Publications by authors named "Marie Hélène Verlhac"

49 Publications

Myosin-X is dispensable for spindle morphogenesis and positioning in the mouse oocyte.

Development 2021 Mar 15. Epub 2021 Mar 15.

CIRB, Collège de France, UMR7241/U1050, 75005 Paris, France

Off-center spindle positioning in mammalian oocytes enables asymmetric divisions in size, important for subsequent embryogenesis. The migration of the meiosis I spindle from the oocyte center to its cortex is mediated by F-actin. Specifically, an F-actin-cage surrounds the microtubule spindle and applies forces to it. To better understand how F-actin transmits forces to the spindle, we studied a potential direct link between F-actin and microtubules. For this, we tested the implication of Myosin-X, a known F-actin and microtubule binder involved in spindle morphogenesis and/or positioning in somatic cells, amphibian oocytes and embryos. Using a mouse strain conditionally invalidated for Myosin-X in oocytes and by live cell imaging, we show that Myosin-X is not localized on the spindle and dispensable for spindle and F-actin assembly. It is not required for force transmission since spindle migration and chromosome alignment occur normally. More broadly, Myosin-X is dispensable for oocyte developmental potential and female fertility. We therefore exclude a role for Myosin-X in transmitting F-actin-mediated forces to the spindle, opening new perspectives regarding this mechanism in mouse oocytes, which differs from most mitotic cells.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1242/dev.199364DOI Listing
March 2021

A new mode of mechano-transduction shakes the oocyte nucleus, thereby fine tunes gene expression modulating the developmental potential.

C R Biol 2021 Feb 4;343(3):223-234. Epub 2021 Feb 4.

CIRB, Collège de France, Université PSL, CNRS, Inserm, Equipe Labellisée FRM, 75005 Paris, France.

Understanding the mechanism of nucleus positioning and the information conveyed by it constitute important research axes in Developmental and Reproductive Biology. In most species, the position of the oocyte nucleus predefines the axes of the future embryo. In the mouse oocyte, the nucleus is centered by a pressure gradient generated by a cytoplasmic actin meshwork nucleated by Formin 2. The discovery of this centering mechanism allowed to better understanding its biological significance. Centering the nucleus in mouse oocytes involves a novel mechano-transduction process, which promotes agitation of the nucleus and of its content, including chromatin, thereby modulating gene expression. This fine regulation of the maternal RNA stores explains why nucleus centering is predictive of the quality of the female gamete and of its developmental potential after fertilization.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.5802/crbiol.24DOI Listing
February 2021

[Cortical tension of the oocyte and euploidy: the right balance].

Med Sci (Paris) 2020 Nov 5;36(11):965-968. Epub 2020 Nov 5.

Centre interdisciplinaire de recherche en biologie (CIRB), UMR7241/U1050, Collège de France, 11 place Marcelin Berthelot, 75005 Paris, France.

View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1051/medsci/2020180DOI Listing
November 2020

Artificially decreasing cortical tension generates aneuploidy in mouse oocytes.

Nat Commun 2020 04 3;11(1):1649. Epub 2020 Apr 3.

CIRB, Collège de France, UMR7241/U1050, 75005, Paris, France.

Human and mouse oocytes' developmental potential can be predicted by their mechanical properties. Their development into blastocysts requires a specific stiffness window. In this study, we combine live-cell and computational imaging, laser ablation, and biophysical measurements to investigate how deregulation of cortex tension in the oocyte contributes to early developmental failure. We focus on extra-soft cells, the most common defect in a natural population. Using two independent tools to artificially decrease cortical tension, we show that chromosome alignment is impaired in extra-soft mouse oocytes, despite normal spindle morphogenesis and dynamics, inducing aneuploidy. The main cause is a cytoplasmic increase in myosin-II activity that could sterically hinder chromosome capture. We describe here an original mode of generation of aneuploidies that could be very common in oocytes and could contribute to the high aneuploidy rate observed during female meiosis, a leading cause of infertility and congenital disorders.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/s41467-020-15470-yDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7125192PMC
April 2020

Active diffusion in oocytes nonspecifically centers large objects during prophase I and meiosis I.

J Cell Biol 2020 03;219(3)

Center for Interdisciplinary Research in Biology, Collège de France, CNRS, INSERM, Paris Sciences et Lettres Research University, Equipe Labellisée Fondation pour la Recherche Médicale, Paris, France.

