MUTATIONAL ANALYSIS OF CELL-CYCLE ARREST, NUCLEAR-LOCALIZATION, AND VIRION PACKAGING OF HUMAN-IMMUNODEFICIENCY-VIRUS TYPE-1 VPR
MUTATIONAL ANALYSIS OF CELL-CYCLE ARREST, NUCLEAR-LOCALIZATION, AND VIRION PACKAGING OF HUMAN-IMMUNODEFICIENCY-VIRUS TYPE-1 VPR
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DOI：
10.1128/jvi.69.12.7909-7916.1995
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发表时间：
1995-12-01
影响因子：
5.4
通讯作者：
LANDAU, NR
中科院分区：
文献类型：
--
作者：
DIMARZIO, P;CHOE, S;LANDAU, NR
人类免疫缺陷病毒1型Vpr是病毒粒子相关的调节蛋白，是病毒在单核/巨噬细胞中有效复制所必需的。该蛋白被认为与Gag基质蛋白一起作用，允许病毒预整合复合体在未分裂的细胞中输入。在细胞中，VPR定位于细胞核。最近，我们发现VPR可以抑制p34(Cdc2)-细胞周期蛋白B的激活，使表达VPR的细胞停滞在细胞周期的G(2)/M期。在这里，我们使用一组编码在氨基末端α-螺旋区域、中心疏水区域或羧基末端碱性区域突变的VPR分子的表达载体来定义蛋白质的功能结构域。结果表明，细胞周期停滞在很大程度上受蛋白质的羧基末端碱性结构域的控制。相反，VPR的氨基末端α-螺旋区域是核定位和包装成病毒粒子所必需的。在α-螺旋区，Ala-30突变为Pro导致蛋白质定位于细胞质。令人惊讶的是，VPR与荧光素酶的融合导致了一个未能定位到细胞核的分子。此外，我们还发现，猴免疫缺陷病毒VPR，而不是VPX，能诱导G(2)期停滞。我们推测VPR有两个与细胞因子相互作用的部位：一个在指定核定位的α螺旋区域，另一个在抑制CDC2所需的羧基末端区域。
Human immunodeficiency virus type 1 Vpr is a virion-associated, regulatory protein that is required for efficient viral replication in monocytes/macrophages. The protein is believed to act in conjunction with the Gag matrix protein to allow import of the viral preintegration complex in nondividing cells. In cells, Vpr localizes to the nucleus. Recently, we showed that Vpr prevents the activation of p34(cdc2)-cyclin B. This results in arrest of Vpr-expressing cells in the G(2)/M phase of the cell cycle. Here, we use a panel of expression vectors encoding Vpr molecules mutated in the amino-terminal alpha-helical region, the central hydrophobic region, or the carboxy-terminal basic region to define the functional domains of the protein. The results showed cell cycle arrest was largely controlled by the carboxy-terminal basic domain of the protein. In contrast, the aminoterminal alpha-helical region of Vpr was required for nuclear localization and packaging into virions. The carboxy terminus appeared to be unnecessary for nuclear localization, In the alpha-helical region, mutation of Ala-30 to Pro resulted in a protein that localized to the cytoplasm. Surprisingly, fusion of Vpr to luciferase resulted in a molecule that failed to localize to the nucleus. In addition, we show that simian immunodeficiency virus Vpr, but not Vpx, induces G(2) arrest. We speculate that Vpr has two sites for interaction with cellular factors: one in the alpha-helical region that specifies nuclear localization and one in the carboxy-terminal domain that is required for Cdc2 inhibition.