture. These results suggest not only a relatively higher stability of the methylation modification but also some difference between the oocyte and somatic chromatin. The ability to deacetylate the chromatin of transferred somatic nuclei disappears rapidly after the oocyte activation. Our results indicate that at least some reprogramming activity appears in the oocyte cytoplasm almost immediately after GV breakdown (GVBD), and that this activity rapidly disappears after the oocyte activation. Mol. Reprod. Dev. 75: 556,564, 2008. © 2007 Wiley-Liss, Inc. [source]

Factors affecting the in vitro action of cumulus cells on the maturing mouse oocytes

MOLECULAR REPRODUCTION & DEVELOPMENT, Issue 1 2008
Li Ge
Abstract The removal of cumulus cells (CCs) from oocytes at the germinal vesicle (GV) stage still represents a major limitation in such embryo techniques as GV transfer, somatic cell haploidization, and oocyte cryopreservation. However, no efficient in vitro maturation (IVM) system for CC-denuded oocytes (DOs) has been established in mammalian species. Although follicular cells are considered to play an important role in oocyte maturation, the specific role and mechanisms of action of different cell types are poorly understood. Reports on whether junctional association between CCs and the oocyte is essential for the beneficial effect of CC co-culture on oocyte maturation are in conflict. Our objective was to try to address these issues using the mouse oocyte model. The results indicated that while co-culture with the CC monolayer could only partially restore the developmental potential of DOs without corona cells, it restored the competence of corona-enclosed DOs completely. Culture in medium conditioned with CC monolayer also promoted maturation of DOs. However, co-culture with the monolayer of mural granulosa cells had no effect. The efficiency of CC co-culture was affected by various factors such as density and age of the CCs, the presence of gonadotropin in the maturation medium and the duration for in vivo (IVO) gonadotropin priming. It is concluded that mouse CCs produce a diffusible factor(s) that support DO maturation in a CC-oocyte junctional communication dependent manner. The data will contribute to our understanding the mechanisms by which CCs promote oocyte maturation and to the establishment of an efficient DO IVM system. Mol. Reprod. Dev. 75: 136,142, 2008. © 2007 Wiley-Liss, Inc. [source]

Reduced oxygen concentration improves the developmental competence of mouse oocytes following in vitro maturation

MOLECULAR REPRODUCTION & DEVELOPMENT, Issue 7 2007
Kimberly A. Preis
Abstract Reduced atmospheric oxygen concentration is beneficial to embryo development; however, optimal oxygen concentration for oocyte maturation remains undetermined. Likewise, there is no consensus of appropriate medium supplementation during maturation. The objective of this study was to determine whether oxygen tension (20% or 5% O2) and epidermal growth factor (EGF) affect oocyte metabolism and subsequent embryo development. Cumulus-oocyte complexes (COCs) were collected from 28-day-old equine chorionic gonadotropin (eCG) primed or unprimed F1 (C57BL/6xCBA) mice. COCs were matured in defined medium in one of four groups: 20% O2, 20% O2,+,EGF, 5% O2, 5% O2,+,EGF. In vivo matured COCs were also collected for analysis. COCs from unprimed mice, matured in 5% O2,±,EGF or 20% O2,+,EGF had higher metabolic rates than COCs matured in 20% O2 (P,<,0.05). COCs from primed mice had higher metabolic rates when matured in the presence of EGF, regardless of oxygen tension (P,<,0.01). Oxygen uptake and mitochondrial membrane potential were higher for in vivo matured oocytes and oocytes matured under 5% O2 compared to oocytes matured under 20% O2 (P,<,0.05). Blastocyst formation was not different between maturation groups (primed or unprimed); however, embryo cell numbers were 20,45% significantly higher when COCs were matured at 5% O2 (P,<,0.05). Results suggest that oocytes matured in physiological concentrations of oxygen have improved development and metabolic activity, more closely resembling in vivo maturation. These findings have implications for oocyte maturation in both clinical and research laboratories. Mol. Reprod. Dev. 74: 893,903, 2007. © 2006 Wiley-Liss, Inc. [source]

Inactivation of MAPK affects centrosome assembly, but not actin filament assembly, in mouse oocytes maturing in vitro

