Recherche

Page web : https://www.college-de-france.fr/​site/​en-cirb/​Terret-Verlhac.htm.

1We study the last stages of murine oogenesis, a process that terminates with the production of functional female gametes required for sexual reproduction. The last step of oogenesis, named meiotic maturation, corresponds to two successive asymmetric divisions without intervening DNA replication. Mammalian meiotic maturation takes place at puberty under periodical hormonal influence. It can be reproduced and followed in vitro on synchronized population of cells. Oocytes are gigantic cells, up to 1000 times larger than most somatic cells, neuron excepted. Oocyte meiotic divisions are extremely asymmetric in size of the daughter cells, which allows the preservation of maternal stores required for embryo development. Oocytes have thus to accomplish two opposite tasks: segregate their chromosomes equally while partitioning their cytoplasm unequally. Furthermore, this tour de force is challenged by the lack of canonical centres of microtubule nucleation, namely centrosomes containing a pair of centrioles. Canonical centrosomes of mitotic cells organize spindle. During 2017/2018, we have identified mechanisms regulating the formation of meiotic spindles in the absence of canonical centrosomes, processes that might have their share in the innate susceptibility of the female gamete to produce errors in chromosome segregation.

1) Shifting meiotic to mitotic spindle assembly in oocytes disrupts chromosome alignment

2In mouse oocytes, devoid of centrioles, spindle microtubules are nucleated from multiple acentriolar MTOCs (MicroTubule Organizing Centers) 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 have shown that HSET levels must be tightly gated in meiosis I and that even a slight overexpression of HSET forces spindle morphogenesis to become more mitotic-like; which means rapid spindle bipolarization and pole assembly coupled with focused spindle 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 prevent the production of aneuploid gametes (Bennabi, 2018).

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

3We showed in the past that acentriolar MTOCs (aMTOCs) fragment into discrete foci (Łuksza, 2013). These aMTOCs are further sorted and clustered to form spindle poles, thus providing balanced forces for faithful chromosome segregation. To assess the impact of aMTOCs biogenesis on spindle assembly, we genetically induced their precocious fragmentation in mouse oocytes using conditional overexpression of Plk4, a master MTOC 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 diminished significantly 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 (Manil-Segalen, 2018).

Publications

4Manil-Ségalen M., Łuksza M., Kanaan J., Marthiens V., Lane S.I.R., Jones K.T., Terret M.-E., Basto R. et Verlhac M.-H., « Chromosome structural anomalies due to aberrant spindle forces exerted at gene editing sites in meiosis », The Journal of Cell Biology, vol. 217, n^o 10, 2018, p. 3416-3430, DOI : 10.1083/jcb.201806072.

Simerly C., Manil-Ségalen M., Castro C., Hartnett C., Kong D., Verlhac M.-H., Loncarek J. et Schatten G., « Separation and loss of centrioles from primordidal germ cells to mature oocytes in the mouse », Scientific Reports, vol. 8, n^o 1, 2018, p. 12791, DOI : 10.1038/s41598-018-31222-x.

Ahmed W.W., Fodor É., Almonacid M., Bussonnier M., Verlhac M.-H., Gov N., Visco P., van Wijland F. et Betz T., « Active mechanics reveal molecular-scale force kinetics in living oocytes », Biophysical Journal, vol. 114, n^o 7, 2018, p. 1667-1679, DOI : 10.1016/j.bpj.2018.02.009.

Bennabi I., Quéguiner I., Kolano A., Boudier T., Mailly P., Verlhac M.-H.* et Terret M.-É.*, « Shifting meiotic to mitotic spindle assembly in oocytes disrupts chromosome alignment », EMBO reports, vol. 19, n^o 2, 2018, p. 368-381, DOI : 10.15252/embr.201745225. *Co-senior authors.

Al Jord A.* et Verlhac M.-H., « Spindle assembly: Two spindles for two genomes in a mammalian zygote », Current biology, vol. 28, n^o 17, 2018, R948-R951, DOI : 10.1016/j.cub.2018.07.044. *Corresponding author.

Verlhac M.-H., « An actin shell delays oocyte chromosome capture by microtubules », The Journal of Cell Biology, vol. 217, n^o 8, 2018, p. 2601-2603, DOI : 10.1083/jcb.201807016.

Verlhac M.-H. et Terret M.-É. (dir.), Mouse Oocyte Development: Methods and Protocols, New York, Humana Press/Springer Protocols, coll. « Methods in Molecular Biology », 2018, DOI : 10.1007/978-1-4939-8603-3.

Almonacid M.*, Terret M.-É. et Verlhac M.-H., « Control of nucleus positioning in mouse oocytes », Seminars in Cell & Developmental Biology, vol. 82, 2018, p. 34-40, DOI : 10.1016/j.semcdb.2017.08.010. *Corresponding author.

Chaigne A., Terret M.-É. et Verlhac M.-H., « Asymmetries and symmetries in the mouse oocyte and zygote », Results and Problems in Cell Differentiation, vol. 61, 2017, p. 285-299, DOI : 10.1007/978-3-319-53150-2_13.

Verlhac M.-H., « An actin shell delays oocyte chromosome capture by microtubules », Journal of Cell Biology, vol. 217, n^o 8, 2018, p. 2601-2603, DOI : 10.1083/jcb.201807016.