Nucleus centering in mouse oocytes results from a gradient of actin-positive vesicle activity and is essential for developmental success. Here, we analyze 3D model simulations to demonstrate how a gradient in the persistence of actin-positive vesicles can center objects of different sizes. We test model predictions by tracking the transport of exogenous passive tracers. The gradient of activity induces a centering force, akin to an effective pressure gradient, leading to the centering of oil droplets with velocities comparable to nuclear ones. Simulations and experimental measurements show that passive particles subjected to the gradient exhibit biased diffusion toward the center. Strikingly, we observe that the centering mechanism is maintained in meiosis I despite chromosome movement in the opposite direction; thus, it can counteract a process that specifically off-centers the spindle. In conclusion, our findings reconcile how common molecular players can participate in the two opposing functions of chromosome centering versus off-centering.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1083/jcb.201908195DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7054987PMC
March 2020

Active Fluctuations of the Nuclear Envelope Shape the Transcriptional Dynamics in Oocytes.

Dev Cell 2019 10 10;51(2):145-157.e10. Epub 2019 Oct 10.

CIRB, Collège de France/CNRS-UMR7241/INSERM-U1050, PSL Research University, Equipe Labellisée FRM, Paris 75005, France. Electronic address:

Nucleus position in cells can act as a developmental cue. Mammalian oocytes position their nucleus centrally using an F-actin-mediated pressure gradient. The biological significance of nucleus centering in mammalian oocytes being unknown, we sought to assess the F-actin pressure gradient effect on the nucleus. We addressed this using a dedicated computational 3D imaging approach, biophysical analyses, and a nucleus repositioning assay in mouse oocytes mutant for cytoplasmic F-actin. We found that the cytoplasmic activity, in charge of nucleus centering, shaped the nucleus while promoting nuclear envelope fluctuations and chromatin motion. Off-centered nuclei in F-actin mutant oocytes were misshaped with immobile chromatin and modulated gene expression. Restoration of F-actin in mutant oocytes rescued nucleus architecture fully and gene expression partially. Thus, the F-actin-mediated pressure gradient also modulates nucleus dynamics in oocytes. Moreover, this study supports a mechano-transduction model whereby cytoplasmic microfilaments could modulate oocyte transcriptome, essential for subsequent embryo development.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.devcel.2019.09.010DOI Listing
October 2019

A computational model of the early stages of acentriolar meiotic spindle assembly.

Mol Biol Cell 2019 03 16;30(7):863-875. Epub 2019 Jan 16.

CIRB, Collège de France, UMR7241/U1050, F-75005 Paris, France.

The mitotic spindle is an ensemble of microtubules responsible for the repartition of the chromosomal content between the two daughter cells during division. In metazoans, spindle assembly is a gradual process involving dynamic microtubules and recruitment of numerous associated proteins and motors. During mitosis, centrosomes organize and nucleate the majority of spindle microtubules. In contrast, oocytes lack canonical centrosomes but are still able to form bipolar spindles, starting from an initial ball that self-organizes in several hours. Interfering with early steps of meiotic spindle assembly can lead to erroneous chromosome segregation. Although not fully elucidated, this process is known to rely on antagonistic activities of plus end- and minus end-directed motors. We developed a model of early meiotic spindle assembly in mouse oocytes, including key factors such as microtubule dynamics and chromosome movement. We explored how the balance between plus end- and minus end-directed motors, as well as the influence of microtubule nucleation, impacts spindle morphology. In a refined model, we added spatial regulation of microtubule stability and minus-end clustering. We could reproduce the features of early stages of spindle assembly from 12 different experimental perturbations and predict eight additional perturbations. With its ability to characterize and predict chromosome individualization, this model can help deepen our understanding of spindle assembly.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1091/mbc.E18-10-0644DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6589792PMC
March 2019

Nuclear positioning as an integrator of cell fate.

Curr Opin Cell Biol 2019 02 26;56:122-129. Epub 2018 Dec 26.

CIRB, Collège de France, and CNRS-UMR7241 and INSERM-U1050, Equipe Labellisée FRM, Paris F-75005, France.

