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> Stages de M2 > Liste des stages proposés pour l’année 2014-2015 > Cohesin protection and deprotection in mammalian oocytes

Cohesin protection and deprotection in mammalian oocytes

proposé par Katja WASSMANN , IBPS - UPMC - UMR7622 Biologie du Developpement,75005 Paris

Projet de stage :

In the first division, named meiosis I, paired chromosomes are segregated and distributed into two daughter cells, whereas in meiosis II and mitosis, sister chromatids are segregated. This meiosis-specific segregation pattern is brought about through the step-wise removal of Cohesin, a protein complex holding sister chromatis together. The M2 project will focus on cell cycle control of cohesin removal in meiosis I and II. We have recently shown that Cyclin A2/Cdk activity and PP2A inhibition trhough I2PP2A are required for sister chromatid separation in meiosis II. These roles of Cyclin A2 and I2PP2A are specific for meiosis. Localization of Cyclin A2 to kinetochores (the attachment sites for microtubules) induces sister chromatid separation. Forced localization of Cyclin A2 to kinetochores in meiosis I leads to precocious sister separation. For up to now we do not know the identity of substrates of Cyclin A2/Cdk1 at kinetochores in meiosis, and we do not know how Cyclin A2 is regulated to induce sister separation only in meiosis II and not meiosis I. We will ask whether Cyclin A2 affects I2PP2A activity for Cohesin removal in mouse oocytes. The dynamics of Cyclin A2 and I2PP2A localization will be analysed throughout meiosis by live imaging with a spinning disk confocal microscope. The project may also require the use of alternative model systems for biochemical approaches such as Xenopus laevis and ascidian oocytes, if the student is interested to pursue this direction.

Techniques mises en œuvre par le stagiaire : Mouse oocyte culture, Chromosome spreads, Immunofluorescence and Confocal microscopy of fixed samples, Live imaging (Spinning Disk Confocal, Epifluorescence), Microinjection, Cloning, mRNA production, possibility of using Xenopus and ascidian oocytes for western blots and co-immunoprecipitations.

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