Influenza A viruses use multivalent sialic acid clusters for cell binding and receptor activation
Influenza A viruses use multivalent sialic acid clusters for cell binding and receptor activation
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DOI：
10.1371/journal.ppat.1008656
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发表时间：
2020-07-01
期刊：
影响因子：
6.7
通讯作者：
Manley, Suliana
中科院分区：
文献类型：
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作者：
Sieben, Christian;Sezgin, Erdinc;Manley, Suliana
甲型流感病毒 (IAV) 使用主要病毒表面蛋白血凝素 (HA) 结合其宿主细胞。 HA 识别唾液酸，这是一种质膜聚糖，充当特定的主要附着因子 (AF)。由于单独的唾液酸不能实现信号传导功能，因此病毒需要激活下游因子来触发内吞摄取。最近，表皮生长因子受体 (EGFR)（受体酪氨酸激酶家族的成员）被证明可以被 IAV 激活并传递细胞进入信号。然而，IAV 与唾液酸的结合如何导致 EGFR 的结合和激活仍不清楚。我们使用多色超分辨率显微镜在 IAV 颗粒尺度上研究 IAV AF 及其功能受体 EGFR 的横向组织。有趣的是，定量聚类分析表明，AF 和 EGFR 在 A549 细胞质膜中以部分重叠的亚微米簇的形式组织。在 AF 域内，局部 AF 浓度平均达到背景浓度的 10 倍，并且趋向于向聚类中心增加，从而代表了多价病毒结合平台。利用我们通过实验测量的簇特征，我们模拟了病毒在平板膜上的扩散。结果预测，局部 AF 浓度强烈影响 IAV 的独特移动模式，其方式与活细胞单病毒跟踪数据一致。与 AF 相比，EGFR 存在于较小的簇中。病毒结合会激活 EGFR，但有趣的是，这一过程的发生并没有发生主要的横向 EGFR 重新分布，这表明预先形成的簇被激活，我们证明其寿命较长。总而言之，我们的结果提供了对流感病毒感染初始步骤的定量了解。 AF 和 EGFR 的共簇允许在特定平台上产生结合和信号传导的协同效应，从而将它们的空间组织与其在病毒-细胞结合和受体激活过程中的功能作用联系起来。作者摘要 质膜是细胞与其环境之间的主要界面。这种复杂而动态的细胞器需要作为屏障进行保护，但也需要将微妙的信号传入和传出细胞。对于包膜病毒 IAV，质膜既是感染过程中需要克服的主要障碍，也是子代病毒颗粒组装的场所。然而，质膜的组织——理解病毒进入如何发挥作用的关键——在感染颗粒的尺度上（长度尺度< 100 nm）仍然很大程度上未知。唾液酸化聚糖可作为 IAV 附着因子，但不能跨质膜传递信号。受体酪氨酸激酶被鉴定为在病毒结合后被激活并充当功能性受体。 IAV 在最初结合聚糖时如何参与并激活其功能受体仍然是推测。在这里，我们使用超分辨率显微镜来研究参与 IAV 感染的质膜结合分子的横向组织及其功能关系。我们发现分子以亚微米纳米域的形式组织，并与病毒扩散模拟相结合，提出了 IAV 如何首先与质膜中的 AF 结合，随后与进入相关的膜受体结合并触发的机制模型。
Influenza A virus (IAV) binds its host cell using the major viral surface protein hemagglutinin (HA). HA recognizes sialic acid, a plasma membrane glycan that functions as the specific primary attachment factor (AF). Since sialic acid alone cannot fulfill a signaling function, the virus needs to activate downstream factors to trigger endocytic uptake. Recently, the epidermal growth factor receptor (EGFR), a member of the receptor-tyrosine kinase family, was shown to be activated by IAV and transmit cell entry signals. However, how IAV's binding to sialic acid leads to engagement and activation of EGFR remains largely unclear. We used multicolor super-resolution microscopy to study the lateral organization of both IAV's AFs and its functional receptor EGFR at the scale of the IAV particle. Intriguingly, quantitative cluster analysis revealed that AFs and EGFR are organized in partially overlapping submicrometer clusters in the plasma membrane of A549 cells. Within AF domains, the local AF concentration reaches on average 10-fold the background concentration and tends to increase towards the cluster center, thereby representing a multivalent virus-binding platform. Using our experimentally measured cluster characteristics, we simulated virus diffusion on a flat membrane. The results predict that the local AF concentration strongly influences the distinct mobility pattern of IAVs, in a manner consistent with live-cell single-virus tracking data. In contrast to AFs, EGFR resides in smaller clusters. Virus binding activates EGFR, but interestingly, this process occurs without a major lateral EGFR redistribution, indicating the activation of pre-formed clusters, which we show are long-lived. Taken together, our results provide a quantitative understanding of the initial steps of influenza virus infection. Co-clustering of AF and EGFR permit a cooperative effect of binding and signaling at specific platforms, thus linking their spatial organization to their functional role during virus-cell binding and receptor activation.Author summary The plasma membrane is the major interface between a cell and its environment. This complex and dynamic organelle needs to protect, as a barrier, but also transmit subtle signals into and out of the cell. For the enveloped virus IAV, the plasma membrane represents both a major obstacle to overcome during infection, and the site for the assembly of progeny virus particles. However, the organisation of the plasma membrane-a key to understanding how viral entry works-at the scale of an infecting particle (length scales < 100 nm) remains largely unknown. Sialylated glycans serve as IAV attachment factors but are not able to transmit signals across the plasma membrane. Receptor tyrosine kinases were identified to be activated upon virus binding and serve as functional receptor. How IAV engages and activates its functional receptors while initially binding glycans still remains speculative. Here, we use super resolution microscopy to study the lateral organization of plasma membrane-bound molecules involved in IAV infection, as well as their functional relationship. We find that molecules are organized in submicrometer nanodomains and, in combination with virus diffusion simulations, present a mechanistic model for how IAV first engages with AFs in the plasma membrane to subsequently engage and trigger entry-associated membrane receptors.