Autophagy is a starvation-induced transport pathway delivering parts of the cytosol into the lysosome (vacuole) for degradation. Autophagy significantly differs from other transport pathways by using double membrane layered transport intermediates. Based on the identification of autophagy genes in Saccharomyces cerevisiae, which served as a pacemaker for higher cells, our mechanistic knowledge of autophagy notably increased over the past few years. We here identify AUT8 as a novel gene essential for autophagy by screening a collection of approximately 5000 yeast deletion strains, each containing a defined deletion in an individual gene. This collection is a result of the world-wide Saccharomyces deletion project and covers the non-essential genes of the whole yeast genome. Homozygous aut8 Delta cells are impaired in maturation of proaminopeptidase I, and they fail to undergo the cell differentiation process of sporulation. The essential function of AUT8 for autophagy is further demonstrated by the lack of accumulation of autophagic vesicles in the vacuoles of aut8 Delta cells starved of nitrogen in the presence of the proteinase B inhibitor phenylmethylsulfonyl fluoride.