Cells are the building units of living organisms and consequently adapt to their environment by modulating their intracellular architecture, in particular the position of their nucleus. Important efforts have been made to decipher the molecular mechanisms involved in nuclear positioning. The LINC complex at the nuclear envelope is a very important part of the molecular connectivity between the cell outside and the intranuclear compartment, and thus emerged as a central player in nuclear mechanotransduction. More recent concepts in nuclear mechanotransduction came from studies involving nuclear confined migration, compression or swelling. Also, the effect of nuclear mechanosensitive properties in driving cell differentiation raises the question of nuclear mechanotransduction and gene expression and recent efforts have been done to tackle it, even though it remains difficult to address in a direct manner. Eventually, an original mechanism of nucleus positioning, mechanotransduction and regulation of gene expression in the non-adherent, non-polarized mouse oocyte, highlights the fact that nuclear positioning is an important developmental issue.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.ceb.2018.12.002DOI Listing
February 2019

Spindle Assembly: Two Spindles for Two Genomes in a Mammalian Zygote.

Curr Biol 2018 09;28(17):R948-R951

CIRB, Collège de France, and CNRS-UMR7241 and INSERM-U1050, Equipe Labellisée FRM, Paris F-75005, France.

A single bipolar spindle was thought to form around both parental genomes in zygotes initiating the first division. A recent study challenges this predominant view by showing that two independent spindles assemble to prevent parental genome mixing in mouse zygotes.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.cub.2018.07.044DOI Listing
September 2018

Separation and Loss of Centrioles From Primordidal Germ Cells To Mature Oocytes In The Mouse.

Sci Rep 2018 08 24;8(1):12791. Epub 2018 Aug 24.

Departments of Cell Biology; Obstetrics, Gynecology and Reproductive Sciences; and Bioengineering, University of Pittsburgh School of Medicine, Pittsburgh, PA, 15213, USA.

Oocytes, including from mammals, lack centrioles, but neither the mechanism by which mature eggs lose their centrioles nor the exact stage at which centrioles are destroyed during oogenesis is known. To answer questions raised by centriole disappearance during oogenesis, using a transgenic mouse expressing GFP-centrin-2 (GFP CETN2), we traced their presence from e11.5 primordial germ cells (PGCs) through oogenesis and their ultimate dissolution in mature oocytes. We show tightly coupled CETN2 doublets in PGCs, oogonia, and pre-pubertal oocytes. Beginning with follicular recruitment of incompetent germinal vesicle (GV) oocytes, through full oocyte maturation, the CETN2 doublets separate within the pericentriolar material (PCM) and a rise in single CETN2 pairs is identified, mostly at meiotic metaphase-I and -II spindle poles. Partial CETN2 foci dissolution occurs even as other centriole markers, like Cep135, a protein necessary for centriole duplication, are maintained at the PCM. Furthermore, live imaging demonstrates that the link between the two centrioles breaks as meiosis resumes and that centriole association with the PCM is progressively lost. Microtubule inhibition shows that centriole dissolution is uncoupled from microtubule dynamics. Thus, centriole doublets, present in early G2-arrested meiotic prophase oocytes, begin partial reduction during follicular recruitment and meiotic resumption, later than previously thought.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/s41598-018-31222-xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6109097PMC
August 2018

Chromosome structural anomalies due to aberrant spindle forces exerted at gene editing sites in meiosis.

J Cell Biol 2018 10 6;217(10):3416-3430. Epub 2018 Aug 6.

Collège de France, Centre for Interdisciplinary Research in Biology, UMR CNRS 7241/INSERM-U1050, Paris, France

Mouse female meiotic spindles assemble from acentriolar microtubule-organizing centers (aMTOCs) that fragment into discrete foci. These are further sorted and clustered to form spindle poles, thus providing balanced forces for faithful chromosome segregation. To assess the impact of aMTOC biogenesis on spindle assembly, we genetically induced their precocious fragmentation in mouse oocytes using conditional overexpression of Plk4, a master microtubule-organizing center regulator. Excessive microtubule nucleation from these fragmented aMTOCs accelerated spindle assembly dynamics. Prematurely formed spindles promoted the breakage of three different fragilized bivalents, generated by the presence of recombined Lox P sites. Reducing the density of microtubules significantly diminished the extent of chromosome breakage. Thus, improper spindle forces can lead to widely described yet unexplained chromosomal structural anomalies with disruptive consequences on the ability of the gamete to transmit an uncorrupted genome.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1083/jcb.201806072DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6168266PMC
October 2018

An actin shell delays oocyte chromosome capture by microtubules.

J Cell Biol 2018 08 6;217(8):2601-2603. Epub 2018 Jul 6.

