HUMAN-IMMUNODEFICIENCY-VIRUS TYPE-1 VPU PROTEIN INDUCES DEGRADATION OF CD4 INVITRO - THE CYTOPLASMIC DOMAIN OF CD4 CONTRIBUTES TO VPU SENSITIVITY
HUMAN-IMMUNODEFICIENCY-VIRUS TYPE-1 VPU PROTEIN INDUCES DEGRADATION OF CD4 INVITRO - THE CYTOPLASMIC DOMAIN OF CD4 CONTRIBUTES TO VPU SENSITIVITY
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DOI：
10.1128/jvi.67.7.3877-3884.1993
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发表时间：
1993-07-01
影响因子：
5.4
通讯作者：
STREBEL, K
中科院分区：
文献类型：
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作者：
CHEN, MY;MALDARELLI, F;STREBEL, K
CD4 是一种完整的膜糖蛋白，作为人类免疫缺陷病毒 (HIV) 受体感染人类宿主细胞。我们最近证明，Vpu（一种 HIV 1 型 (HIV-1) 编码的整合膜磷蛋白）可诱导内质网中 CD4 的快速降解。在本报告中，我们描述了一个体外模型系统，该系统使我们能够定义 Vpu 依赖性 CD4 降解的重要参数。兔网织红细胞裂解物中 CD4 的衰变率大约是之前在 Vpu 存在的组织培养实验中观察到的值的三分之一（40 分钟与 12 分钟），并且不需要其他 HIV-1 编码的蛋白质。降解取决于测定中微粒体膜的存在以及同一膜区室中 Vpu 和 CD4 的共表达。通过体外降解试验，分析了 CD4 特定突变（包括 C 末端截短和糖基化突变体）的影响。这些实验的结果表明Vpu具有诱导糖基化以及非糖基化膜相关CD4降解的能力。 CD4 C 端 13 个氨基酸的截断并不影响 Vpu 诱导其降解的能力。然而，从 CD4 C 末端去除 32 个氨基酸就完全消除了对 Vpu 的敏感性。这表明 Vpu 靶向 CD4 胞质结构域中的特定序列以诱导其降解。我们还在体外 CD4 降解实验中分析了 Vpu 突变对其生物活性的影响。这些实验的结果表明，对于 Vpu 的这种功能至关重要的序列位于其亲水性 C 末端结构域。
CD4 is an integral membrane glycoprotein which functions as the human immunodeficiency virus (HIV) receptor for infection of human host cells. We have recently demonstrated that Vpu, an HIV type 1 (HIV-1) encoded integral membrane phosphoprotein, induces rapid degradation of CD4 in the endoplasmic reticulum. In this report, we describe an in vitro model system that allowed us to define important parameters for Vpu-dependent CD4 degradation. The rate of CD4 decay in rabbit reticulocyte lysate was approximately one-third of that observed previously in tissue culture experiments in the presence of Vpu (40 versus 12 min) and required no other HIV-1 encoded proteins. Degradation was contingent on the presence of microsomal membranes in the assay and the coexpression of Vpu and CD4 in the same membrane compartment. By using the in vitro degradation assay, the effects of specific mutations in CD4, including C-terminal truncations and glycosylation mutants, were analyzed. The results of these experiments indicate that Vpu has the capacity to induce degradation of glycosylated as well as nonglycosylated membrane-associated CD4. Truncation of 13 C-terminal amino acids of CD4 did not affect the ability of Vpu to induce its degradation. However, the removal of 32 amino acids from the C-terminus of CD4 completely abolished sensitivity to Vpu. This suggests that Vpu targets specific sequences in the cytoplasmic domain of CD4 to induce its degradation. We also analyzed the effects of mutations in Vpu on its biological activity in the in vitro CD4 degradation assay. The results of these experiments suggest that sequences critical for this function of Vpu are located in its hydrophilic C-terminal domain.