MOLECULAR REPRODUCTION & DEVELOPMENT, Issue 7 2007
Seung-Eun Lee
Abstract Mitogen-activated protein kinase (MAPK) plays a crucial role in meiotic maturation of mouse oocytes. In order to understand the mechanism by which MAPK regulates meiotic maturation, we examined the effects of the MAPK pathway inhibitor U0126 on microtubule organization, ,-tubulin and nuclear mitotic apparatus protein (NuMA) distribution, and actin filament assembly in mouse oocytes maturing in vitro. Western blotting with antibodies that detect active, phosphorylated MAPK revealed that MAPK was inactive in fully grown germinal vesicle (GV) oocytes. Phosphorylated MAPK was first detected 3 hr after the initiation of maturation cultures, was fully active at 6 hr, and remained active until metaphase II. Treatment of GV stage oocytes with 20 µM U0126 completely blocked MAPK phosphorylation, but did not affect GV breakdown (GVBD). However, the oocytes did not progress to the Metaphase I stage, which would normally occur after 9 hr in the maturation cultures. The inhibition of MAPK resulted in abnormal spindles and abnormal distributions of ,-tubulin and NuMA, but did not affect actin filament assembly. In oocytes treated with U0126 after GVBD, polar body extrusion was normal, but the organization of the metaphase plate and chromosome segregation were abnormal. In conclusion, the meiotic abnormalities caused by U0126, a specific inhibitor of MAPK signaling, indicate that MAPK plays an important regulatory role in microtubule and centrosome assembly, but not actin filament assembly. Mol. Reprod. Dev. © 2006 Wiley-Liss, Inc. Mol. Reprod. Dev. 74: 904,911, 2007. © 2007 Wiley-Liss, Inc. [source]

Proteomic profiling of murine oocyte maturation,

MOLECULAR REPRODUCTION & DEVELOPMENT, Issue 5 2007
Alejandra M. Vitale
Abstract In an effort to better understand oocyte function, we utilized two-dimensional (2D) electrophoresis and mass spectrometry to identify proteins that are differentially expressed during murine oocyte maturation. Proteins from 500 germinal vesicle (GV) and metaphase II-(MII) arrested oocytes were extracted, resolved on 2D electrophoretic gels, and stained with silver. Analysis of the gels indicated that 12 proteins appeared to be differentially expressed between the GV and MII stage. These proteins were then cored from the 2D gels and identified by mass spectrometry as: transforming acidic coiled-coil protein 3 (TACC3), heat shock protein 105 (HSP105), programmed cell death six-interacting protein (PDCD6IP), stress-inducible phosphoprotein (STI1), importin ,2, adenylsuccinate synthase (ADDS), nudix, spindlin, lipocalin, lysozyme, translationally controlled tumor protein (TCTP), and nucleoplasmin 2 (NPM2). Interestingly, PDCD6IP, importin ,2, spindlin, and NPM2 appear slightly larger in mass and more acidic on the MII oocyte gel compared to the GV oocyte gel, suggesting that they may be post-translationally modified during oocyte maturation. Given NPM2 is an oocyte-restricted protein, we chose to further investigate its properties during oocyte maturation and preimplantation development. Real-Time RT-PCR showed that NPM2 mRNA levels rapidly decline at fertilization. Indirect immunofluorescence analysis showed that, with the exception of cortical localization in MII-arrested oocytes, NPM2 is localized to the nucleus of both GV stage oocytes and all stages of preimplantation embryos. We then performed one-dimensional (1D) western blot analysis of mouse oocytes and preimplantation embryos and found that, as implicated by the 2D gel comparison, NPM2 undergoes a phosphatase-sensitive electrophoretic mobility shift during the GV to MII transition. The slower migrating NPM2 form is also present in pronuclear embryos but by the two-cell stage, the majority of NPM2 exists as the faster migrating form, which persists to the blastocyst stage. Mol. Reprod. Dev. 74: 608,616, 2007. © 2006 Wiley-Liss, Inc. [source]

