scholarly journals Functionality of the spindle checkpoint during the first meiotic division of mammalian oocytes

Reproduction ◽  
2003 ◽  
pp. 443-450 ◽  
Author(s):  
S Brunet ◽  
G Pahlavan ◽  
S Taylor ◽  
B Maro
Keyword(s):  
Chromosome Segregation ◽  
Meiotic Division ◽  
Mapk Pathway ◽  
Spindle Checkpoint ◽  
Meiotic Maturation ◽  
Metaphase Arrest ◽  
Eukaryotic Evolution ◽  
Microtubule Depolymerization ◽  
Mouse Oocytes ◽  
M Phase

The spindle checkpoint ensures accurate chromosome segregation by delaying anaphase until all chromosomes are correctly aligned on the microtubule spindle. Although this mechanism is conserved throughout eukaryotic evolution, it is unclear whether it operates during meiosis in female mammals. The results of the present study show that in mouse oocytes spindle alterations prevent both chromosome segregation and MPF (M phase promoting factor) inactivation during the first meiotic M phase. Moreover, the spindle checkpoint component budding uninhibited by benzimidazole 1 (BUB1) localizes to kinetochores and is phosphorylated until anaphase of both meiotic M phases. Both localization and phosphorylation are similar to those observed in oocytes at microtubule depolymerization. In addition, the kinetochore localization and phosphorylation of BUB1 do not depend on the MOS/.../MAPK pathway. These data indicate that the spindle checkpoint is probably active during meiotic maturation in mouse oocytes. BUB1 remains associated with kinetochores and is phosphorylated during the metaphase arrest of the second meiotic M phase, indicating that this protein may also play a role in the natural metaphase II arrest in mammalian oocytes.

scholarly journals Cell cycle modification during the transitions between meiotic M-phases in mouse oocytes

1992 ◽  
Vol 102 (3) ◽  
pp. 457-467 ◽  
Author(s):  
J.Z. Kubiak ◽  
M. Weber ◽  
G. Geraud ◽  
B. Maro
Keyword(s):  
Kinase Activity ◽  
Polar Body ◽  
Metaphase Arrest ◽  
Phosphorylated Proteins ◽  
Mouse Oocytes ◽  
Sequence Of Events ◽  
M Phase ◽  
Second Polar Body ◽  
Polar Body Extrusion ◽  
Centrosomal Proteins

When metaphase II-arrested mouse oocytes (M II) are activated very soon after ovulation, they respond abortively by second polar body extrusion followed by another metaphase arrest (metaphase III, M III; Kubiak, 1989). The M II/M III transition resembles the natural transition between the first and second meiotic metaphases (M I/M II). We observed that a similar sequence of events takes place during these two transitions: after anaphase, a polar body is extruded, the microtubules of the midbody disappear rapidly and a new metaphase spindle forms. The MPM-2 monoclonal antibody (which reacts with phosphorylated proteins associated with the centrosome during M-phase) stains discrete foci of peri-centriolar material only in metaphase arrested oocytes; during both transitional periods, a diffuse staining is observed, suggesting that these centrosomal proteins are dephosphorylated, as in a normal interphase. However, the chromosomes always remain condensed and an interphase network of microtubules is never observed during the transitional periods. Incorporation of 32P into proteins increases specifically during the transitional periods. Pulse-chase experiments, after labeling of the oocytes in M phase with 32P, showed that a 62 kDa phosphoprotein band disappears at the time of polar body extrusion. Histone H1 kinase activity (which reflects the activity of the maturation promoting factor) drops during both transitional periods to the level characteristic of interphase and then increases when the new spindle forms. Both the M I/M II and M II/M III transitions require protein synthesis as demonstrated by the effect of puromycin. These results suggest that the two M-phase/M-phase transitions are probably driven by the same molecular mechanism.

scholarly journals The spindle checkpoint protein Mad2 regulates APC/C activity during prometaphase and metaphase of meiosis I in Saccharomyces cerevisiae

2011 ◽  
Vol 22 (16) ◽  
pp. 2848-2861 ◽  
Author(s):  
Dai Tsuchiya ◽  
Claire Gonzalez ◽  
Soni Lacefield
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Cell Cycle ◽  
Chromosome Segregation ◽  
Meiotic Division ◽  
Spindle Checkpoint ◽  
Yeast Cells ◽  
Meiotic Cell ◽  
Checkpoint Protein ◽  
Sister Chromatids ◽  
Meiosis I

