scholarly journals RhoA- and Cdc42-induced antagonistic forces underlie symmetry breaking and spindle rotation in mouse oocytes

PLoS Biology ◽  
2021 ◽  
Vol 19 (9) ◽  
pp. e3001376
Author(s):  
Benoit Dehapiot ◽  
Raphaël Clément ◽  
Bourdais Anne ◽  
Virginie Carrière ◽  
Huet Sébastien ◽  
...  
Keyword(s):  
Symmetry Breaking ◽  
Polar Body ◽  
Break Symmetry ◽  
Ring Closure ◽  
Rotation Direction ◽  
Mouse Oocytes ◽  
Polar Bodies ◽  
Chromatid Segregation ◽  
Spindle Rotation ◽  
Dependent Polarization

Mammalian oocyte meiotic divisions are highly asymmetric and produce a large haploid gamete and 2 small polar bodies. This relies on the ability of the cell to break symmetry and position its spindle close to the cortex before anaphase occurs. In metaphase II–arrested mouse oocytes, the spindle is actively maintained close and parallel to the cortex, until fertilization triggers sister chromatid segregation and the rotation of the spindle. The latter must indeed reorient perpendicular to the cortex to enable cytokinesis ring closure at the base of the polar body. However, the mechanisms underlying symmetry breaking and spindle rotation have remained elusive. In this study, we show that spindle rotation results from 2 antagonistic forces. First, an inward contraction of the cytokinesis furrow dependent on RhoA signaling, and second, an outward attraction exerted on both sets of chromatids by a Ran/Cdc42-dependent polarization of the actomyosin cortex. By combining live segmentation and tracking with numerical modeling, we demonstrate that this configuration becomes unstable as the ingression progresses. This leads to spontaneous symmetry breaking, which implies that neither the rotation direction nor the set of chromatids that eventually gets discarded are biologically predetermined.

scholarly journals RhoA- and Ran-induced antagonistic forces underlie symmetry breaking and spindle rotation in mouse oocytes

2020 ◽  
Author(s):  
Benoit Dehapiot ◽  
Raphaël Clément ◽  
Anne Bourdais ◽  
Sébastien Huet ◽  
Guillaume Halet
Keyword(s):  
Symmetry Breaking ◽  
Polar Body ◽  
Break Symmetry ◽  
Ring Closure ◽  
Rotation Direction ◽  
Mouse Oocytes ◽  
Polar Bodies ◽  
Spindle Rotation ◽  
Dependent Polarization ◽  
Rhoa Signaling

AbstractMammalian oocyte meiotic divisions are highly asymmetric and produce a large haploid gamete and two small polar bodies. This relies on the ability of the cell to break symmetry and position its spindle close to the cortex before the anaphase occurs. In metaphase II arrested mouse oocytes, the spindle is actively maintained close and parallel to the cortex, until the fertilization triggers the sister chromatids segregation and the rotation of the spindle. The latter must indeed reorient perpendicular to the cortex to enable the cytokinesis ring closure at the base of the polar body. However, the mechanisms underlying symmetry breaking and spindle rotation have remained elusive. In this study, we show that the spindle rotation results from two antagonistic forces. First, an inward contraction of the cytokinesis furrow dependent on RhoA signaling and second, an outward attraction exerted on both lots of chromatids by a RanGTP dependent polarization of the actomyosin cortex. By combining live segmentation and tracking with numerical modelling, we demonstrate that this configuration becomes unstable as the ingression progresses. This leads to spontaneous symmetry breaking, which implies that neither the rotation direction nor the lot of chromatids that eventually gets discarded are biologically predetermined.

Inhibition of protein kinases by 6-dimethylaminopurine accelerates the transition to interphase in activated mouse oocytes

1993 ◽  
Vol 104 (3) ◽  
pp. 861-872 ◽  
Author(s):  
M.S. Szollosi ◽  
J.Z. Kubiak ◽  
P. Debey ◽  
H. de Pennart ◽  
D. Szollosi ◽  
...  
Keyword(s):  
Protein Phosphorylation ◽  
Kinase Inhibitor ◽  
Polar Body ◽  
Histone H1 ◽  
Sperm Chromatin ◽  
Oocyte Activation ◽  
Mouse Oocytes ◽  
Spindle Rotation ◽  
Polar Body Extrusion ◽  
Pronuclear Formation

