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Screening for new actors implicated in Piecemeal Microautophagy of the Nucleus (PMN) in S.cerevisae

par Dawaliby, Rosie ;Mayer, Andreas
Référence USGEB annual meeting, Membranes in Motion (29-30 January 2009: Interlaken, Switzerland)
Publication Non publié, 2009-01

Poster de conférence

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Résumé :

Autophagy is a mechanism of transferring cytosolic proteins or organelles intolysosomes/vacuoles to be recycled. This recycling process is activated under starvation conditions in all eukaryotic cells or upon rapamycin treatment. Two different forms of autophagy can be observed: macro- and microautophagy. Few years ago, a novel form of autophagy had been discovered in yeast cells: Piecemeal Microautophagy of the Nucleus (PMN). PMN is driven by formation of a nucleus-vacuole (NV) junction via direct interaction of the nuclear membrane protein Nvj1 and the peripheral vacuolar membrane protein Vac8 (step I). NV junction formation is followed by the emergence of a bulge carrying a part of the nucleus and invaginating into the vacuole (step II). The bulge develops into a tear drop like bleb (step III). Finally, a vesicle carrying nuclear material is released into the vacuolar lumen (step IV) to be degraded (step V).In order to discover novel actors in the PMN process, we performed a large scale screening of the complete yeast KO collection from Euroscarf. The screen was based on the observation that the nucleolus (represented by nucleolar marker Nop1-GFP) was the preferential substrate for PMN in microscopy experiments upon rapamycin treatment. We took advantage of that observation to search in the KO collection for mutants defective in Nop1-GFP transfer to the vacuole. 318 gene deletions affected PMN to different extents, and we classified them into functional families. Here we present several complexes acting on the nuclear side, vacuolar side or implicated in lipid metabolism that showed strong inhibition of PMN. We describe at which step of PMN process these complexes are needed.