The phosphorylation of HIV-1 Gag by atypical protein kinase C facilitates viral infectivity by promoting Vpr incorporation into virions.
The phosphorylation of HIV-1 Gag by atypical protein kinase C facilitates viral infectivity by promoting Vpr incorporation into virions.
复制标题
DOI：
10.1186/1742-4690-11-9
复制
发表时间：
2014-01-22
期刊：
影响因子：
3.3
通讯作者：
Ryo A
中科院分区：
文献类型：
--
作者：
Kudoh A;Takahama S;Sawasaki T;Ode H;Yokoyama M;Okayama A;Ishikawa A;Miyakawa K;Matsunaga S;Kimura H;Sugiura W;Sato H;Hirano H;Ohno S;Yamamoto N;Ryo A
人类免疫缺陷病毒1型（HIV-1）Gag是介导病毒样颗粒（VLP）从感染细胞膜组装和释放的主要结构蛋白。Gag C-末端p6结构域含有促进病毒从质膜释放并介导病毒Vpr蛋白掺入的短序列基序。Gag p6也被发现在HIV-1感染期间被磷酸化，并且该事件可能影响病毒复制。然而，指导Gag p6磷酸化作用于病毒复制的激酶仍有待鉴定。在我们目前的研究中，我们使用蛋白质组学方法鉴定了这种激酶，并进一步描述了其在HIV-1复制中的作用。设计了一种蛋白质组学方法来系统地鉴定与HIV-1 Gag有效相互作用的人类蛋白激酶，并成功地鉴定了22种候选蛋白。在这组中，发现非典型蛋白激酶C（aPKC）磷酸化HIV-1 Gag p6。随后的LC-MS/MS和使用磷酸化特异性抗体的免疫印迹分析在体外和体内证实了aPKC在Ser 487处磷酸化HIV-1 Gag。计算机辅助的结构建模和随后的基于细胞的测定显示，这种磷酸化事件是必需的Gag和Vpr之间的相互作用，并导致Vpr掺入到病毒体中。此外，抑制aPKC活性降低了病毒体中的Vpr水平，并削弱了人原代巨噬细胞的HIV-1感染性。我们目前的研究结果首次表明，HIV-1 Gag的丝氨酸487磷酸化是由aPKC介导的，这种激酶可以调节Vpr掺入HIV-1病毒粒子，从而支持病毒的感染性。此外，aPKC抑制有效抑制巨噬细胞中的HIV-1感染性。因此，aPKC可能是HIV-1感染的一个有趣的治疗靶点。
Human immunodeficiency virus type 1 (HIV-1) Gag is the main structural protein that mediates the assembly and release of virus-like particles (VLPs) from an infected cell membrane. The Gag C-terminal p6 domain contains short sequence motifs that facilitate virus release from the plasma membrane and mediate incorporation of the viral Vpr protein. Gag p6 has also been found to be phosphorylated during HIV-1 infection and this event may affect virus replication. However, the kinase that directs the phosphorylation of Gag p6 toward virus replication remains to be identified. In our present study, we identified this kinase using a proteomic approach and further delineate its role in HIV-1 replication. A proteomic approach was designed to systematically identify human protein kinases that potently interact with HIV-1 Gag and successfully identified 22 candidates. Among this panel, atypical protein kinase C (aPKC) was found to phosphorylate HIV-1 Gag p6. Subsequent LC-MS/MS and immunoblotting analysis with a phospho-specific antibody confirmed both in vitro and in vivo that aPKC phosphorylates HIV-1 Gag at Ser487. Computer-assisted structural modeling and a subsequent cell-based assay revealed that this phosphorylation event is necessary for the interaction between Gag and Vpr and results in the incorporation of Vpr into virions. Moreover, the inhibition of aPKC activity reduced the Vpr levels in virions and impaired HIV-1 infectivity of human primary macrophages. Our current results indicate for the first time that HIV-1 Gag phosphorylation on Ser487 is mediated by aPKC and that this kinase may regulate the incorporation of Vpr into HIV-1 virions and thereby supports virus infectivity. Furthermore, aPKC inhibition efficiently suppresses HIV-1 infectivity in macrophages. aPKC may therefore be an intriguing therapeutic target for HIV-1 infection.
登录
查看更多内容
影响因子：
5.6
作者：
Chugh P;Fan S;Planelles V;Maggirwar SB;Dewhurst S;Kim B
通讯作者：
Kim B
影响因子：
3.7
作者：
BALLIET, JW;KOLSON, DL;COLLMAN, R
通讯作者：
COLLMAN, R
影响因子：
4.8
作者：
Cartier, C;Sivard, P;Boyer, V
通讯作者：
Boyer, V
影响因子：
5.4
作者：
Demirov, DG;Orenstein, JM;Freed, EO
通讯作者：
Freed, EO
影响因子：
5.4
作者：
Folgueira, L;McElhinny, JA;Paya, CV
通讯作者：
Paya, CV