Mouse oocyte activation is followed by a peculiar period during which the interphase network of microtubules does not form and the chromosomes remain condensed despite the inactivation of MPF. To evaluate the role of protein phosphorylation during this period, we studied the effects of the protein kinase inhibitor 6-dimethylaminopurine (6-DMAP) on fertilization and/or parthenogenetic activation of metaphase II-arrested mouse oocytes. 6-DMAP by itself does not induce the inactivation of histone H1 kinase in metaphase II-arrested oocytes, and does not influence the dynamics of histone H1 kinase inactivation during oocyte activation. However, 6-DMAP inhibits protein phosphorylation after oocyte activation. In addition, the phosphorylated form of some proteins disappear earlier in oocytes activated in the presence of 6-DMAP than in the activated control oocytes. This is correlated with the acceleration of some post-fertilization morphological events, such as sperm chromatin decondensation and its transient recondensation, formation of the interphase network of microtubules and pronuclear formation. In addition, numerous abnormalities could be observed: (1) the spindle rotation and polar body extrusion are inhibited; (2) the exchange of protamines into histones seems to be impaired, as judged by the morphology of DNA fibrils by electron microscopy; (3) the formation of a new nuclear envelope around the sperm chromatin proceeds prematurely, while recondensation is not yet completed. These observations suggest that the 6-DMAP-sensitive kinase(s) is (are) involved in the control of post-fertilization events such as the formation of the interphase network of microtubules, the remodelling of sperm chromatin and pronucleus formation.

scholarly journals Specific deletion of Cdc42 does not affect meiotic spindle organization/migration and homologous chromosome segregation but disrupts polarity establishment and cytokinesis in mouse oocytes

2013 ◽  
Vol 24 (24) ◽  
pp. 3832-3841 ◽  
Author(s):  
Zhen-Bo Wang ◽  
Zong-Zhe Jiang ◽  
Qing-Hua Zhang ◽  
Meng-Wen Hu ◽  
Lin Huang ◽  
...  
Keyword(s):  
Chromosome Segregation ◽  
Homologous Chromosome ◽  
Polar Body ◽  
Conditional Knockout ◽  
Meiotic Spindle ◽  
Spindle Positioning ◽  
Mouse Oocytes ◽  
Polar Bodies ◽  
And Migration ◽  
Specific Deletion

Mammalian oocyte maturation is distinguished by highly asymmetric meiotic divisions during which a haploid female gamete is produced and almost all the cytoplasm is maintained in the egg for embryo development. Actin-dependent meiosis I spindle positioning to the cortex induces the formation of a polarized actin cap and oocyte polarity, and it determines asymmetric divisions resulting in two polar bodies. Here we investigate the functions of Cdc42 in oocyte meiotic maturation by oocyte-specific deletion of Cdc42 through Cre-loxP conditional knockout technology. We find that Cdc42 deletion causes female infertility in mice. Cdc42 deletion has little effect on meiotic spindle organization and migration to the cortex but inhibits polar body emission, although homologous chromosome segregation occurs. The failure of cytokinesis is due to the loss of polarized Arp2/3 accumulation and actin cap formation; thus the defective contract ring. In addition, we correlate active Cdc42 dynamics with its function during polar body emission and find a relationship between Cdc42 and polarity, as well as polar body emission, in mouse oocytes.

In vitro fertilization and cleavage of mouse oocytes recombined with the first polar body

2008 ◽  
Vol 5 (2) ◽  
pp. 169-173 ◽  
Author(s):  
Wang Gong-Jin ◽  
Tan Xiao-Dong ◽  
Zhou Xiao-Long ◽  
Xu Xiao-Bo ◽  
Fan Bi-Qin
Keyword(s):  
Polar Body ◽  
Farm Animals ◽  
Blue Staining ◽  
Vitro Fertilization ◽  
Mouse Oocytes ◽  
Polar Bodies ◽  
First Polar Body ◽  
Different Temperatures ◽  
Further Development

