SAC proteins in oocyte meiosis.

proposé par Katja WASSMANN, UMR7622 Biologie du Développement, UPMC, 9 quai Saint Bernard, 75005 Paris.

Projet de stage :

This M2 project will focus on SAC proteins in oocyte meiosis. 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.We want to establish how monopolar attachment is recognized as correct in meiosis I, and incorrect in meiosis II. We will generate oocytes that enter meiosis II with non-separated homologous chromosomes, and through live imaging we will ask how these bivalent chromosomes are segregated and whether the checkpoint can recognize these bivalents as wrong in meiosis II. We will ask whether and how the SAC promotes monopolar attachment and centromeric cohesin protection in meiosis I, but not meiosis II. It has been shown that Histone modifications such as H3 phosphorylation by Haspin, and Histone H2A phosphorylation by the checkpoint kinase Bub1 are required for recruitment of proteins required for error correction, and for centromere localization of the cohesin protector Sgo1 in mitosis [5]. Our unpublished data indicate that the phosphorylation events required for microtubule attachment and localization of the meiotic cohesin protector Sgo2 are different in meiosis I and II compared to mitosis. Our work aims at elucidating those differences and the molecular circuits at work at the centromere and kinetochore in meiosis, to understand how the specific meiotic segregation pattern is brought about [6, 7]. A combination of conditional knock-out mouse models and specific inhibitors are at our disposition to dissect the molecular mechanisms of checkpoint control, centromeric cohesin protection, and error correction during both meiotic division, and this M2 project will address how one component of the SAC functions in Cohesin protection and proper orientation of chromosomes in meiosis.

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, western blots and co-immunoprecipitations.

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