Autophagosome requires specific early Sec proteins for its formation and NSF/SNARE for vacuolar fusion

Double membrane structure, autophagosome, is formed de novo in the process of autophagy in the yeast Saccharomyces cerevisiae, and many Apg, proteins participate in this process. To further understand autophagy, we analyzed t he involvement of factors engaged in the secretory pathway. First, we showe d that Sec18p (N-ethylmaleimide-sensitive fusion protein, NSF) and Vti1p (s oluble N-ethylmaleimide-sensitive fusion protein attachment protein, SNARE) , and soluble N-ethylmaleimide-sensitive fusion protein receptor are requir ed for fusion of the autophagosome to the vacuole but are not involved in a utophagosome formation. Second, Sec12p was shown to be essential for autoph agy but not for the cytoplasm to vacuole-targeting (Cvt) (pathway, which sh ares mostly the same machinery with autophagy. Subcellular fractionation an d electron microscopic analyses showed that Cvt vesicles,, but not autophag osomes, can be formed in sec12 cells. Three other coatmer protein (COPII) m utants, sec16, sec23, and sec24, were also defective in autophagy. The bloc kage of autophagy in these mutants was not dependent on transport from endo plasmic reticulum-to-Golgi, because mutations in two other COPII genes, SEC 13 and SEC31, did not affect autophagy. These results demonstrate the requi rement for subgroup of COPII proteins in autophagy. This evidence demonstra ting the involvement of Sec proteins in the mechanism of autophagosome form ation is crucial for understanding membrane flow during the process.