AbstractThe developmental functions of oocytes of three strains of mice (Kunming, ICR and C57BL/6-Tg(CAG-EGFP)C14-Y01-FM131Osb) recombined with the nuclei of first polar bodies (Pbs I) were explored. Cumulus oocyte complexes (COCs) from the mice were collected after superovulation, then Pbs I were obtained from the COCs by 2% pronase treatment. The survival of Pbs I under different temperatures was identified by morphology and trypan blue staining. Later, the polar body I (Pb 1) nucleus and a little cytoplasm was injected into each oocyte, the nuclei of which had been enucleated by micromanipulation. Oocytes recombined with Pbs I were fertilized, then cultured in vitro in order to observe their further development. The results showed that the vigour of Pbs I was maintained for 12–14 h after superovulation, and was still maintained after 48 h at 4 °C. A total of 13 out of 117 recombined oocytes from Kunming and ICR mice, as well as 3 out of 38 recombined oocytes from C57BL/6-Tg(CAG-EGFP)C14-Y01-FM131Osb mice, developed into two-cell embryos. The experiments confirmed that mouse oocytes recombined with the nuclei of Pbs I could maintain fertilization and development. These results present valuable references for further utilization of genetic resources for farm animals

scholarly journals PAR-1 and the microtubule-associated proteins CLASP2 and dynactin-p50 have specific localisation on mouse meiotic and first mitotic spindles

Reproduction ◽  
2005 ◽  
Vol 130 (3) ◽  
pp. 311-320 ◽  
Author(s):  
Catherine A Moore ◽  
Magdalena Zernicka-Goetz
Keyword(s):  
Polar Body ◽  
Model Systems ◽  
Regulatory Proteins ◽  
Meiotic Spindle ◽  
Mitotic Spindles ◽  
Microtubule Associated Proteins ◽  
Mouse Oocytes ◽  
Polar Bodies ◽  
Associated Proteins ◽  
Second Polar Body

The site of second meiotic division, marked by the second polar body, is an important reference point in the early mouse embryo. To study its formation, we look at the highly asymmetric meiotic divisions. For extrusion of the small polar bodies during meiosis, the spindles must be located cortically. The positioning of meiotic spindles is known to involve the actin cytoskeleton, but whether microtubules are also involved is not clear. In this study we investigated the patterns of localisation of microtubule regulatory proteins in mouse oocytes. PAR-1 is a member of the PAR (partitioning-defective) family with known roles in regulation of microtubule stability and spindle positioning in other model systems. Here we show its specific localisation on mouse meiotic and first mitotic spindles. In addition, the microtubule-associated proteins CLASP2 (a CLIP associating protein) and dynactin-p50 are found on kinetochores and a subset of microtubule-organising centres. Thus we show specific localisation of microtubule regulatory proteins in mouse oocytes, which could indicate roles in meiotic spindle organisation.

scholarly journals Ral GTPase is essential for actin dynamics and Golgi apparatus distribution in mouse oocyte maturation

Cell Division ◽  
2021 ◽  
Vol 16 (1) ◽  
Author(s):  
Ming-Hong Sun ◽  
Lin-Lin Hu ◽  
Chao-Ying Zhao ◽  
Xiang Lu ◽  
Yan-Ping Ren ◽  
...  
Keyword(s):  
Golgi Apparatus ◽  
Oocyte Maturation ◽  
Polar Body ◽  
Mouse Oocyte ◽  
Actin Dynamics ◽  
Mouse Oocytes ◽  
Oocyte Meiosis ◽  
Ral Gtpase ◽  
Polar Body Extrusion ◽  
Cytoskeleton Dynamics

Abstract Background Ral family is a member of Ras-like GTPase superfamily, which includes RalA and RalB. RalA/B play important roles in many cell biological functions, including cytoskeleton dynamics, cell division, membrane transport, gene expression and signal transduction. However, whether RalA/B involve into the mammalian oocyte meiosis is still unclear. This study aimed to explore the roles of RalA/B during mouse oocyte maturation. Results Our results showed that RalA/B expressed at all stages of oocyte maturation, and they were enriched at the spindle periphery area after meiosis resumption. The injection of RalA/B siRNAs into the oocytes significantly disturbed the polar body extrusion, indicating the essential roles of RalA/B for oocyte maturation. We observed that in the RalA/B knockdown oocytes the actin filament fluorescence intensity was significantly increased at the both cortex and cytoplasm, and the chromosomes were failed to locate near the cortex, indicating that RalA/B regulate actin dynamics for spindle migration in mouse oocytes. Moreover, we also found that the Golgi apparatus distribution at the spindle periphery was disturbed after RalA/B depletion. Conclusions In summary, our results indicated that RalA/B affect actin dynamics for chromosome positioning and Golgi apparatus distribution in mouse oocytes.

