A narrow window of cortical tension guides asymmetric spindle positioning in the mouse oocyteby A. Chaigne, C. Campillo, N. S. Gov, R. Voituriez, C. Sykes, M. H. Verlhac, M. E. Terret

Nature Communications

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Received 25 Jul 2014 | Accepted 2 Dec 2014 | Published 19 Jan 2015

A narrow window of cortical tension guides asymmetric spindle positioning in the mouse oocyte

A. Chaigne1, C. Campillo2, N.S. Gov3,*, R. Voituriez4,*, C. Sykes5,6,7, M.H. Verlhac1 & M.E. Terret1

Cell mechanics control the outcome of cell division. In mitosis, external forces applied on a stiff cortex direct spindle orientation and morphogenesis. During oocyte meiosis on the contrary, spindle positioning depends on cortex softening. How changes in cortical organization induce cortex softening has not yet been addressed. Furthermore, the range of tension that allows spindle migration remains unknown. Here, using artificial manipulation of mouse oocyte cortex as well as theoretical modelling, we show that cortical tension has to be tightly regulated to allow off-center spindle positioning: a too low or too high cortical tension both lead to unsuccessful spindle migration. We demonstrate that the decrease in cortical tension required for spindle positioning is fine-tuned by a branched F-actin network that triggers the delocalization of myosin-II from the cortex, which sheds new light on the interplay between actin network architecture and cortex tension.

DOI: 10.1038/ncomms7027 1 CIRB, Colle`ge de France, and CNRS-UMR7241 and INSERM-U1050, Equipe labellise´e Ligue contre le Cancer, Paris F-75005, France. 2 Universite´ Evry Val d’Essonne, LAMBE, Boulevard F Mitterrand, Evry 91025, France. 3 Department of Chemical Physics, Weizmann Institute of Science, Rehovot 76100, Israel. 4 UMR7600-CNRS/UPMC, 4 Place Jussieu, Paris F-75005, France. 5 CNRS-UMR168, Paris F-75248, France. 6 UPMC, 4 Place Jussieu, Paris F-75248, France. 7 Institut Curie, Centre de Recherche, Laboratoire Physico-Chimie, Paris F-75248, France. * These authors contributed equally to this work. Correspondence and requests for materials should be addressed to M.H.V. (email: marie-helene.verlhac@college-de-france.fr) or to M.E.T. (email: marieemilie.terret@college-de-france.fr).

NATURE COMMUNICATIONS | 6:6027 | DOI: 10.1038/ncomms7027 | www.nature.com/naturecommunications 1 & 2015 Macmillan Publishers Limited. All rights reserved.

D uring mitosis, forces applied on the cortex by the cell environment can control spindle orientation1,2, but the cell itself generates forces essential for its division. Indeed, at mitosis onset, before centrosome separation, cells round up3 following an increase in osmotic pressure4 and an increase in cortical tension mediated by the cell cortex3,5. This process is essential to allow proper spindle morphogenesis, stability and positioning, as it creates an optimal volume for chromosome capture and allows interaction between the cell cortex and astral microtubules emanating from the centrosomes at spindle poles in all directions of the space5–8. In mouse oocytes on the contrary, off-center positioning of the spindle following its migration, required to allow the asymmetry in size of the first meiotic division, depends on a reduced cortical tension which is correlated with both increased cortical actin nucleation and myosin-II delocalization from the cell cortex9.

Mouse oocytes are arrested in Prophase I of the first meiotic division (Meiosis I) in the ovary. Upon hormonal stimulation meiosis resumes, triggering Nuclear Envelope BreakDown (NEBD) and the two consecutive meiotic divisions. Rapidly, the first meiotic spindle forms, rarely at the perfect cell center but slightly off-centered10,11. At that stage, the oocyte bears no apparent sign of polarization12–14. The spindle then migrates along its long axis towards the closest cortex10. Mouse oocytes lacking true centrosomes and astral microtubules15, spindle positioning in female mouse meiosis does not depend on microtubules10 as in mitosis where astral microtubules connect the spindle to the cortex to actively position it within the cell8.

Spindle positioning in Meiosis I instead depends on the progressive assembly of two actin networks: a cytoplasmic meshwork nucleated by Formin-2 (refs 16–19) and Spire1/2 (ref. 20) together with a cortical actin thickening nucleated by the

Arp2/3 complex9. Mos, a MAPKKK (MAP Kinase Kinase Kinase) expressed specifically in oocytes after NEBD and activating

MAPKs around 3 h after NEBD10,21,22 controls both the nucleation of this actin thickening and the removal of myosinII from the cortex, two events responsible for the drop in cortical tension required for spindle migration9,23.

However, to which extent and why cortical tension needs to decrease in mouse oocytes to trigger spindle migration has so far remained elusive. Here using mathematical modelling and cortical tension manipulation, we show that cortical tension is strictly gated to allow spindle migration and that the decrease in cortical tension is mediated by F-actin dependent myosin-II exclusion from the cortex.

Results

Spindle position correlates with F-actin density asymmetry. To investigate how the regulation of cortical tension affects spindle movement to the cortex, we developed a theoretical model of spindle migration based on the following observations. During spindle positioning, F-actin forms an actin cage that surrounds the microtubule spindle and connects it to the thick actin cortex18,19 (Fig. 1a). At that stage, myosin-II is located at both spindle poles19,24 and its inhibition impairs spindle migration9,19.

In addition, a local stretching of the cortical actin thickening can be observed at the cortex above the approaching spindle pole9 (Figs 1a,2a). This suggests that myosin-II pulls the spindle towards the cortex from both poles by pulling on filaments emanating from the cortical thickening overlapping with filaments emanating from the actin cage. Therefore, the pulling force should depend on the overlap between cortical and cytoplasmic filaments, which should be more important on the side where the spindle is migrating, accounting for the directionality of spindle migration10,11.