This study characterizes Apg8p, an essential protein for autophagy in yeast. Apg8p is transcriptionally upregulated in response to starvation and is primarily bound to membranes under both growing and starvation conditions. Immunofluorescence microscopy reveals that Apg8p localization changes drastically upon shifting cells to starvation, residing on tiny dot structures in the cytoplasm during growth and on large punctate structures during starvation, which are confirmed to be autophagosomes and autophagic bodies by immuno-EM. Apg8p is also enriched on isolation membranes and electron less-dense regions, suggesting the presence of intermediate structures during autophagosome formation. The study also shows that microtubules do not play a crucial role in autophagy in yeast, contradicting previous proposals. Additionally, the apg8 null mutant exhibits severe defects in autophagosome formation, indicating that Apg8p plays a vital role in this process. The findings provide insights into the formation of autophagosomes and suggest that Apg8p may regulate the efficiency and morphogenesis of autophagosomes.This study characterizes Apg8p, an essential protein for autophagy in yeast. Apg8p is transcriptionally upregulated in response to starvation and is primarily bound to membranes under both growing and starvation conditions. Immunofluorescence microscopy reveals that Apg8p localization changes drastically upon shifting cells to starvation, residing on tiny dot structures in the cytoplasm during growth and on large punctate structures during starvation, which are confirmed to be autophagosomes and autophagic bodies by immuno-EM. Apg8p is also enriched on isolation membranes and electron less-dense regions, suggesting the presence of intermediate structures during autophagosome formation. The study also shows that microtubules do not play a crucial role in autophagy in yeast, contradicting previous proposals. Additionally, the apg8 null mutant exhibits severe defects in autophagosome formation, indicating that Apg8p plays a vital role in this process. The findings provide insights into the formation of autophagosomes and suggest that Apg8p may regulate the efficiency and morphogenesis of autophagosomes.