The role of Atg20 in the regulation of selective autophagy pathways

• Main Authors: Ying-Jung Hsieh, 謝瑩蓉
• Other Authors: Wei-Pang Huang
• Format: Others
• Language: en_US
• Published: 2011
• Online Access: http://ndltd.ncl.edu.tw/handle/99894799481729286304

碩士 === 國立臺灣大學 === 動物學研究所 === 99 === Autophagy is an evolutionarily conserved catabolic pathway in eukaryotic cells, which mediates the turnover of cytosolic components sequestered by double membrane vesicles and delivered to the lysosome/vacuole to be degraded to maintain cellular homeostasis. Saccharomyces cerevisiae has adapted this mechanism for delivery of peroxisome by pexophagy or transport specific vacuolar hydrolases, including aminopeptidase I (Ape1), α-mannosidase (Ams1), and aspartyl aminopeptidase (Ape4), via the cytoplasm-to-vacuole targeting (Cvt) pathway. The mechanism underlying the biogenesis of the Cvt vesicle and pexophagosome are barely understood. Atg20 is a Cvt pathway-specific component with a conserved phox homology domain (PX domain) involved in binding to phosphatidylinositol 3-phosphate [PtdIns(3)P], and this binding ability is necessary for its distribution and function. Here, I show that Atg20 participates in the Cvt vesicle formation, and this process is dependent on its two coiled-coil domains. Since the Cvt pathway is blocked in atg20 mutant cells maintained in growing conditions, early-action Atg proteins, including Atg1, Atg9, and Atg11, are seen accumulated at the pre-autophagosomal structure (PAS). Detailed analyses showed that deletion of the coiled-coil domains of Atg20 abolishes the PAS localization of Atg20 and Atg24, while the interaction between Atg20 and its known interacting partner, Atg11, Atg17, and Atg24, is not affected. Furthermore, the PAS localization of Atg20 requires not only its PtdIns(3)P-binding ability but also the presence of Atg24 and Trs85, which is also required for the formation of the Cvt vesicle, indicating they may function together for the completion of the Cvt vesicle via unknown mechanism. In addition, Atg20 belongs to the sorting nexin (SNX) family, which is involved in the retrieval of proteins from the early endosomes to the late Golgi apparatus. Mutation of atg20, however, only causes partial defects in the retrieving of Snc1, suggesting that the Cvt pathway defect is not due to the disruption of the endomembrane trafficking. Moreover, I also find deletion of the coiled-coil domains of Atg20 causes pexophagy defect. Together, it is concluded that the coiled-coil domains of Atg20 are essential for the Cvt vesicle formation in cooperation with Atg24 and Trs85 under vegetative growth conditions.