Center for Interdisciplinary Research in Biology, Collège de France, CNRS-UMR7241 and INSERM-U1050, Equipe Labellisée FRM, Paris, France

The large nuclei and tiny spindles of oocytes create a challenge for chromosome capture at M-phase entry. A contractile F-actin mesh in starfish oocytes delivers chromosomes to the spindle and Burdyniuk et al. (2018. https://doi.org/10.1083/jcb.201802080) show that F-actin delays the capture of chromosomes until they are within reach of microtubules.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1083/jcb.201807016DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6080928PMC
August 2018

Active Mechanics Reveal Molecular-Scale Force Kinetics in Living Oocytes.

Biophys J 2018 04;114(7):1667-1679

Laboratoire Physico-Chimie Curie, Institut Curie, PSL Research University, Paris, France; Sorbonne Universités, UPMC Université Paris 06, Paris, France; Institute of Cell Biology, Center for Molecular Biology of Inflammation, Cells-in-Motion Cluster of Excellence, Münster University, Münster, Germany.

Active diffusion of intracellular components is emerging as an important process in cell biology. This process is mediated by complex assemblies of molecular motors and cytoskeletal filaments that drive force generation in the cytoplasm and facilitate enhanced motion. The kinetics of molecular motors have been precisely characterized in vitro by single molecule approaches, but their in vivo behavior remains elusive. Here, we study the active diffusion of vesicles in mouse oocytes, where this process plays a key role in nuclear positioning during development, and combine an experimental and theoretical framework to extract molecular-scale force kinetics (force, power stroke, and velocity) of the in vivo active process. Assuming a single dominant process, we find that the nonequilibrium activity induces rapid kicks of duration τ ∼ 300 μs resulting in an average force of F ∼ 0.4 pN on vesicles in in vivo oocytes, remarkably similar to the kinetics of in vitro myosin-V. Our results reveal that measuring in vivo active fluctuations allows extraction of the molecular-scale activity in agreement with single-molecule studies and demonstrates a mesoscopic framework to access force kinetics.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.bpj.2018.02.009DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5954280PMC
April 2018

Shifting meiotic to mitotic spindle assembly in oocytes disrupts chromosome alignment.

EMBO Rep 2018 02 12;19(2):368-381. Epub 2018 Jan 12.

Center for Interdisciplinary Research in Biology (CIRB) College de France, CNRS, INSERM, PSL Research University, Equipe labellisée FRM, Paris, France

Mitotic spindles assemble from two centrosomes, which are major microtubule-organizing centers (MTOCs) that contain centrioles. Meiotic spindles in oocytes, however, lack centrioles. In mouse oocytes, spindle microtubules are nucleated from multiple acentriolar MTOCs that are sorted and clustered prior to completion of spindle assembly in an "inside-out" mechanism, ending with establishment of the poles. We used HSET (kinesin-14) as a tool to shift meiotic spindle assembly toward a mitotic "outside-in" mode and analyzed the consequences on the fidelity of the division. We show that HSET levels must be tightly gated in meiosis I and that even slight overexpression of HSET forces spindle morphogenesis to become more mitotic-like: rapid spindle bipolarization and pole assembly coupled with focused poles. The unusual length of meiosis I is not sufficient to correct these early spindle morphogenesis defects, resulting in severe chromosome alignment abnormalities. Thus, the unique "inside-out" mechanism of meiotic spindle assembly is essential to prevent chromosomal misalignment and production of aneuploidy gametes.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.15252/embr.201745225DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5797964PMC
February 2018

Control of nucleus positioning in mouse oocytes.

Semin Cell Dev Biol 2018 10 12;82:34-40. Epub 2017 Aug 12.

CIRB, Collège de France, and CNRS-UMR7241 and INSERM-U1050, Equipe Labellisée FRM, Paris F-75005, France.

The position of the nucleus in a cell can instruct morphogenesis in some cases, conveying spatial and temporal information and abnormal nuclear positioning can lead to disease. In oocytes from worm, sea urchin, frog and some fish, nucleus position regulates embryo development, it marks the animal pole and in Drosophila it defines the future dorso-ventral axis of the embryo and of the adult body plan. However, in mammals, the oocyte nucleus is centrally located and does not instruct any future embryo axis. Yet an off-center nucleus correlates with poor outcome for mouse and human oocyte development. This is surprising since oocytes further undergo two extremely asymmetric divisions in terms of the size of the daughter cells (enabling polar body extrusion), requiring an off-centering of their chromosomes. In this review we address not only the bio-physical mechanism controlling nucleus positioning via an actin-mediated pressure gradient, but we also speculate on potential biological relevance of nuclear positioning in mammalian oocytes and early embryos.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.semcdb.2017.08.010DOI Listing
October 2018

Asymmetries and Symmetries in the Mouse Oocyte and Zygote.