In many eukaryotes, disruption of the spindle checkpoint protein Mad2 results in an increase in meiosis I nondisjunction, suggesting that Mad2 has a conserved role in ensuring faithful chromosome segregation in meiosis. To characterize the meiotic function of Mad2, we analyzed individual budding yeast cells undergoing meiosis. We find that Mad2 sets the duration of meiosis I by regulating the activity of APCCdc20. In the absence of Mad2, most cells undergo both meiotic divisions, but securin, a substrate of the APC/C, is degraded prematurely, and prometaphase I/metaphase I is accelerated. Some mad2Δ cells have a misregulation of meiotic cell cycle events and undergo a single aberrant division in which sister chromatids separate. In these cells, both APCCdc20 and APCAma1 are prematurely active, and meiosis I and meiosis II events occur in a single meiotic division. We show that Mad2 indirectly regulates APCAma1 activity by decreasing APCCdc20 activity. We propose that Mad2 is an important meiotic cell cycle regulator that ensures the timely degradation of APC/C substrates and the proper orchestration of the meiotic divisions.

Activation of p90rsk during meiotic maturation and first mitosis in mouse oocytes and eggs: MAP kinase-independent and -dependent activation

Development ◽  
1996 ◽  
Vol 122 (6) ◽  
pp. 1957-1964 ◽  
Author(s):  
P. Kalab ◽  
J.Z. Kubiak ◽  
M.H. Verlhac ◽  
W.H. Colledge ◽  
B. Maro
Keyword(s):  
Kinase Activity ◽  
Polar Body ◽  
Ribosomal Subunit ◽  
Germinal Vesicle ◽  
Maximum Level ◽  
Meiotic Maturation ◽  
S6 Kinase ◽  
Mouse Oocytes ◽  
M Phase ◽  
Second Polar Body

Mitogen-activated protein kinases (MAPK) become activated during the meiotic maturation of oocytes from many species; however, their molecular targets remain unknown. This led us to characterize the activation of the ribosomal subunit S6 kinase of Mr 82 X 10(3) - 92 X 10(3) (p90rsk; a major substrate of MAPK in somatic cells) in maturing mouse oocytes and during the first cell cycle of the mouse embryo. We assessed the phosphorylation state of p90rsk by examining the electrophoretic mobility shifts on immunoblots and measured the kinase activity of immunoprecipitated p90rsk on a S6-derived peptide. Germinal vesicle stage (GV) oocytes contained a doublet of Mr 82 × 10(3) and 84 × 10(3) with a low S6 peptide kinase activity (12% of the maximum level found in metaphase II oocytes). A band of Mr 86 × 10(3) was first observed 30 minutes after GV breakdown (GVBD) and became prominent within 2 to 3 hours. MAPK was not phosphorylated 1 hour after GVBD, when the p90rsk-specific S6 kinase activity reached 37 % of the M II level. 2 hours after GVBD, MAPK became phosphorylated and p90rsk kinase activity reached 86% of the maximum level. The p90rsk band of Mr 88 × 10(3), present in mature M II oocytes when S6 peptide kinase activity is maximum, appeared when MAPK phosphorylation was nearly complete (2.5 hours after GVBD). In activated eggs, the dephosphorylation of p90rsk to Mr 86 X 10(3) starts about 1 hour after the onset of pronuclei formation and continues very slowly until the beginning of mitosis, when the doublet of Mr 82 X 10(3) and 84 X 10(3) reappears. A role for a M-phase activated kinase (like p34cdc2) in p90rsk activation was suggested by the reappearance of the Mr 86 X 10(3) band during first mitosis and in 1-cell embryos arrested in M phase by nocodazole. The requirement of MAPK for the full activation of p90rsk during meiosis was demonstrated by the absence of the fully active Mr 88 X 10(3) band in maturing c-mos −/− oocytes, where MAPK is not activated. The inhibition of kinase activity in activated eggs by 6-DMAP after second polar body extrusion provided evidence that both MAPK- and p90rsk-specific phosphatases are activated at approximately the same time prior to pronuclei formation.

Mos is required for MAP kinase activation and is involved in microtubule organization during meiotic maturation in the mouse

Development ◽  
1996 ◽  
Vol 122 (3) ◽  
pp. 815-822 ◽  
Author(s):  
M.H. Verlhac ◽  
J.Z. Kubiak ◽  
M. Weber ◽  
G. Geraud ◽  
W.H. Colledge ◽  
...  
Keyword(s):  
Map Kinase ◽  
Meiotic Maturation ◽  
Microtubule Organization ◽  
Kinase Activation ◽  
Mouse Oocytes ◽  
M Phase ◽  
In The Wild ◽  
Cytostatic Factor ◽  
Map Kinase Kinase Kinase ◽  
Map Kinase Kinase

