Apg7p/Cvt2p is required for the cytoplasm-to-vacuole targeting, macroautophagy, and peroxisome degradation pathways
Apg7p/Cvt2p is required for the cytoplasm-to-vacuole targeting, macroautophagy, and peroxisome degradation pathways
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DOI：
10.1091/mbc.10.5.1337
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发表时间：
1999-05-01
影响因子：
3.3
通讯作者：
Klionsky, DJ
中科院分区：
文献类型：
--
作者：
Kim, J;Dalton, VM;Klionsky, DJ
细胞器的正常功能需要有效的蛋白质靶向适当的亚细胞位置。例如，真菌液泡中的降解依赖于一系列针对常驻水解酶及其底物的靶向机制。所使用的特定过程取决于可用的营养素。在饥饿条件下，大自噬是主要的方法，通过它大量的胞质溶胶被隔离到自噬囊泡（自噬体）注定这个细胞器。分子遗传学、形态学和生物化学证据表明，巨自噬与通过细胞质至液泡靶向（CVT）途径递送液泡水解酶氨肽酶I的生物合成途径具有许多相同的细胞机制。这两种途径所需的机制包括一种新的蛋白质修饰系统，涉及两种自噬蛋白Apg12p和Apg5p的缀合。缀合反应被证明依赖于Apg7p，其与泛素活化酶的Fl家族共享同源性。在这项研究中，我们证明了Apg7p在Cvt囊泡和自噬体形成的隔离步骤中发挥作用。Apg7 p融合到绿色荧光蛋白（GFP）的亚细胞定位表明Apg7 pGFP的亚群以Apg12 p依赖的方式与膜结合。亚细胞分级分离实验还表明，Apg7p池的一部分在饥饿条件下可沉淀。最后，我们证明了毕赤酵母同源Gsa7p的过氧化物酶体降解所需的功能相似的Apg7p，这表明这种新的共轭系统可能代表了一个一般的非经典的靶向机制，是跨物种的保守。
Proper functioning of organelles necessitates efficient protein targeting to the appropriate subcellular locations. For example, degradation in the fungal vacuole relies on an array of targeting mechanisms for both resident hydrolases and their substrates. The particular processes that are used vary depending on the available nutrients. Under starvation conditions, macroautophagy is the primary method by which bulk cytosol is sequestered into autophagic vesicles (autophagosomes) destined for this organelle. Molecular genetic, morphological, and biochemical evidence indicates that macroautophagy shares much of the same cellular machinery as a biosynthetic pathway for the delivery of the vacuolar hydrolase, aminopeptidase I, via the cytoplasm-to-vacuole targeting (Cvt) pathway. The machinery required in both pathways includes a novel protein modification system involving the conjugation of two autophagy proteins, Apg12p and Apg5p. The conjugation reaction was demonstrated to be dependent on Apg7p, which shares homology with the Fl family of ubiquitin-activating enzymes. In this study, we demonstrate that Apg7p functions at the sequestration step in the formation of Cvt vesicles and autophagosomes. The subcellular localization of Apg7p fused to green fluorescent protein (GFP) indicates that a subpopulation of Apg7pGFP becomes membrane associated in an Apg12p-dependent manner. Subcellular fractionation experiments also indicate that a portion of the Apg7p pool is pelletable under starvation conditions. Finally, we demonstrate that the Pichia pastoris homologue Gsa7p that is required for peroxisome degradation is functionally similar to Apg7p, indicating that this novel conjugation system may represent a general nonclassical targeting mechanism that is conserved across species.