The role of WWP1-Gag interaction and gag ubiquitination in assembly and release of human T-Cell leukemia virus type 1
The role of WWP1-Gag interaction and gag ubiquitination in assembly and release of human T-Cell leukemia virus type 1
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DOI：
10.1128/jvi.00642-07
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发表时间：
2007-09-01
影响因子：
5.4
通讯作者：
Derse, David
中科院分区：
文献类型：
--
作者：
Heidecker, Gisela;Lloyd, Patricia A.;Derse, David
人T细胞白血病病毒1型（HTLV-1）Gag的基质（MA）结构域中的PPPY基序与WWP 1相关，WWP 1是含HECT结构域的E3泛素连接酶家族的成员。PPPY基序的突变在早期阶段阻止了颗粒组装并消除了MA的泛素化。当Gag在缺少催化活性HECT结构域（C2 WW）的截短形式的WNW 1的存在下表达时，观察到类似的效果。为了理解泛素化在出芽中的作用，我们将MA中的四个赖氨酸突变为赖氨酸，并确定赖氨酸74为泛素化的独特位点。由K74 R突变体产生的病毒样颗粒不含泛素化的MA，并显示出感染性颗粒释放的四倍减少。此外，K74 R突变使组装对C2 WW抑制超敏感;与野生型Gag相比，K74 R Gag出芽在C2 WW的表达水平显著较低时被抑制。这一发现表明Gag和WWPI之间的相互作用是Gag泛素化以外的功能所必需的。此外，我们发现PPPY突变体Gag对野生型Gag的出芽产生强烈的显性负效应，进一步支持了募集WWP 1以实现颗粒组装的重要性。
The PPPY motif in the matrix (MA) domain of human T-cell leukemia virus type 1 (HTLV-1) Gag associates with WWP1, a member of the HECT domain containing family of E3 ubiquitin ligases. Mutation of the PPPY motif arrests particle assembly at an early stage and abolishes ubiquitination of MA. Similar effects are seen when Gag is expressed in the presence of a truncated form of WNW1 that lacks the catalytically active HECT domain (C2WW). To understand the role of ubiquitination in budding, we mutated the four lysines in MA to arginines and identified lysine 74 as the unique site of ubiquitination. Virus-like particles produced by the K74R mutant did not contain ubiquitinated MA and showed a fourfold reduction in the release of infectious particles. Furthermore, the K74R mutation rendered assembly hypersensitive to C2WW inhibition; K74R Gag budding was inhibited at significantly lower levels of expression of C2WW compared with wild-type Gag. This finding indicates that the interaction between Gag and WWPI is required for functions other than Gag ubiquitination. Additionally, we show that the PPPY- mutant Gag exerts a strong dominant-negative effect on the budding of wild-type Gag, further supporting the importance of recruitment of WWP1 to achieve particle assembly.