Mos is normally expressed during oocyte meiotic maturation in vertebrates. However, apart from its cytostatic factor (CSF) activity, its precise role during mouse meiosis is still unknown. First, we analyzed its role as a MAP kinase kinase kinase. Mos is synthesized concomitantly with the activation of MAP kinase in mouse oocytes. Moreover, MAP kinase is not activated during meiosis in oocytes from mos −/− mice. This result implies that Mos is necessary for MAP kinase activation in mouse oocytes. Raf-1, another MAP kinase kinase kinase, is already present in immature oocytes, but does not seem to be active when MAP kinase is activated. Moreover, the absence of MAP kinase activation in mos −/− oocytes demonstrates that Raf-1 cannot compensate for the lack of Mos. These results suggest that Raf-1 is not involved in MAP kinase activation. Second, we analyzed the organization of the microtubules and chromosomes in oocytes from mos −/− mice. We observed that during the transition between two meiotic M-phases, the microtubules and chromosomes evolve towards an interphase-like state in mos −/− oocytes, while in the control mos +/− oocytes they remain in an M-phase configuration, as in the wild type. Moreover, after spontaneous activation, the majority of mos −/− oocytes are arrested for at least 10 hours in a third meiotic M-phase where they exhibit monopolar half-spindles. These observations present the first evidence, in intact oocytes, of a role for the Mos/…/MAP kinase cascade in the control of microtubule and chromatin organization during meiosis.

348 INACTIVATION OF MAPK AFFECTS CENTROSOME ASSEMBLY, BUT NOT ACTIN FILAMENT ASSEMBLY, IN MOUSE OOCYTES MATURING IN VITRO

2007 ◽  
Vol 19 (1) ◽  
pp. 289
Author(s):  
S. E. Lee ◽  
J. H. Kim ◽  
Y. K. Ji ◽  
N. H. Kim
Keyword(s):  
Actin Filament ◽  
Mapk Pathway ◽  
Polar Body ◽  
Metaphase Plate ◽  
Specific Inhibitor ◽  
Mapk Signaling ◽  
Meiotic Maturation ◽  
Mouse Oocytes ◽  
Filament Assembly

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, γ-tubulin and NuMA distribution, and actin filament assembly in mouse oocytes maturing in vitro. Significant differences between control and treatment were determined using Tukey's multiple range test; P < 0.05 was considered significant. 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 h after the initiation of maturation cultures, was fully active at 6 h, and remained active until metaphase II (MII). When GV oocytes were cultured in medium containing 20 �M U0126 for 9 h, the oocytes underwent geminal vesicle breakdown (GVBD), but were significantly inhibited in progression to the MI stage as compared to controls (control: 76.7 � 5.66%; 10 �M U0126: 29.0 � 7.35%; 20 �M U0126: 26.4 � 5.74%). Polar body extrusion was observed in most oocytes when cultured in the presence of 20 �M U0126 from 4 h after initiation of in vitro oocyte maturation to 14 h into the in vitro maturation incubation (control: 76.6 � 4.21%; 20 �M U0126: 70.7 � 4.61%). However, there was a decrease in normal MII-stage oocytes when cultured for 4 to 14 h (control: 87.7 � 5.12%; 20 �M U0126: 32.8 � 3.79%); abnormality included, for example, a polar body lacking a nucleus, as well as an abnormal spindle or metaphase plate. The inhibition of MAPK resulted in abnormal spindles and abnormal distributions of γ-tubulin and NuMA, but did not affect actin filament assembly. 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.

Meiotic CENP-C is a shepherd: bridging the space between the centromere and the kinetochore in time and space

2020 ◽  
Vol 64 (2) ◽  
pp. 251-261
Author(s):  
Jessica E. Fellmeth ◽  
Kim S. McKim
Keyword(s):  
Chromosome Segregation ◽  
New Technologies ◽  
Early Prophase ◽  
Unique Role ◽  
Kinetochore Assembly ◽  
M Phase ◽  
In Vivo Analysis ◽  
Meiosis I

Abstract While many of the proteins involved in the mitotic centromere and kinetochore are conserved in meiosis, they often gain a novel function due to the unique needs of homolog segregation during meiosis I (MI). CENP-C is a critical component of the centromere for kinetochore assembly in mitosis. Recent work, however, has highlighted the unique features of meiotic CENP-C. Centromere establishment and stability require CENP-C loading at the centromere for CENP-A function. Pre-meiotic loading of proteins necessary for homolog recombination as well as cohesion also rely on CENP-C, as do the main scaffolding components of the kinetochore. Much of this work relies on new technologies that enable in vivo analysis of meiosis like never before. Here, we strive to highlight the unique role of this highly conserved centromere protein that loads on to centromeres prior to M-phase onset, but continues to perform critical functions through chromosome segregation. CENP-C is not merely a structural link between the centromere and the kinetochore, but also a functional one joining the processes of early prophase homolog synapsis to late metaphase kinetochore assembly and signaling.

