NOVEL SYSTEM FOR MONITORING AUTOPHAGY IN THE YEAST SACCHAROMYCES-CEREVISIAE

The yeast S. cerevisiae imports cytosolic components into the vacuole non-selectively by autophagy and degrades them by vacuolar hydrolases under nutrient starvation conditions. We developed a novel system for monitoring autophagy by constructing cells in which modified vacuolar alkaline phosphatase is expressed as an inactive precursor form in the cytosol. Under starvation conditions, the processing of the precursor to the mature form and phosphatase activity appeared gradually, and t he mature form was located in the vacuole. Disruption of APG1, an esse ntial gene for autophagy, resulted in no processing or phosphatase act ivity. These results indicate that the precursor form in the cytosol i s transferred to the vacuole by autophagy and converted to the active form by vacuolar proteinases. Thus, autophagy could be determined easi ly and accurately by measuring the phosphatase activity. (C) 1995 Acad emic Press, Inc.