Human immunodeficiency virus type 1 nef activates p21-activated kinase via recruitment into lipid rafts
Human immunodeficiency virus type 1 nef activates p21-activated kinase via recruitment into lipid rafts
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DOI：
10.1128/jvi.78.8.4085-4097.2004
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发表时间：
2004-04-01
影响因子：
5.4
通讯作者：
Fackler, OT
中科院分区：
文献类型：
--
作者：
Krautkrämer, E;Giese, SI;Fackler, OT
人类免疫缺陷病毒1型的Nef蛋白是艾滋病发病的一个重要因素。除了从细胞表面下调CD4和主要组织相容性复合体I类分子，以及增加病毒粒子的传染性外，Nef还触发t细胞受体（TCR）级联激活，促进病毒传播。已经确定了Nef诱导的信号通路；然而，目前尚不清楚Nef是如何以及在哪个亚细胞区室触发信号的。Nef招募一种多蛋白复合物来激活介导下游效应功能的细胞Pak激酶。由于Nef的一个亚群存在于洗涤剂不溶性微结构域（脂筏）中，生理TCR信号从那里开始，我们测试了脂筏是否有助于Nef介导的Pak激活。在浮选分析中，nef相关的Pak活性仅与脂筏分馏。在Nef存在下，Pak的激活与激酶的脂质筏募集相一致，否则它被排除在洗涤剂不溶性微结构域之外。脂筏的实验增溶干扰了Pak活性与Nef的关联。为了更严格地分析筏定位对Nef函数的重要性，我们生成了一个棕榈酰化的Nef （PalmNef）。棕榈Nef在脂筏中高度富集，Pak活性水平显著高于Nef。值得注意的是，Pak被其生理激活剂Cdc42和Rac激活也发生在脂筏中，并且需要脂筏的完整性。综上所述，这些数据表明Nef通过招募包括Pak在内的信号机制进入脂筏诱导信号转导，从而模仿生理细胞机制启动TCR级联。
The Nef protein of human immunodeficiency virus type 1 is an important factor in AIDS pathogenesis. In addition to downregulating CD4 and major histocompatibility complex class I molecules from the cell surface, as well as increasing virion infectivity, Nef triggers activation of the T-cell receptor (TCR) cascade to facilitate virus spread. Signaling pathways that are induced by Nef have been identified; however, it is unclear how and in which subcellular compartment Nef triggers signaling. Nef recruits a multiprotein complex to activate the cellular Pak kinase that mediates downstream effector functions. Since a subpopulation of Nef is present in detergent-insoluble microdomains (lipid rafts) from where physiological TCR signaling is initiated, we tested whether lipid rafts are instrumental for Nef-mediated Pak activation. In flotation analysis, Nef-associated Pak activity exclusively fractionated with lipid rafts. Activation of Pak in the presence of Nef coincided with lipid raft recruitment of the kinase, which was otherwise excluded from detergent-insoluble microdomains. Experimental solubilization of lipid rafts interfered with the association of Pak activity with Nef. To analyze the importance of the raft localization for Nef function more rigorously, we generated a palmitoylated Nef (PalmNef). PalmNef was highly enriched in lipid rafts and associated with significantly higher levels of Pak activity than Nef. Notably, activation of Pak by its physiological activators, Cdc42 and Rac, also occurred in lipid rafts and required raft integrity. Together, these data suggest that Nef induces signal transduction via the recruitment of a signaling machinery including Pak into lipid rafts, thereby mimicking a physiological cellular mechanism to initiate the TCR cascade.