The TP53INP2 Protein Is Required for Autophagy in Mammalian Cells
The TP53INP2 Protein Is Required for Autophagy in Mammalian Cells
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DOI：
10.1091/mbc.e08-07-0671
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发表时间：
2009-02-01
影响因子：
3.3
通讯作者：
Iovanna, Juan Lucio
中科院分区：
文献类型：
--
作者：
Nowak, Jonathan;Archange, Cendrine;Iovanna, Juan Lucio
利用生物信息学方法，我们鉴定了一个TP 53 INP 1相关基因，该基因编码的蛋白与肿瘤蛋白53诱导的核蛋白1（TP 53 INP 1）具有30%的同一性，命名为TP 53 INP 2。TP 53 INP 1和TP 53 INP 2序列在从智人到黑腹果蝇的几个物种中发现，但在早期的真核生物和原核生物中都没有发现直系同源物。为了深入了解TP 53 INP 2蛋白的功能，我们进行了酵母双杂交筛选，显示TP 53 INP 2与LC 3相关蛋白GABARAP和GABARAP样2结合，然后我们通过免疫共沉淀证明TP 53 INP 2与这些蛋白相互作用，以及与LC 3和自噬体跨膜蛋白VMP 1相互作用。TP 53 INP 2在雷帕霉素或饥饿激活自噬后从细胞核易位到自噬体结构。此外，我们发现TP 53 INP 2的表达是自噬体发育所必需的，因为它的小干扰RNA介导的敲低强烈降低了哺乳动物细胞对自噬的敏感性。最后，我们发现TP 53 INP 2和LC 3或LC 3相关蛋白GABARAP和GABARAP样2之间的相互作用需要自噬，并通过生物发光共振能量转移测定判断由渥曼青霉素调制。我们认为TP 53 INP 2是一种支架蛋白，通过与跨膜蛋白VMP 1相互作用，将LC 3和/或LC 3相关蛋白招募到自噬体膜上。结论TP 53 INP 2是一个新的参与哺乳动物细胞自噬的基因。
Using a bioinformatic approach, we identified a TP53INP1-related gene encoding a protein with 30% identity with tumor protein 53-induced nuclear protein 1 (TP53INP1), which was named TP53INP2. TP53INP1 and TP53INP2 sequences were found in several species ranging from Homo sapiens to Drosophila melanogaster, but orthologues were found neither in earlier eukaryotes nor in prokaryotes. To gain insight into the function of the TP53INP2 protein, we carried out a yeast two-hybrid screening that showed that TP53INP2 binds to the LC3-related proteins GABARAP and GABARAP-like2, and then we demonstrated by coimmunoprecipitation that TP53INP2 interacts with these proteins, as well as with LC3 and with the autophagosome transmembrane protein VMP1. TP53INP2 translocates from the nucleus to the autophagosome structures after activation of autophagy by rapamycin or starvation. Also, we showed that TP53INP2 expression is necessary for autophagosome development because its small interfering RNA-mediated knockdown strongly decreases sensitivity of mammalian cells to autophagy. Finally, we found that interactions between TP53INP2 and LC3 or the LC3-related proteins GABARAP and GABARAP-like2 require autophagy and are modulated by wortmannin as judged by bioluminescence resonance energy transfer assays. We suggest that TP53INP2 is a scaffold protein that recruits LC3 and/or LC3-related proteins to the autophagosome membrane by interacting with the transmembrane protein VMP1. It is concluded that TP53INP2 is a novel gene involved in the autophagy of mammalian cells.