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> Stages de M1 > Stages proposés pour l’année 2014-2015 > Consequences of de novo centriole assembly on mouse oocyte meiotic (...)

Consequences of de novo centriole assembly on mouse oocyte meiotic divisions

proposé par Marie-Hélène Verlhac , CIRB, UMR7241/INSERM-U1050, Collège de France, 75005 Paris

Projet de stage :

Oocytes from higher eukaryotes undergo asymmetric divisions in size, leading to the formation of a large cell, the oocyte and two small ones, the polar bodies. This allows most of the maternal stores to be retained in the oocyte, a vital condition for further embryo development. We study meiotic spindle morphogenesis and positioning, key events ensuring the asymmetry of meiotic divisions in oocytes. In somatic cells, the spindle axis is determined by the position of the two opposing centrosomes before entry into mitosis. In these cells, movement and orientation of the spindle are in close relationship with the polarity axis of the mother cell, and occurs via astral microtubules emanating from the centrosomes located at spindle poles. However, prophase oocytes do not have a pre-defined polarity, and meiotic spindles organize in the absence of canonical centrosomes, thus in absence of their associated astral microtubules. Spindles are highly dynamic structures assembling around chromosomes to distribute them equally into daughter cells. Errors in spindle formation can lead to aneuploidies. It is therefore essential that bipolar spindle assembly occurs correctly. We want to study the consequences of adding centrioles back during oogenesis. Proteins involved in centriole duplication can also trigger de novo centriole assembly. This is notably the case of Sas4/Plk4 (Polo Like Kinase 4). In collaboration with Renata Basto lab (Curie Institute, Paris), we have produced a transgenic line, which overexpresses Plk4 conditionally during oocyte growth. We have preliminary results that show that we can indeed organize de novo assembly of centrin-positive structures and that the process of meiotic divisions is compromised in these oocytes. Using live imaging as well as super-resolution microscopy, we will analyse in greater details the defects observed in oocytes, which overexpress Plk4 during oogenesis. With this project, we can thus test the biological significance of centriole loss during mouse oogenesis.

Techniques mises en œuvre par le stagiaire :

Mouse oocyte isolation and culture ; Live Imaging ; Super-resolution microscopy

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