The RING-CH ligase K5 antagonizes restriction of KSHV and HIV-1 particle release by mediating ubiquitin-dependent endosomal degradation of tetherin.
The RING-CH ligase K5 antagonizes restriction of KSHV and HIV-1 particle release by mediating ubiquitin-dependent endosomal degradation of tetherin.
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DOI：
10.1371/journal.ppat.1000843
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发表时间：
2010-04-15
期刊：
影响因子：
6.7
通讯作者：
Neil SJ
中科院分区：
文献类型：
--
作者：
Pardieu C;Vigan R;Wilson SJ;Calvi A;Zang T;Bieniasz P;Kellam P;Towers GJ;Neil SJ
Tetherin (CD317/BST2) 是一种干扰素诱导的膜蛋白，可抑制多种有包膜病毒颗粒的释放。几种哺乳动物病毒已经进化出了使系链蛋白失活的对策，其原型是 HIV-1 Vpu 蛋白。在这里，我们证明人类疱疹病毒卡波西肉瘤相关疱疹病毒（KSHV）对系链蛋白限制敏感，并且其活性被 KSHV 编码的 RING-CH E3 泛素连接酶 K5 抵消。 KSHV 感染细胞中 Tetherin 的表达会抑制病毒颗粒的释放，就像使用 RNA 干扰消除 K5 蛋白一样。 K5 诱导细胞表面人类系链蛋白的物种特异性下调，随后发生内体降解。我们发现，K5 靶向 tetherin 细胞质尾部的单个赖氨酸 (K18) 进行泛素化，导致 tetherin 重新定位到 CD63 阳性内体区室。系链蛋白降解依赖于 ESCRT 介导的内体分选，但不需要系链蛋白胞质尾部基于酪氨酸的分选信号。重要的是，我们还表明，K5 在 HIV-1 释放中替代 Vpu 的能力完全依赖于 K18 和 K5 的 RING-CH 结构域。相比之下，虽然Vpu诱导tetherin细胞质尾部赖氨酸残基泛素化，但这些位置的突变对其tetherin功能的拮抗作用没有影响，并且残留的tetherin与表达Vpu的细胞中的反式高尔基体网络（TGN）相关。综上所述，我们的结果表明，K5 是一种机制独特的针对系链蛋白介导的限制的病毒对策，并且疱疹病毒颗粒的释放对这种抗病毒抑制模式敏感。为了在宿主中有效复制，病毒必须避开构成先天免疫系统一部分的抗病毒细胞防御。 Tetherin 是一种抗病毒膜蛋白，可抑制受感染细胞释放多种包膜病毒，可被 HIV-1 Vpu 蛋白拮抗。人类病原体卡波西肉瘤相关疱疹病毒 (KSHV) 的 K5 蛋白调节多种宿主蛋白（包括系链蛋白）的细胞表面水平。我们证明 KSHV 释放对系链蛋白敏感，并且 K5 表达是在表达系链蛋白的细胞中有效生产病毒所必需的。 K5 还能够挽救 Vpu 缺陷的 HIV-1 病毒从系链蛋白中释放。 K5 表达诱导细胞表面系链蛋白水平下调和晚期内涵体降解，这取决于系链蛋白细胞质尾部的单个赖氨酸残基。最后，我们表明，ESCRT 途径促进细胞表面受体的运输以进行降解，是 K5 介导的系链蛋白从质膜去除所必需的。因此，我们证明疱疹病毒对系链蛋白的抗病毒作用敏感，并且 KSHV 已经进化出一种破坏其的机制。这些发现扩大了对系链蛋白敏感的病毒列表，表明系链蛋白对策是包膜病毒中广泛的防御机制。
Tetherin (CD317/BST2) is an interferon-induced membrane protein that inhibits the release of diverse enveloped viral particles. Several mammalian viruses have evolved countermeasures that inactivate tetherin, with the prototype being the HIV-1 Vpu protein. Here we show that the human herpesvirus Kaposi's sarcoma-associated herpesvirus (KSHV) is sensitive to tetherin restriction and its activity is counteracted by the KSHV encoded RING-CH E3 ubiquitin ligase K5. Tetherin expression in KSHV-infected cells inhibits viral particle release, as does depletion of K5 protein using RNA interference. K5 induces a species-specific downregulation of human tetherin from the cell surface followed by its endosomal degradation. We show that K5 targets a single lysine (K18) in the cytoplasmic tail of tetherin for ubiquitination, leading to relocalization of tetherin to CD63-positive endosomal compartments. Tetherin degradation is dependent on ESCRT-mediated endosomal sorting, but does not require a tyrosine-based sorting signal in the tetherin cytoplasmic tail. Importantly, we also show that the ability of K5 to substitute for Vpu in HIV-1 release is entirely dependent on K18 and the RING-CH domain of K5. By contrast, while Vpu induces ubiquitination of tetherin cytoplasmic tail lysine residues, mutation of these positions has no effect on its antagonism of tetherin function, and residual tetherin is associated with the trans-Golgi network (TGN) in Vpu-expressing cells. Taken together our results demonstrate that K5 is a mechanistically distinct viral countermeasure to tetherin-mediated restriction, and that herpesvirus particle release is sensitive to this mode of antiviral inhibition. To replicate efficiently in their hosts, viruses must avoid antiviral cellular defenses that comprise part of the innate immune system. Tetherin, an antiviral membrane protein that inhibits the release of several enveloped viruses from infected cells, is antagonized by the HIV-1 Vpu protein. The K5 protein of the human pathogen Kaposi's sarcoma-associated herpesvirus (KSHV) modulates the cell surface levels of several host proteins including tetherin. We show that KSHV release is sensitive to tetherin, and that K5 expression is required for efficient virus production in tetherin-expressing cells. K5 is also capable of rescuing Vpu-defective HIV-1 virus release from tetherin. K5 expression induces a down-regulation of cell-surface tetherin levels and degradation in late endosomes, which depends on a single lysine residue in the tetherin cytoplasmic tail. Finally, we show that the ESCRT pathway, which promotes the trafficking of cell surface receptors for degradation, is required for K5-mediated tetherin removal from the plasma membrane. Thus, we demonstrate that herpesviruses are sensitive to the antiviral effects of tetherin and that KSHV has evolved a mechanism for its destruction. These findings extend the list of viruses sensitive to tetherin, suggesting that tetherin counter-measures are widespread defense mechanisms amongst enveloped viruses.
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