Cell Biology of Viral Infection Lab (CBV), Instituto Gulbenkian de Ciência (IGC)-Fundação Calouste Gulbenkian, Oeiras, Portugal.
Electron Microscopy Facility (EMF), Instituto Gulbenkian de Ciência (IGC)-Fundação Calouste Gulbenkian, Oeiras, Portugal.
PLoS Biol. 2023 Nov 20;21(11):e3002290. doi: 10.1371/journal.pbio.3002290. eCollection 2023 Nov.
It is now established that many viruses that threaten public health establish condensates via phase transitions to complete their lifecycles, and knowledge on such processes may offer new strategies for antiviral therapy. In the case of influenza A virus (IAV), liquid condensates known as viral inclusions, concentrate the 8 distinct viral ribonucleoproteins (vRNPs) that form IAV genome and are viewed as sites dedicated to the assembly of the 8-partite genomic complex. Despite not being delimited by host membranes, IAV liquid inclusions accumulate host membranes inside as a result of vRNP binding to the recycling endocytic marker Rab11a, a driver of the biogenesis of these structures. We lack molecular understanding on how Rab11a-recycling endosomes condensate specifically near the endoplasmic reticulum (ER) exit sites upon IAV infection. We show here that liquid viral inclusions interact with the ER to fuse, divide, and slide. We uncover that, contrary to previous indications, the reported reduction in recycling endocytic activity is a regulated process rather than a competition for cellular resources involving a novel role for the host factor ATG9A. In infection, ATG9A mediates the removal of Rab11a-recycling endosomes carrying vRNPs from microtubules. We observe that the recycling endocytic usage of microtubules is rescued when ATG9A is depleted, which prevents condensation of Rab11a endosomes near the ER. The failure to produce viral inclusions accumulates vRNPs in the cytosol and reduces genome assembly and the release of infectious virions. We propose that the ER supports the dynamics of liquid IAV inclusions, with ATG9A facilitating their formation. This work advances our understanding on how epidemic and pandemic influenza genomes are formed. It also reveals the plasticity of recycling endosomes to undergo condensation in response to infection, disclosing new roles for ATG9A beyond its classical involvement in autophagy.
现在已经确定，许多威胁公共健康的病毒通过相变形成凝聚体来完成它们的生命周期，而对这些过程的了解可能为抗病毒治疗提供新的策略。以甲型流感病毒（IAV）为例，液滴凝聚体被称为病毒包含体，它们浓缩了形成 IAV 基因组的 8 种独特的病毒核糖核蛋白（vRNP），并被视为专门用于组装 8 分基因组复合物的场所。尽管不被宿主膜限制，但由于 vRNP 与再循环内吞标记物 Rab11a 结合，IAV 液滴包含体在内部积累宿主膜，Rab11a 是这些结构生物发生的驱动因素。我们缺乏分子理解，即在 IAV 感染时，Rab11a 再循环内吞体如何特异性地在内质网（ER）出口位点附近凝聚。我们在这里表明，液滴状病毒包含体与 ER 相互作用融合、分裂和滑动。我们发现，与之前的指示相反，报道的再循环内吞活性降低是一个受调控的过程，而不是涉及宿主因子 ATG9A 的新角色的细胞资源竞争。在感染过程中，ATG9A 介导从微管上去除携带 vRNP 的 Rab11a 再循环内吞体。我们观察到，当 ATG9A 耗尽时，微管的再循环内吞作用得到挽救，从而阻止 Rab11a 内吞体在 ER 附近凝聚。无法产生病毒包含体导致 vRNP 在细胞质中积累，从而减少基因组组装和传染性病毒粒子的释放。我们提出 ER 支持液滴状 IAV 包含体的动态，ATG9A 促进其形成。这项工作提高了我们对流行和大流行流感基因组形成的理解。它还揭示了再循环内吞体在响应感染时发生凝聚的可塑性，揭示了 ATG9A 超越其在自噬中的经典作用的新角色。
