ALIX Rescues Budding of a Double PTAP/PPEY L-Domain Deletion Mutant of Ebola VP40: A Role for ALIX in Ebola Virus Egress
ALIX Rescues Budding of a Double PTAP/PPEY L-Domain Deletion Mutant of Ebola VP40: A Role for ALIX in Ebola Virus Egress
复制标题
DOI：
10.1093/infdis/jiu838
复制
发表时间：
2015-10-01
影响因子：
6.4
通讯作者：
Harty, Ronald N.
中科院分区：
文献类型：
--
作者：
Han, Ziying;Madara, Jonathan J.;Harty, Ronald N.
埃博拉病毒（EBOV）是一种有包膜的负义RNA病毒，属于丝状病毒科，可引起高死亡率的出血热综合征。到目前为止，还没有获得许可的疫苗或治疗剂来控制EBOV感染和预防传播。因此，需要更好地了解调节病毒传播的机制，这对制定对策至关重要。EBOV VP 40基质蛋白在病毒体组装和排出的后期阶段中起核心作用，并且VP 40的独立表达通过精确模拟活病毒出芽的机制导致病毒样颗粒（VLP）的产生。VP 40晚期（L）出芽结构域通过募集宿主ESCRT和ESCRT相关蛋白以完成膜分裂过程来介导有效的病毒-细胞分离。L-结构域由核心共有氨基酸基序组成，包括PPxY、P（T/S）AP和YPx（（n））L/I，并且EBOV VP 40含有重叠的PPxY和PTAP基序，其分别与Nedd 4和Tsg 101的相互作用已被广泛表征。在这里，我们首次提供的数据表明，EBOV VP 40具有第三个L-结构域YPx（（n））L/I共有基序，与ESCRT-III蛋白阿利克斯相互作用。我们发现定位于EBOV VP 40氨基酸18-26的YPx（（n））L/I基序与阿利克斯Bro 1-V片段相互作用，并且内源性阿利克斯表达的siRNA敲低抑制EBOV VP 40 VLP的排出。此外，过表达阿利克斯Bro 1-V将出芽缺陷型EBOV VP 40双PTAP/PPEY L结构域缺失突变体的VLP产生拯救至野生型水平。总之，这些发现表明EBOV VP 40通过YPx（（n））L/I基序招募宿主阿利克斯，该基序可以作为替代L结构域来促进病毒外出。
Ebola (EBOV) is an enveloped, negative-sense RNA virus belonging to the family Filoviridae that causes hemorrhagic fever syndromes with high-mortality rates. To date, there are no licensed vaccines or therapeutics to control EBOV infection and prevent transmission. Consequently, the need to better understand the mechanisms that regulate virus transmission is critical to developing countermeasures. The EBOV VP40 matrix protein plays a central role in late stages of virion assembly and egress, and independent expression of VP40 leads to the production of virus-like particles (VLPs) by a mechanism that accurately mimics budding of live virus. VP40 late (L) budding domains mediate efficient virus-cell separation by recruiting host ESCRT and ESCRT-associated proteins to complete the membrane fission process. L-domains consist of core consensus amino acid motifs including PPxY, P(T/S) AP, and YPx((n)) L/I, and EBOV VP40 contains overlapping PPxY and PTAP motifs whose interactions with Nedd4 and Tsg101, respectively, have been characterized extensively. Here, we present data demonstrating for the first time that EBOV VP40 possesses a third L-domain YPx((n)) L/I consensus motif that interacts with the ESCRT-III protein Alix. We show that the YPx((n)) L/I motif mapping to amino acids 18-26 of EBOV VP40 interacts with the Alix Bro1-V fragment, and that siRNA knockdown of endogenous Alix expression inhibits EBOV VP40 VLP egress. Furthermore, overexpression of Alix Bro1-V rescues VLP production of the budding deficient EBOV VP40 double PTAP/PPEY L-domain deletion mutant to wild-type levels. Together, these findings demonstrate that EBOV VP40 recruits host Alix via a YPx((n)) L/I motif that can function as an alternative L-domain to promote virus egress.