A Two-Pronged Structural Analysis of Retroviral Maturation Indicates that Core Formation Proceeds by a Disassembly-Reassembly Pathway Rather than a Displacive Transition
A Two-Pronged Structural Analysis of Retroviral Maturation Indicates that Core Formation Proceeds by a Disassembly-Reassembly Pathway Rather than a Displacive Transition
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DOI：
10.1128/jvi.01408-13
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发表时间：
2013-12-01
影响因子：
5.4
通讯作者：
Steven, Alasdair C.
中科院分区：
文献类型：
--
作者：
Keller, Paul W.;Huang, Rick K.;Steven, Alasdair C.
逆转录病毒的成熟过程包括Gag多蛋白的连续裂解，Gag多蛋白最初排列在一个球形壳中，导致形成多面体或圆锥形的衣壳。有证据表明，衣壳在成熟的病毒粒子内由解离衣壳（CA）蛋白重新组装，但可能存在一种置换途径，其中CA壳保持组装但被重塑。抑制CA与间隔肽SP1/SP之间的最终裂解可阻断成熟衣壳的产生。我们通过检测劳斯肉瘤病毒（RSV） CA-SP在体外组装成二十面体衣壳的能力来研究SP的保留是否会导致CA组装不合格。衣壳确实组装，并且与单独由CA形成的衣壳难以区分，表明SP是无序的。我们还使用冷冻电子断层扫描来表征在成熟抑制剂PF-46396或切割阻断CA5突变存在下产生的HIV-1颗粒。抑制剂处理的病毒粒子有一个类似于未成熟Gag壳的CA层的壳，但不太完整。生成一些CA蛋白，但通常不足以形成成熟的核心。我们提出，像PF-46396这样的抑制剂与Gag晶格结合，在那里它们阻止蛋白酶进入CA- sp1切割位点，并阻止CA的释放。CA5颗粒在CA- sp1位点没有切割，具有相对薄壁的球形壳。这种晶格似乎向着成熟状态发展，但既不产生圆锥形核，也不产生感染性病毒粒子。这些观察结果支持岩心形成的拆解-重组途径。
Retrovirus maturation involves sequential cleavages of the Gag polyprotein, initially arrayed in a spherical shell, leading to formation of capsids with polyhedral or conical morphology. Evidence suggests that capsids assemble de novo inside maturing virions from dissociated capsid (CA) protein, but the possibility persists of a displacive pathway in which the CA shell remains assembled but is remodeled. Inhibition of the final cleavage between CA and spacer peptide SP1/SP blocks the production of mature capsids. We investigated whether retention of SP might render CA assembly incompetent by testing the ability of Rous sarcoma virus (RSV) CA-SP to assemble in vitro into icosahedral capsids. Capsids were indeed assembled and were indistinguishable from those formed by CA alone, indicating that SP was disordered. We also used cryo-electron tomography to characterize HIV-1 particles produced in the presence of maturation inhibitor PF-46396 or with the cleavage-blocking CA5 mutation. Inhibitor-treated virions have a shell that resembles the CA layer of the immature Gag shell but is less complete. Some CA protein is generated but usually not enough for a mature core to assemble. We propose that inhibitors like PF-46396 bind to the Gag lattice where they deny the protease access to the CA-SP1 cleavage site and prevent the release of CA. CA5 particles, which exhibit no cleavage at the CA-SP1 site, have spheroidal shells with relatively thin walls. It appears that this lattice progresses displacively toward a mature-like state but produces neither conical cores nor infectious virions. These observations support the disassembly- reassembly pathway for core formation.