Results Probl Cell Differ 2017 ;61:285-299

CIRB, Collège de France, CNRS-UMR7241, INSERM-U1050, Paris, 75005, France.

Mammalian oocytes grow periodically after puberty thanks to the dialogue with their niche in the follicle. This communication between somatic and germ cells promotes the accumulation, inside the oocyte, of maternal RNAs, proteins and other molecules that will sustain the two gamete divisions and early embryo development up to its implantation. In order to preserve their stock of maternal products, oocytes from all species divide twice minimizing the volume of their daughter cells to their own benefit. For this, they undergo asymmetric divisions in size where one main objective is to locate the division spindle with its chromosomes off-centred. In this chapter, we will review how this main objective is reached with an emphasis on the role of actin microfilaments in this process in mouse oocytes, the most studied example in mammals. This chapter is subdivided into three parts: I-General features of asymmetric divisions in mouse oocytes, II-Mechanism of chromosome positioning by actin in mouse oocytes and III-Switch from asymmetric to symmetric division at the oocyte-to-embryo transition.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1007/978-3-319-53150-2_13DOI Listing
July 2017

Meiotic spindle assembly and chromosome segregation in oocytes.

J Cell Biol 2016 Dec 22;215(5):611-619. Epub 2016 Nov 22.

Centre for Interdisciplinary Research in Biology, Collège de France, Centre National de la Recherche Scientifique, Institut National de la Santé et de la Recherche Médicale, PSL Research University, Paris 75006, France

Oocytes accumulate maternal stores (proteins, mRNAs, metabolites, etc.) during their growth in the ovary to support development after fertilization. To preserve this cytoplasmic maternal inheritance, they accomplish the difficult task of partitioning their cytoplasm unequally while dividing their chromosomes equally. Added to this complexity, most oocytes, for reasons still speculative, lack the major microtubule organizing centers that most cells use to assemble and position their spindles, namely canonical centrosomes. In this review, we will address recent work on the mechanisms of meiotic spindle assembly and chromosome alignment/segregation in female gametes to try to understand the origin of errors of oocyte meiotic divisions. The challenge of oocyte divisions appears indeed not trivial because in both mice and humans oocyte meiotic divisions are prone to chromosome segregation errors, a leading cause of frequent miscarriages and congenital defects.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1083/jcb.201607062DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5147004PMC
December 2016

Mother centrioles are kicked out so that starfish zygote can grow.

J Cell Biol 2016 Mar 21;212(7):759-61. Epub 2016 Mar 21.

Center for Interdisciplinary Research in Biology, Collège de France, Centre National de la Recherche Scientifique-UMR7241, and Institut National de la Santé et de la Recherche Médicale-U1050, Paris F-75005, France

Most oocytes eliminate their centrioles during meiotic divisions through unclear mechanisms. In this issue, Borrego-Pinto et al. (2016. J Cell. Biol. http://dx.doi.org/10.1083/jcb.201510083) show that mother centrioles need to be eliminated from starfish oocytes by extrusion into the polar bodies for successful embryo development.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1083/jcb.201602053DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4810309PMC
March 2016

Oocyte Maturation and Development.

F1000Res 2016 9;5. Epub 2016 Mar 9.

CIRB, Collège de France, Paris, France.

Sexual reproduction is essential for many organisms to propagate themselves. It requires the formation of haploid female and male gametes: oocytes and sperms. These specialized cells are generated through meiosis, a particular type of cell division that produces cells with recombined genomes that differ from their parental origin. In this review, we highlight the end process of female meiosis, the divisions per se, and how they can give rise to a functional female gamete preparing itself for the ensuing zygotic development. In particular, we discuss why such an essential process in the propagation of species is so poorly controlled, producing a strong percentage of abnormal female gametes in the end. Eventually, we examine aspects related to the lack of centrosomes in female oocytes, the asymmetry in size of the mammalian oocyte upon division, and in mammals the direct consequences of these long-lived cells in the ovary.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.12688/f1000research.7892.1DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4786908PMC
March 2016

F-actin mechanics control spindle centring in the mouse zygote.

Nat Commun 2016 Jan 4;7:10253. Epub 2016 Jan 4.

CIRB, Collège de France, and CNRS-UMR7241 and INSERM-U1050, Equipe labellisée Ligue contre le Cancer, Paris F-75005, France.