Effects of delayed excision of oviducts/ovaries on mouse oocytes and embryos

MOLECULAR REPRODUCTION & DEVELOPMENT, Issue 4 2007
De-Qiang Miao
Abstract To achieve the best and reproducible results of experiments, effects of delayed excision of oviducts/ovaries on mouse ovarian/ovulated oocytes and embryos have been studied. Oviducts/ovaries were excised at different times after death of mice and effects of the postmortem interval on ovarian/ovulated oocytes and embryos were analyzed. When oviduct excision was delayed 10 min, many ovulated oocytes lysed or underwent in vitro spontaneous activation, and this postmortem effect aggravated with the extension of postmortem interval and oocyte aging. Oocytes from different mouse strains responded differently to delayed oviduct removal. Delayed oviduct excision did not cause lysis of zygotes or embryos but compromised their developmental potential. When ovaries were excised at 30 min after death, percentages of atretic follicles increased while blastocyst cell number declined significantly after oocyte maturation in vitro. Preservation of oviducts in vitro, in intact or opened abdomen at different temperatures and histological analysis of oviducts from different treatments suggested that toxic substance(s) were secreted from the dying oviducts which induced oocyte lysis and spontaneous activation and both this effect itself and the sensitivity of oocytes to this effect was temperature dependent. It is concluded that a short delay of oviduct/ovary removal had marked detrimental effects on oocytes and embryos. This must be taken into account in experiments using oocytes or embryos from slaughtered animals. The data may also be important for estimation of the time of death in forensic medicine and for rescue of oocytes from deceased valuable or endangered mammals. Mol. Reprod. Dev. 74: 468,477, 2007. © 2006 Wiley-Liss, Inc. [source]

Eomesodermin is expressed in mouse oocytes and pre-implantation embryos

MOLECULAR REPRODUCTION & DEVELOPMENT, Issue 4 2005
Josie McConnell
Abstract T-box genes are a highly conserved family of genes encoding transcription factors, which share a conserved DNA binding domain (the T-box). Appropriate temporal and spatial expression of this gene family is critical for gastrulation and organogenesis in a number of species. The T-box containing gene Eomesodermin was first identified in Xenopus, where it plays a critical role in mesoderm formation. In situ analyses in mice have described the expression patterns of the mouse ortholog of this gene mEomesodermin (mEomes) at the time of implantation and during fetal development. Additional studies involving the disruption of the mEomes gene, have demonstrated an additional role for mEomes in trophoblast formation. However, these analyses did not address the possibility that maternally encoded or pre-blastocyst zygotic transcription of mEomes may also contribute to embryonic development. We show here that mEomes mRNA is present prior to blastocyst formation, and that the protein product of mEomes is associated with nuclear DNA during oocyte development and persistently localizes within all nuclei of the preimplantation embryo until the early blastocyst stage. mEomes protein is associated with the meiotic spindle in the unfertilized egg and with the mitotic spindle at each cell division. Our results are consistent with mEomesodermin having a role in early preimplantation development and inner cell mass formation in addition to its function in the trophoblast lineage. Mol. Reprod. Dev. © 2005 Wiley-Liss, Inc. [source]

Specific maternal transcripts in bovine oocytes and cleavaged embryos: Identification with novel DDRT-PCR methods

MOLECULAR REPRODUCTION & DEVELOPMENT, Issue 3 2005
Kyu-Chan Hwang
Abstract We used annealing control primer (ACP)-based differential display reverse transcription polymerase chain reaction (DDRT-PCR) to isolate differentially expressed amplicons in bovine germinal vesicle (GV) stage oocytes, 8-cell stage embryos produced in vitro, and blastocyst stage embryos produced in vitro. Four expressed sequence tags (ESTs) of genes that were specifically and predominantly expressed in GV oocytes were cloned and sequenced. We have used a fluorescence monitored real-time quantitative PCR (qPCR) to quantify and analyzed the temporal expression of the target differentially expressed transcripts throughout the preimplantation stages from oocytes to blastocysts. The cloned genes or ESTs all exhibited significant sequence similarity with known bovine genes (98%,100%; DNCL1 and ZP2) or ESTs (81%,97%; FANK1 and GTL3) of other species. As revealed by real-time qRT-PCR, DNCL1, FANK1, GTL3, and ZP2 transcripts were observed in the GV stage oocytes and expression gradually decreased up to the 8-cell stage embryo and the transcripts were not detected in later stages. Similarly, upregulation was observed in GV stage mouse oocytes and metaphase II, suggesting that these four differentially expressed orthologous genes play important roles in early preimplantation, as maternally-derived transcripts. Mol. Reprod. Dev. © 2005 Wiley-Liss, Inc. [source]

Maternal chromatin remodeling during maturation and after fertilization in mouse oocytes