Regulation of spontaneous and induced resumption of meiosis in mouse oocytes by different intracellular pathways

Reproduction ◽  
2000 ◽  
pp. 377-383 ◽  
Author(s):  
L Leonardsen ◽  
A Wiersma ◽  
M Baltsen ◽  
AG Byskov ◽  
CY Andersen
Keyword(s):  
Signal Transduction ◽  
Protein Kinase ◽  
Protein Kinase A ◽  
Mitogen Activated Protein Kinase ◽  
Meiotic Maturation ◽  
Mouse Oocytes ◽  
Mitogen Activated Protein ◽  
Dependent Signal ◽  
Kinase Pathway ◽  
Kinase A

The mitogen-activated protein kinase-dependent and the cAMP-protein kinase A-dependent signal transduction pathways were studied in cultured mouse oocytes during induced and spontaneous meiotic maturation. The role of the mitogen-activated protein kinase pathway was assessed using PD98059, which specifically inhibits mitogen-activated protein kinase 1 and 2 (that is, MEK1 and MEK2), which activates mitogen-activated protein kinase. The cAMP-dependent protein kinase was studied by treating oocytes with the protein kinase A inhibitor rp-cAMP. Inhibition of the mitogen-activated protein kinase pathway by PD98059 (25 micromol l(-1)) selectively inhibited the stimulatory effect on meiotic maturation by FSH and meiosis-activating sterol (that is, 4,4-dimethyl-5alpha-cholest-8,14, 24-triene-3beta-ol) in the presence of 4 mmol hypoxanthine l(-1), whereas spontaneous maturation in the absence of hypoxanthine was unaffected. This finding indicates that different signal transduction mechanisms are involved in induced and spontaneous maturation. The protein kinase A inhibitor rp-cAMP induced meiotic maturation in the presence of 4 mmol hypoxanthine l(-1), an effect that was additive to the maturation-promoting effect of FSH and meiosis-activating sterol, indicating that induced maturation also uses the cAMP-protein kinase A-dependent signal transduction pathway. In conclusion, induced and spontaneous maturation of mouse oocytes appear to use different signal transduction pathways.

The Effects of Butyric Acid on the Differentiation, Proliferation, Apoptosis, and Autophagy of IPEC-J2 Cells

2020 ◽  
Vol 20 (4) ◽  
pp. 307-317
Author(s):  
Yuan Yang ◽  
Jin Huang ◽  
Jianzhong Li ◽  
Huansheng Yang ◽  
Yulong Yin
Keyword(s):  
Cell Viability ◽  
P38 Mapk ◽  
Butyric Acid ◽  
Cell Apoptosis ◽  
Mapk Pathway ◽  
Mrna Level ◽  
Dependent Manner ◽  
M Phase ◽  
High Concentrations ◽  
Underlying Mechanisms

Background: Butyric acid (BT), a short-chain fatty acid, is the preferred colonocyte energy source. The effects of BT on the differentiation, proliferation, and apoptosis of small intestinal epithelial cells of piglets and its underlying mechanisms have not been fully elucidated. Methods: In this study, it was found that 0.2-0.4 mM BT promoted the differentiation of procine jejunal epithelial (IPEC-J2) cells. BT at 0.5 mM or higher concentrations significantly impaired cell viability in a dose- and time-dependent manner. In addition, BT at high concentrations inhibited the IPEC-J2 cell proliferation and induced cell cycle arrest in the G2/M phase. Results: Our results demonstrated that BT triggered IPEC-J2 cell apoptosis via the caspase8-caspase3 pathway accompanied by excess reactive oxygen species (ROS) and TNF-α production. BT at high concentrations inhibited cell autophagy associated with increased lysosome formation. It was found that BT-reduced IPEC-J2 cell viability could be attenuated by p38 MAPK inhibitor SB202190. Moreover, SB202190 attenuated BT-increased p38 MAPK target DDIT3 mRNA level and V-ATPase mRNA level that were responsible for normal acidic lysosomes. Conclusion: In conclusion, 1) at 0.2-0.4 mM, BT promotes the differentiation of IPEC-J2 cells; 2) BT at 0.5 mM or higher concentrations induces cell apoptosis via the p38 MAPK pathway; 3) BT inhibits cells autophagy and promotes lysosome formation at high concentrations.

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