Yamauchi Keiko, Wada Keiji, Tanji Kunikazu, Tanaka Makoto, Kamitani Tetsu
Department of Cardiology, The University of Texas M. D. Anderson Cancer Center, Houston, TX, USA.
FEBS J. 2008 Apr;275(7):1540-1555. doi: 10.1111/j.1742-4658.2008.06313.x. Epub 2008 Feb 25.
HIV-1 efficiently infects susceptible cells and causes AIDS in humans. Although HIV can also enter the cells of Old World monkeys, it encounters a block before reverse transcription. Data have shown that this species-specific restriction is mediated by tripartite motif (TRIM)5alpha, whose molecular function is still undefined. Here, we show that TRIM5alpha functions as a RING-finger-type E3 ubiquitin ligase both in vitro and in vivo and ubiquitinates itself in cooperation with the E2 ubiquitin-conjugating enzyme UbcH5B. In addition to the self-ubiquitination, we show that TRIM5alpha is ubiquitinated by another E3 ubiquitin ligase, Ro52, and deubiquitinated by YopJ, one of the pathogenic proteins derived from Yersinia species. Thus, the ubiquitination of TRIM5alpha is catalyzed by itself and Ro52 and downregulated by YopJ. Unexpectedly, although TRIM5alpha is ubiquitinated, our results have revealed that the proteasome inhibitors MG115 and MG132 do not stabilize it in HeLa cells, suggesting that the ubiquitination of TRIM5alpha does not lead to proteasomal degradation. Importantly, TRIM5alpha is clearly conjugated by a single ubiquitin molecule (monoubiquitination). Our monoubiquitin-fusion assay suggests that monoubiquitination is a signal for TRIM5alpha to translocate from cytoplasmic bodies to the cytoplasm.
HIV-1能有效感染易感细胞并在人类中引发艾滋病。虽然HIV也能进入旧世界猴的细胞，但在逆转录之前会遇到障碍。数据表明，这种物种特异性限制是由三联基序（TRIM）5α介导的，其分子功能仍不明确。在此，我们表明TRIM5α在体外和体内均作为一种环状结构域型E3泛素连接酶发挥作用，并与E2泛素结合酶UbcH5B协同作用使自身泛素化。除了自我泛素化，我们还表明TRIM5α被另一种E3泛素连接酶Ro52泛素化，并被源自耶尔森菌属的致病蛋白之一YopJ去泛素化。因此，TRIM5α的泛素化由其自身和Ro52催化，并被YopJ下调。出乎意料的是，尽管TRIM5α被泛素化，但我们的结果显示蛋白酶体抑制剂MG115和MG132在HeLa细胞中并不能使其稳定，这表明TRIM5α的泛素化不会导致蛋白酶体降解。重要的是，TRIM5α明显被单个泛素分子缀合（单泛素化）。我们的单泛素融合试验表明，单泛素化是TRIM5α从细胞质小体转运到细胞质的信号。