MOLECULAR REPRODUCTION & DEVELOPMENT, Issue 2 2004
Marcella Spinaci
Abstract Immunofluorescence staining with antibodies against acetylated histone H4 and 5-methylcytosine was carried out to investigate female chromatin remodeling throughout oocyte maturation and chromatin rearrangement involving both male and female genomes after fertilization. Oocyte cytoplasm remodels female chromatin in preparation of the fertilizing event and the subsequent chromatin rearrangement. Histone H4 are in fact progressively deacetylated whereas demethylating enzymes do not seem to be active over this period. The acetylase/deacetylase balance seems to be cell cycle dependent as female chromatin is deacetylated during maturation and reacetylated at telophase II stage both after fertilization and activation. On the contrary, DNA demethylation seems to be strictly selective. It is in fact confined to the remodeling of paternal genome after fertilization of mature oocytes as the ooplasm is not effective in demethylating either paternal chromatin in germinal vesicle breakdown (GVBD) fertilized oocytes or maternal genome of partenogenetically activated oocytes. Surprisingly, we induced maternal chromatin demethylation after fertilization by treating oocytes with a combination of a methyltransferase inhibitor, 5-azacytidine (5-AzaC), and a reversible and specific inhibitor of histone deacetylase, trichostatin A (TSA). This treatment likely induces a hyperacetylation of histones (thus favoring the access to demethylating enzymes by opening female chromatin structure) associated with a block of reparative methylation by inhibiting methytransferases. This manipulation of chromatin remodeling may have applications regarding the biological significance of aberrant DNA methylation. Mol. Reprod. Dev. 69: 215,221, 2004. © 2004 Wiley-Liss, Inc. [source]

Phosphorylation of inositol 1,4,5-triphosphate receptor 1 during in vitro maturation of porcine oocytes

ANIMAL SCIENCE JOURNAL, Issue 1 2010
Junya ITO
ABSTRACT During fertilization in mammalian species, a sperm-induced intracellular Ca2+ signal ([Ca2+]i) mediates both exit of meiosis and oocyte activation. Recently, we demonstrated in mouse oocytes that the phosphorylation levels of inositol 1,4,5 trisphosphate receptor type1 (IP3R1), the channel responsible for Ca2+ release and oscillations during fertilization, changed during maturation and fertilization. Therefore, we examined the expression and phosphorylation of IP3R1 during in vitro maturation of pig oocytes. Here, our present study shows that expression of IP3R1 protein did not change during maturation, although the phosphorylation status of the receptor, specifically at an MPM-2 epitope, did. We found that while at the beginning of maturation IP3R1 lacked MPM-2 immunoreactivity, it became MPM-2 reactive by 24 h and reached maximal reactivity by 36 h. Interestingly, the acquisition of MPM-2 reactivity coincided with the activation of p34cdc2 kinase and mitogen-activated protein kinase (MAPK), which are involved in meiotic progression. Following completion of maturation, inactivation of MAPK by U0126 did not affect IP3R1 phosphorylation, although inactivation of p34cdc2 kinase by roscovitine dramatically reduced IP3R1 phosphorylation. Neither inhibitor affected total expression of IP3R1. Altogether, our results show that IP3R1 undergoes dynamic phosphorylation during maturation and this might underlie the generation of oscillations at fertilization. [source]

Retrotransposons and regulatory suites

BIOESSAYS, Issue 2 2005
James A. Shapiro
Cellular differentiation and multicellular development require the programmed expression of coregulated suites of genetic loci dispersed throughout the genome. How do functionally diverse loci come to share common regulatory motifs? A new paper finds that retrotransposons (RTEs) may play a role in providing common regulation to a group of functions expressed during the development of oocytes and preimplantation embryos. Examining cDNA libraries, Peaston et al.1 find that 13% of all processed transcripts in full-grown mouse oocytes contain RTE sequences, mostly from the MT family of retroviral-like elements. Smaller but still significant percentages of RTE sequences are found in cDNA libraries from 2-cell embryos and blastocysts. A quarter of these RTE sequences are at the 5, ends of chimeric transcripts that also contain exons from endogenous mouse loci. These chimeric transcripts display restricted expression in oocytes and preimplantation embryos and presumably originate from developmentally regulated LTR promoters. Some, but not all, chimeric transcripts encode novel protein products. BioEssays 27:122,125, 2005. © 2005 Wiley Periodicals, Inc. [source]