Abstract: Monitoring Editor: Howard Riezman Autophagy is a potent intracellular degradation process with pivotal roles in health and disease. Atg8, a lipid conjugated ubiquitin-like protein, is required for the formation of autophagosomes, double-membrane vesicles responsible for the delivery of cytoplasmic material to lysosomes. How and when Atg8 functions in this process, however, is not clear. Here we show that Atg8 controls the expansion of the autophagosome precursor, the phagophore, and give the first real-time, observation-based temporal dissection of the autophagosome formation process. We demonstrate that the amount of Atg8 determines the size of autophagosomes. During autophagosome biogenesis, Atg8 forms an expanding structure and later dissociates from the site of vesicle formation. Based on the dynamics of Atg8, we present a multi-stage model of autophagosome formation. This model provides a foundation for future analyses of the functions and dynamics of known autophagy-related proteins and for screening new genes.