Mitotic spindle position relies on interactions between astral microtubules nucleated by centrosomes and a rigid cortex. Some cells, such as mouse oocytes, do not possess centrosomes and astral microtubules. These cells rely only on actin and on a soft cortex to position their spindle off-centre and undergo asymmetric divisions. While the first mouse embryonic division also occurs in the absence of centrosomes, it is symmetric and not much is known on how the spindle is positioned at the exact cell centre. Using interdisciplinary approaches, we demonstrate that zygotic spindle positioning follows a three-step process: (1) coarse centring of pronuclei relying on the dynamics of an F-actin/Myosin-Vb meshwork; (2) fine centring of the metaphase plate depending on a high cortical tension; (3) passive maintenance at the cell centre. Altogether, we show that F-actin-dependent mechanics operate the switch between asymmetric to symmetric division required at the oocyte to embryo transition.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/ncomms10253DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4725770PMC
January 2016

SKAP, an outer kinetochore protein, is required for mouse germ cell development.

Reproduction 2016 Mar 14;151(3):239-51. Epub 2015 Dec 14.

CNRSIGH (UPR1142), 141 rue de la Cardonille, 34396 Montpellier, FranceCNRSCRBM (UMR5237), 1919 route de Mende, 34293 Montpellier, FranceINRADépartement Biologie et Amélioration des Plantes, route de Saint-Cyr, 78026 Versailles, FranceCollège de FranceCIRB (UMR CNRS 7241/INSERM- U1050), 11 Place Marcelin Berthelot, 75005 Paris, FranceDepartamento de BiologíaFacultad de Ciencias, Universidad Autónoma de Madrid, 28049 Madrid, Spain

In sexually reproducing organisms, accurate gametogenesis is crucial for the transmission of genetic material from one generation to the next. This requires the faithful segregation of chromosomes during mitotic and meiotic divisions. One of the main players in this process is the kinetochore, a large multi-protein complex that forms at the interface of centromeres and microtubules. Here, we analyzed the expression profile and function of small kinetochore-associated protein (SKAP) in the mouse. We found that two distinct SKAP isoforms are specifically expressed in the germline: a smaller isoform, which is detected in spermatogonia and spermatocytes and localized in the outer mitotic and meiotic kinetochores from metaphase to telophase, and a larger isoform, which is expressed in the cytoplasm of elongating spermatids. We generated SKAP-deficient mice and found that testis size and sperm production were severely reduced in mutant males. This phenotype was partially caused by defects during spermatogonia proliferation before entry into meiosis. We conclude that mouse SKAP, while being dispensable for somatic cell divisions, has an important role in the successful outcome of male gametogenesis. In germ cells, analogous to what has been suggested in studies using immortalized cells, SKAP most likely stabilizes the interaction between kinetochores and microtubules, where it might be needed as an extra safeguard to ensure the correct segregation of mitotic and meiotic chromosomes.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1530/REP-15-0451DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4738695PMC
March 2016

Active diffusion positions the nucleus in mouse oocytes.

Nat Cell Biol 2015 Apr 16;17(4):470-9. Epub 2015 Mar 16.

CIRB, Collège de France, and CNRS-UMR7241 and INSERM-U1050, Equipe Labellisée Ligue Contre le Cancer, Paris F-75005, France.

In somatic cells, the position of the cell centroid is dictated by the centrosome. The centrosome is instrumental in nucleus positioning, the two structures being physically connected. Mouse oocytes have no centrosomes, yet harbour centrally located nuclei. We demonstrate how oocytes define their geometric centre in the absence of centrosomes. Using live imaging of oocytes, knockout for the formin 2 actin nucleator, with off-centred nuclei, together with optical trapping and modelling, we discover an unprecedented mode of nucleus positioning. We document how active diffusion of actin-coated vesicles, driven by myosin Vb, generates a pressure gradient and a propulsion force sufficient to move the oocyte nucleus. It promotes fluidization of the cytoplasm, contributing to nucleus directional movement towards the centre. Our results highlight the potential of active diffusion, a prominent source of intracellular transport, able to move large organelles such as nuclei, providing in vivo evidence of its biological function.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/ncb3131DOI Listing
April 2015

RHAMM deficiency disrupts folliculogenesis resulting in female hypofertility.

Biol Open 2015 Mar 6;4(4):562-71. Epub 2015 Mar 6.

