Servizio Comunitario di Informazione in materia di Ricerca e Sviluppo - CORDIS


MMAF Sintesi della relazione

Project ID: 260304
Finanziato nell'ambito di: FP7-IDEAS-ERC
Paese: Austria

Final Report Summary - MMAF (Molecular mechanisms of autophagosome formation)

Autophagosomes are double membrane-bound organelles that are formed de novo during a process called autophagy. Autophagosomes mediate the bulk degradation of cytoplasmic material such as aggregated proteins, dysfunctional or surplus mitochondria and intracellular pathogens. Autophagy is conserved from yeast to human and has been shown to protect the organism from conditions such as starvation, neurodegeneration and infectious diseases. During autophagosome formation initially small, membrane structures termed isolation membranes are formed. These isolation membranes expand and thereby gradually enclose cytoplasmic cargo. Finally, isolation membranes close to give rise to mature autophagosomes. After their formation autophagosomes fuse with lysosomes or vacuoles in yeast within which their inner membrane and the content are degraded. Despite their importance little is known about how cells generate autophagosomes and how cargo is specifically captured. In order to gain insight into how the cellular machinery generates autophagosomes we reconstitute crucial steps in vitro.
In particular, we studied the conserved Atg5-Atg12/Atg16 complex that is essential for autophagosome formation. We showed that the yeast Atg5-Atg12/Atg16 complex directly binds membranes. Membrane binding is mediated by Atg5, inhibited by Atg12 and activated by Atg16. In a fully reconstituted system using giant unilamellar vesicles and recombinant proteins we revealed that all components of the complex are required for efficient promotion of Atg8 conjugation to phosphatidylethanolamine and were able to assign precise functions to all of its components during this process. In addition, we found that in vitro the Atg5-Atg12/Atg16 complex is able to tether membranes independently of Atg8. Furthermore, we showed that membrane binding by Atg5 is downstream of its recruitment to the pre-autophagosomal structure but is essential for autophagy and cytoplasm-to-vacuole transport at a stage preceding Atg8 conjugation and vesicle closure. Our findings provide important insights into the mechanism of action of the Atg5-Atg12/Atg16 complex during autophagosome formation. We also established a role for Atg8 family proteins for the shaping of the isolation membrane during selective autophagy.

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