> Stages de M2 > Liste des stages proposés pour l année 2018-2019 > Comparative analysis of sperm and oocyte mitochondria composition

Comparative analysis of sperm and oocyte mitochondria composition

proposé par Vincent Galy , UMR7622 - Laboratoire de Biologie du Développement, Institut de Biologie Paris Seine, UPMC, Paris

Projet de stage : After fertilization, the autophagy machinery selectively recognizes and eliminates the sperm mitochondria. One possible mechanism underlying the recognition of the sperm mitochondria is that they expose a specific mark. In order to reveal the differences between sperm and oocytes mitochondria and identify such mark, we plan to compare their proteomes.

We would isolate and analyse pure population of mitochondria from oocytes and sperm cells. We already generated a line expressing fluorescent mitochondrial proteins in germ cells. As a first step, you will set the conditions to isolate oocytes and sperm cells respectively from feminized hermaphrodites and from males. Then, you will set-up lysis conditions to obtain entire, functional mitochondria in solution that will be purified by flow cytometry. Finally, the sperm and oocyte mitochondria proteomes will be obtained by Mass Spectrometry analysis. The comparison of both proteomes will allow to identify proteins specifically present in sperm mitochondria. These candidate proteins will then be depleted from the worms using RNAi or genome editing in order to characterize the impact of their depletion/mutation on sperm-mitochondria stability in the embryo. Your work will contribute to elucidate the molecular mechanisms of sperm mitochondria clearance and to test for the first time the functional consequences of sperm mitochondria inheritance. Approches et techniques mises en œuvre : Mitochondria purification, protein purifcation, proteomic analysis, RNAi, live imaging

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