The mouse APG10 homologue, an E2-like enzyme for Apg12p conjugation, facilitates MAP-LC3 modification
The mouse APG10 homologue, an E2-like enzyme for Apg12p conjugation, facilitates MAP-LC3 modification
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DOI：
10.1074/jbc.m300550200
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发表时间：
2003-10-10
影响因子：
4.8
通讯作者：
Kominami, E
中科院分区：
文献类型：
--
作者：
Nemoto, T;Tanida, I;Kominami, E
自噬是溶酶体/空泡大量降解胞质区室的过程。自噬体的形成涉及膜的动态重排，其中两个泛素样修饰（Apg 12 p的缀合和MAP-LC 3的可溶形式到膜结合形式的修饰）是必需的。在酵母中，Apg 10 p是Apg 12 p结合所必需的E2样酶。分离的小鼠APG 10基因产物依赖于哺乳动物Apg 7 p（E1样酶）与哺乳动物Apg 12 p相互作用，并促进Apg 12 p结合。Apg 10 p与Apg 12 p的相互作用依赖于Apg 12 p的羧基末端甘氨酸。对小鼠Apg 10 p中预测的活性位点半胱氨酸（Cys（161））的突变分析表明，突变型Apg 10 pC 161 S可与Apg 12 p形成稳定的中间体，即使在存在Apg 7 p的情况下也可抑制Apg 12 p结合，而Apg 7 p的过表达可促进Apg 12 p-Apg 5 p结合物的形成。此外，Apg 10 p与Apg 7 p的共表达促进了MAP-LC 3的可溶性形式向膜结合形式的修饰，这是自噬所必需的第二种修饰。小鼠Apg 10 p与HEK 293细胞中的MAP-LC 3相互作用，而突变型Apg 10 pC 161 S与MAP-LC 3不形成任何中间体。Apg 10 p和MAP-LC 3之间的直接相互作用也通过酵母双杂交分析证明。突变体Apg 10 pC 161 S不能与MAP-LC 3形成任何中间体，排除了MAP-LC 3与Apg 10 p作为底物相互作用的可能性。
Autophagy is a process for the bulk degradation of cytosolic compartments by lysosomes/vacuoles. The formation of autophagosomes involves a dynamic rearrangement of the membrane for which two ubiquitinlike modifications ( the conjugation of Apg12p and the modification of a soluble form of MAP-LC3 to a membrane-bound form) are essential. In yeast, Apg10p is an E2-like enzyme essential for Apg12p conjugation. The isolated mouse APG10 gene product interacts with mammalian Apg12p dependent on mammalian Apg7p ( E1-like enzyme), and facilitates Apg12p conjugation. The interaction of Apg10p with Apg12p is dependent on the carboxyl-terminal glycine of Apg12p. Mutational analysis of the predicted active site cysteine (Cys(161)) within mouse Apg10p shows that mutant Apg10pC161S, which can form a stable intermediate with Apg12p, inhibits Apg12p conjugation even in the presence of Apg7p, while overexpression of Apg7p facilitates formation of an Apg12p-Apg5p conjugate. Furthermore, the coexpression of Apg10p with Apg7p facilitates the modification of a soluble form of MAP-LC3 to a membrane-bound form, a second modification essential for autophagy. Mouse Apg10p interacts with MAP-LC3 in HEK293 cells, while no mutant Apg10pC161S forms any intermediate with MAP-LC3. Direct interaction between Apg10p and MAP-LC3 is also demonstrated by yeast two-hybrid analysis. The inability of mutant Apg10pC161S to form any intermediate with MAP-LC3 has ruled out the possibility that MAP-LC3 interacts with Apg10p as a substrate.