Visualization and Sequencing of Membrane Remodeling Leading to Influenza Virus Fusion
Visualization and Sequencing of Membrane Remodeling Leading to Influenza Virus Fusion
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DOI：
10.1128/jvi.00240-16
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发表时间：
2016-08-01
影响因子：
5.4
通讯作者：
Lee, Kelly K.
中科院分区：
文献类型：
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作者：
Gui, Long;Ebner, Jamie L.;Lee, Kelly K.
蛋白质介导的膜融合是许多基本生物学事件（包括包膜病毒感染）中的重要步骤。蛋白质和膜中间体的性质以及在这些基本过程中膜重塑的顺序仍然知之甚少。在这里，我们使用冷冻电子断层扫描（cryo-ET）成像流感病毒和囊泡与一系列脂质成分之间的相互作用。通过以下的群体动力学的膜融合中间体成像的冷冻ET，我们发现，膜重塑开始与血凝素融合蛋白钉抓到目标膜上，然后由局部的目标膜dimpling作为本地集群的血凝素开始进行构象重折叠。然后，局部凹陷转变为延伸的紧密贴壁接触区，其中两个近端膜瓣叶在大多数情况下彼此无法区分，表明脂质头部基团显著脱水和可能混合。增加目标膜中融合增强胆固醇或双单酰基甘油磷酸的含量导致扩展接触区形成的增加。有趣的是，半融合的中间体被发现在这里研究的流感病毒融合系统中是极其罕见的，最有可能反映了这种状态的不稳定性及其快速转化为融合后复合物，随着时间的推移，人口增加。通过跟踪融合复合物的群体随着时间的推移，结构和序列的膜重组，从而导致有效的包膜病毒fusion.IMPORTANCEEnveloped病毒使用专门的表面蛋白介导的细胞和病毒膜的融合，导致形成的孔，通过该病毒的遗传物质被传递到细胞。对于流感病毒，三聚体血凝素（HA）糖蛋白刺突介导宿主细胞附着和膜融合。虽然已经表征了融合机器的构象和部分的子集的结构，但是在融合期间膜变形的性质和顺序在很大程度上回避了表征。在集中于HA介导的膜重塑的早期阶段的研究的基础上，在此使用冷冻电子断层扫描（cryo-ET）来对处于与脂质体融合的不同阶段的完整流感病毒粒子的三维组织进行成像，从而一路引导至融合反应的完成。通过监测融合中间群体在酸诱导的融合过程中的演变，我们确定了导致包膜病毒有效融合的膜重组的进展。
Protein-mediated membrane fusion is an essential step in many fundamental biological events, including enveloped virus infection. The nature of protein and membrane intermediates and the sequence of membrane remodeling during these essential processes remain poorly understood. Here we used cryo-electron tomography (cryo-ET) to image the interplay between influenza virus and vesicles with a range of lipid compositions. By following the population kinetics of membrane fusion intermediates imaged by cryo-ET, we found that membrane remodeling commenced with the hemagglutinin fusion protein spikes grappling onto the target membrane, followed by localized target membrane dimpling as local clusters of hemagglutinin started to undergo conformational refolding. The local dimples then transitioned to extended, tightly apposed contact zones where the two proximal membrane leaflets were in most cases indistinguishable from each other, suggesting significant dehydration and possible intermingling of the lipid head groups. Increasing the content of fusion-enhancing cholesterol or bis-monoacylglycerophosphate in the target membrane led to an increase in extended contact zone formation. Interestingly, hemifused intermediates were found to be extremely rare in the influenza virus fusion system studied here, most likely reflecting the instability of this state and its rapid conversion to postfusion complexes, which increased in population over time. By tracking the populations of fusion complexes over time, the architecture and sequence of membrane reorganization leading to efficient enveloped virus fusion were thus resolved.IMPORTANCEEnveloped viruses employ specialized surface proteins to mediate fusion of cellular and viral membranes that results in the formation of pores through which the viral genetic material is delivered to the cell. For influenza virus, the trimeric hemagglutinin (HA) glycoprotein spike mediates host cell attachment and membrane fusion. While structures of a subset of conformations and parts of the fusion machinery have been characterized, the nature and sequence of membrane deformations during fusion have largely eluded characterization. Building upon studies that focused on early stages of HA-mediated membrane remodeling, here cryo-electron tomography (cryo-ET) was used to image the three-dimensional organization of intact influenza virions at different stages of fusion with liposomes, leading all the way to completion of the fusion reaction. By monitoring the evolution of fusion intermediate populations over the course of acid-induced fusion, we identified the progression of membrane reorganization that leads to efficient fusion by an enveloped virus.