Leibniz Institute for Age Research - Fritz Lipmann Institute, Beutenbergstrasse 11, D-07745 Jena, Germany

The postnatal mammalian ovary contains the primary follicles, each comprising an immature oocyte surrounded by a layer of somatic granulosa cells. Oocytes reach meiotic and developmental competence via folliculogenesis. During this process, the granulosa cells proliferate massively around the oocyte, form an extensive extracellular matrix (ECM) and differentiate into cumulus cells. As the ECM component hyaluronic acid (HA) is thought to form the backbone of the oocyte-granulosa cell complex, we deleted the relevant domain of the Receptor for HA Mediated Motility (RHAMM) gene in the mouse. This resulted in folliculogenesis defects and female hypofertility, although HA-induced signalling was not affected. We report that wild-type RHAMM localises at the mitotic spindle of granulosa cells, surrounding the oocyte. Deletion of the RHAMM C-terminus in vivo abolishes its spindle association, resulting in impaired spindle orientation in the dividing granulosa cells, folliculogenesis defects and subsequent female hypofertility. These data reveal the first identified physiological function for RHAMM, during oogenesis, and the importance of this spindle-associated function for female fertility.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1242/bio.201410892DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4400598PMC
March 2015

[Cortex softening: a prerequisite for the asymmetry of oocyte first division].

Med Sci (Paris) 2014 Jan 24;30(1):18-21. Epub 2014 Jan 24.

Centre interdisciplinaire de recherche en biologie, CNRS-UMR7241, Inserm-U1050, Équipe labellisée Ligue contre le cancer, Collège de France, 11, place Marcelin Berthelot, 75005 Paris, France - Memolife Laboratory of Excellence and Paris Science Lettre, Paris, F-75005, France.

View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1051/medsci/20143001005DOI Listing
January 2014

Actin-based spindle positioning: new insights from female gametes.

J Cell Sci 2014 Feb 10;127(Pt 3):477-83. Epub 2014 Jan 10.

CIRB, Collège de France, CNRS-UMR7241, INSERM-U1050, 75231 Paris, Cedex 05, France.

Asymmetric divisions are essential in metazoan development, where they promote the emergence of cell lineages. The mitotic spindle has astral microtubules that contact the cortex, which act as a sensor of cell geometry and as an integrator to orient cell division. Recent advances in live imaging revealed novel pools and roles of F-actin in somatic cells and in oocytes. In somatic cells, cytoplasmic F-actin is involved in spindle architecture and positioning. In starfish and mouse oocytes, newly discovered meshes of F-actin control chromosome gathering and spindle positioning. Because oocytes lack centrosomes and astral microtubules, F-actin networks are key players in the positioning of spindles by transmitting forces over long distances. Oocytes also achieve highly asymmetric divisions, and thus are excellent models to study the roles of these newly discovered F-actin networks in spindle positioning. Moreover, recent studies in mammalian oocytes provide a further understanding of the organisation of F-actin networks and their biophysical properties. In this Commentary, we present examples of the role of F-actin in spindle positioning and asymmetric divisions, with an emphasis on the most up-to-date studies from mammalian oocytes. We also address specific technical issues in the field, namely live imaging of F-actin networks and stress the need for interdisciplinary approaches.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1242/jcs.142711DOI Listing
February 2014

Mouse oocyte, a paradigm of cancer cell.

Cell Cycle 2013 Nov 30;12(21):3370-6. Epub 2013 Sep 30.

CIRB; Collège de France and CNRS-UMR7241 and INSERM-U1050; Paris, France; Equipe Labellisée Ligue Contre le Cancer; Paris, France; Memolife Laboratory of Excellence and Paris Science Lettre; Paris, France.

View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.4161/cc.26583DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3895426PMC
November 2013

Rebuilding MTOCs upon centriole loss during mouse oogenesis.

Dev Biol 2013 Oct 14;382(1):48-56. Epub 2013 Aug 14.

Collège de France, Center for Interdisciplinary Research in Biology, CIRB, UMR CNRS 7241/INSERM-U1050, Equipe Labellisée Ligue Contre le Cancer, 11 place Marcelin Berthelot, 75005 Paris, France; Memolife Laboratory of Excellence and Paris Science Lettre, France.

