THE CYTOPLASMIC DOMAIN OF CD4 IS SUFFICIENT FOR ITS DOWN-REGULATION FROM THE CELL-SURFACE BY HUMAN-IMMUNODEFICIENCY-VIRUS TYPE-1 NEF
THE CYTOPLASMIC DOMAIN OF CD4 IS SUFFICIENT FOR ITS DOWN-REGULATION FROM THE CELL-SURFACE BY HUMAN-IMMUNODEFICIENCY-VIRUS TYPE-1 NEF
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DOI：
10.1128/jvi.68.5.3092-3101.1994
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发表时间：
1994-05-01
影响因子：
5.4
通讯作者：
GARCIA, JV
中科院分区：
文献类型：
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作者：
ANDERSON, SJ;LENBURG, M;GARCIA, JV
人类免疫缺陷病毒1型Nef下调小鼠和人类CD4的表面表达，但不下调人类CD8的表达。我们最近报道，Nef下调CD4的胞浆结构域是必需的。利用人CD8的胞外区和跨膜区与人CD4的胞内区融合而成的嵌合分子，我们在这里证明了CD4的胞浆区足以被Nef调控。由于CD4的细胞质结构域也是其与p56(Lck)相关的部位，我们使用了一系列的CD4突变体来确定Nef下调CD4细胞质结构域所需的区域是否与p56(Lck)结合所需的区域相同。我们的结果表明，Nef下调CD4所需的细胞质结构域部分与p56(Lck)的结合部位重叠，但不需要与p56(Lck)结合所必需的半胱氨酸残基。这一观察结果提出了Nef与p56(Lck)竞争结合CD4的可能性。然而，在比CD4和p56(Lck)的免疫共沉淀条件温和得多的条件下，我们没有发现Nef和CD4之间存在关联的证据。虽然表达Nef的细胞裂解物中的总CD4减少，但p56(Lck)的水平没有受到明显影响。脉冲追逐实验进一步表明，表达Nef的细胞中的CD4半衰期减少。这些结果表明，Nef诱导的表面CD4表达的减少至少部分是通过减少CD4蛋白的半衰期来实现的。这些结果还表明，在表达Nef的T细胞中，p56(Lck)的很大一部分没有CD4，这可能对T细胞功能有显著影响。
Human immunodeficiency virus type 1 Nef down-regulates surface expression of murine and human CD4 but not human CD8. We recently reported that the cytoplasmic domain of CD4 is required for its down-regulation by Nef. Using a chimeric molecule composed of the extracellular and transmembrane domains of human CD8 fused to the cytoplasmic domain of human CD4, we show here that the cytoplasmic domain of CD4 is sufficient for dean-regulation by Nef. Since the cytoplasmic domain of CD4 is also the site of its association with p56(lck), we used a series of CD4 mutants to determine whether the regions of the cytoplasmic domain of CD4 required for down-regulation by Nef are the same as those required for p56(lck) binding. Our results indicate that the portion of the cytoplasmic domain required for the down-regulation of CD4 by Nef overlaps with the binding site of p56(lck), but the cysteine residues which are essential for the association of CD4 with p56(lck) are not required. This observation raised the possibility that Nef competes with p56(lck) for binding to CD4. However, under conditions which are considerably milder than those permissive for coimmunoprecipitation of CD4 and p56(lck), we found no evidence for an association between Nef and CD4. While a decrease in total CD4 was observed in lysates of cells expressing Nef, the levels of p56(lck) were not significantly affected. Pulse-chase experiments further revealed a decrease in the half-life of CD4 in Nef-expressing cells. These results show that the decrease in surface CD4 expression induced by Nef is mediated at least in part by a decrease in the half-life of CD4 protein. These results also indicate that a large portion of p56(lck) is free of CD4 in T cells expressing Nef, which could have a significant effect on T-cell function.