Triphenyltin chloride induces spindle microtubule depolymerisation and inhibits meiotic maturation in mouse oocytes

2014 ◽  
Vol 26 (8) ◽  
pp. 1084 ◽  
Author(s):  
Yu-Ting Shen ◽  
Yue-Qiang Song ◽  
Xiao-Qin He ◽  
Fei Zhang ◽  
Xin Huang ◽  
...  
Keyword(s):  
Polar Body ◽  
Meiotic Maturation ◽  
Dependent Manner ◽  
Germinal Vesicle Breakdown ◽  
Mouse Oocytes ◽  
First Polar Body ◽  
Triphenyltin Chloride ◽  
Dose Dependent

Meiosis produces haploid gametes for sexual reproduction. Triphenyltin chloride (TPTCL) is a highly bioaccumulated and toxic environmental oestrogen; however, its effect on oocyte meiosis remains unknown. We examined the effect of TPTCL on mouse oocyte meiotic maturation in vitro and in vivo. In vitro, TPTCL inhibited germinal vesicle breakdown (GVBD) and first polar body extrusion (PBE) in a dose-dependent manner. The spindle microtubules completely disassembled and the chromosomes condensed after oocytes were exposed to 5 or 10 μg mL–1 TPTCL. γ-Tubulin protein was abnormally localised near chromosomes rather than on the spindle poles. In vivo, mice received TPTCL by oral gavage for 10 days. The general condition of the mice deteriorated and the ovary coefficient was reduced (P < 0.05). The number of secondary and mature ovarian follicles was significantly reduced by 10 mg kg–1 TPTCL (P < 0.05). GVBD decreased in a non-significant, dose-dependent manner (P > 0.05). PBE was inhibited with 10 mg kg–1 TPTCL (P < 0.05). The spindles of in vitro and in vivo metaphase II oocytes were disassembled with 10 mg kg–1 TPTCL. These results suggest that TPTCL seriously affects meiotic maturation by disturbing cell-cycle progression, disturbing the microtubule cytoskeleton and inhibiting follicle development in mouse oocytes.

scholarly journals Polarized Cdc42 activation promotes polar body protrusion and asymmetric division in mouse oocytes

2013 ◽  
Vol 377 (1) ◽  
pp. 202-212 ◽  
Author(s):  
Benoit Dehapiot ◽  
Virginie Carrière ◽  
John Carroll ◽  
Guillaume Halet
Keyword(s):  
Polar Body ◽  
Asymmetric Division ◽  
Mouse Oocytes

scholarly journals Kinetochore Fibers Are Not Involved in the Formation of the First Meiotic Spindle in Mouse Oocytes, but Control the Exit from the First Meiotic M Phase

1999 ◽  
Vol 146 (1) ◽  
pp. 1-12 ◽  
Author(s):  
Stéphane Brunet ◽  
Angélica Santa Maria ◽  
Philippe Guillaud ◽  
Denis Dujardin ◽  
Jacek Z. Kubiak ◽  
...  
Keyword(s):  
Polar Body ◽  
Meiotic Spindle ◽  
Homologous Chromosomes ◽  
Mouse Oocytes ◽  
Sister Chromatids ◽  
M Phase ◽  
Continuous Presence ◽  
Set Up ◽  
Polar Body Extrusion ◽  
Reductional Division

During meiosis, two successive divisions occur without any intermediate S phase to produce haploid gametes. The first meiotic division is unique in that homologous chromosomes are segregated while the cohesion between sister chromatids is maintained, resulting in a reductional division. Moreover, the duration of the first meiotic M phase is usually prolonged when compared with mitotic M phases lasting 8 h in mouse oocytes. We investigated the spindle assembly pathway and its role in the progression of the first meiotic M phase in mouse oocytes. During the first 4 h, a bipolar spindle forms and the chromosomes congress near the equatorial plane of the spindle without stable kinetochore– microtubule end interactions. This late prometaphase spindle is then maintained for 4 h with chromosomes oscillating in the central region of the spindle. The kinetochore–microtubule end interactions are set up at the end of the first meiotic M phase (8 h after entry into M phase). This event allows the final alignment of the chromosomes and exit from metaphase. The continuous presence of the prometaphase spindle is not required for progression of the first meiotic M phase. Finally, the ability of kinetochores to interact with microtubules is acquired at the end of the first meiotic M phase and determines the timing of polar body extrusion.

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.

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