Analysis of Bovine Leukemia Virus Gag Membrane Targeting and Late Domain Function
Analysis of Bovine Leukemia Virus Gag Membrane Targeting and Late Domain Function
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牛白血病病毒Gag膜靶向和晚期结构域功能分析
DOI：
10.1128/jvi.76.16.8485-8493.2002
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发表时间：
2002
影响因子：
5.4
通讯作者：
L. Mansky
中科院分区：
文献类型：
--
作者：
Huating Wang;Kendra M. Norris;L. Mansky
逆转录病毒样颗粒的组装只需要Gag多蛋白前体的表达。我们利用这一点开发了一个模型系统来研究牛白血病病毒（BLV）的病毒颗粒组装途径。BLV与人类t细胞白血病病毒（htlv）密切相关，都是逆转录病毒科德尔塔逆转录病毒属的成员。在哺乳动物细胞中，含有流感病毒血凝素（HA）表位标签的羧基末端BLV Gag多蛋白的过表达导致病毒样颗粒（VLPs）的大量产生。将位点定向突变引入ha标记的Gag中，以测试该模型系统在研究病毒组装途径某些方面的有效性。首先，突变破坏了对Gag肉豆蔻化很重要的氨基末端甘氨酸残基，导致VLP的产生急剧减少。可以预见，VLP生产缺陷的性质与Gag膜定位有关。其次，在缺乏病毒蛋白酶的情况下，PPPY基序（位于MA结构域）的突变大大减少了VLP的产生。在存在活性病毒蛋白酶的情况下，VLP产生的减少更为严重。通过电子显微镜检查颗粒显示，大量颗粒开始从质膜上脱落，但没有完全从细胞表面释放，这表明PPPY基序作为一个晚期结构域（L结构域）起作用。
ABSTRACT Assembly of retrovirus-like particles only requires the expression of the Gag polyprotein precursor. We have exploited this in the development of a model system for studying the virus particle assembly pathway for bovine leukemia virus (BLV). BLV is closely related to the human T-cell leukemia viruses (HTLVs), and all are members of the Deltaretrovirus genus of the Retroviridae family. Overexpression of a BLV Gag polyprotein containing a carboxy-terminal influenza virus hemagglutinin (HA) epitope tag in mammalian cells led to the robust production of virus-like particles (VLPs). Site-directed mutations were introduced into HA-tagged Gag to test the usefulness of this model system for studying certain aspects of the virus assembly pathway. First, mutations that disrupted the amino-terminal glycine residue that is important for Gag myristylation led to a drastic reduction in VLP production. Predictably, the nature of the VLP production defect was correlated to Gag membrane localization. Second, mutation of the PPPY motif (located in the MA domain) greatly reduced VLP production in the absence of the viral protease. This reduction in VLP production was more severe in the presence of an active viral protease. Examination of particles by electron microscopy revealed an abundance of particles that began to pinch off from the plasma membrane but were not completely released from the cell surface, indicating that the PPPY motif functions as a late domain (L domain).
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DOI：
10.1006/jmbi.1998.1788
发表时间：
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期刊：
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