The vast majority of animal cells contain canonical centrosomes as a main microtubule-organizing center defined by a central pair of centrioles. As a rare and striking exception to this rule, vertebrate oocytes loose their centrioles at an early step of oogenesis. At the end of oogenesis, centrosomes are eventually replaced by numerous acentriolar microtubule-organizing centers (MTOCs) that shape the spindle poles during meiotic divisions. The mechanisms involved in centrosome and acentriolar MTOCs metabolism in oocytes have not been elucidated yet. In addition, little is known about microtubule organization and its impact on intracellular architecture during the oocyte growth phase following centrosome disassembly. We have investigated this question in the mouse by coupling immunofluorescence and live-imaging approaches. We show that growing oocytes contain dispersed pericentriolar material, responsible for microtubule assembly and distribution all over the cell. The gradual enlargement of PCM foci eventually leads in competent oocytes to the formation of big perinuclear MTOCs and to the assembly of large microtubule asters emanating from the close vicinity of the nucleus. Upon meiosis resumption, perinuclear MTOCs spread around the nuclear envelope, which in parallel is remodelled before breaking-down, via a MT- and dynein-dependent mechanism. Only fully competent oocytes are able to perform this dramatic reorganization at NEBD. Therefore, the MTOC-MT reorganization that we describe is one of key feature of mouse oocyte competency.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.ydbio.2013.07.029DOI Listing
October 2013

A soft cortex is essential for asymmetric spindle positioning in mouse oocytes.

Nat Cell Biol 2013 Aug 14;15(8):958-66. Epub 2013 Jul 14.

1] CIRB, Collège de France, and CNRS-UMR7241 and INSERM-U1050, Equipe Labellisée Ligue Contre le Cancer, Paris F-75005, France.

At mitosis onset, cortical tension increases and cells round up, ensuring correct spindle morphogenesis and orientation. Thus, cortical tension sets up the geometric requirements of cell division. On the contrary, cortical tension decreases during meiotic divisions in mouse oocytes, a puzzling observation because oocytes are round cells, stable in shape, that actively position their spindles. We investigated the pathway leading to reduction in cortical tension and its significance for spindle positioning. We document a previously uncharacterized Arp2/3-dependent thickening of the cortical F-actin essential for first meiotic spindle migration to the cortex. Using micropipette aspiration, we show that cortical tension decreases during meiosis I, resulting from myosin-II exclusion from the cortex, and that cortical F-actin thickening promotes cortical plasticity. These events soften and relax the cortex. They are triggered by the Mos-MAPK pathway and coordinated temporally. Artificial cortex stiffening and theoretical modelling demonstrate that a soft cortex is essential for meiotic spindle positioning.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/ncb2799DOI Listing
August 2013

Using FRET to study RanGTP gradients in live mouse oocytes.

Methods Mol Biol 2013 ;957:107-20

Institut Jacques Monod, CNRS, UMR 7592, Université Paris Diderot, Sorbonne Paris Cité, Paris, France.

Oocytes are extremely large cells that have to coordinate accurate chromosome segregation, asymmetric cytoplasm partitioning together with their own development as fertilizable gametes. For this, they undergo both global (cell cycle progression related) and local changes. It is therefore essential to be able to monitor local changes as they take place in live maturing oocytes. We describe here a method to follow RanGTP gradients using FRET technology in vivo.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1007/978-1-62703-191-2_7DOI Listing
April 2013

Error-prone mammalian female meiosis from silencing the spindle assembly checkpoint without normal interkinetochore tension.

Proc Natl Acad Sci U S A 2012 Jul 2;109(27):E1858-67. Epub 2012 May 2.

Centre Interdisciplinaire de Recherche en Biologie, Unité Mixte de Recherche-Centre National de la Recherche Scientifique 7241/Institut National de la Santé et de la Recherche Médicale U1050, Collège de France, 75005 Paris, France.

It is well established that chromosome segregation in female meiosis I (MI) is error-prone. The acentrosomal meiotic spindle poles do not have centrioles and are not anchored to the cortex via astral microtubules. By Cre recombinase-mediated removal in oocytes of the microtubule binding site of nuclear mitotic apparatus protein (NuMA), which is implicated in anchoring microtubules at poles, we determine that without functional NuMA, microtubules lose connection to MI spindle poles, resulting in highly disorganized early spindle assembly. Subsequently, very long spindles form with hyperfocused poles. The kinetochores of homologs make attachments to microtubules in these spindles but with reduced tension between them and accompanied by alignment defects. Despite this, the spindle assembly checkpoint is normally silenced and the advance to anaphase I and first polar body extrusion takes place without delay. Females without functional NuMA in oocytes are sterile, producing aneuploid eggs with altered chromosome number. These findings establish that in mammalian MI, the spindle assembly checkpoint is unable to sustain meiotic arrest in the presence of one or few misaligned and/or misattached kinetochores with reduced interkinetochore tension, thereby offering an explanation for why MI in mammals is so error-prone.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1073/pnas.1204686109DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3390881PMC
July 2012