For entry, Ebola virus (EBOV) requires the interaction of its viral glycoprotein with the cellular protein Niemann-Pick C1 (NPC1) which resides in late endosomes and lysosomes. How EBOV is trafficked and delivered to NPC1 and whether this is positively regulated during entry remain unclear. Here, we show that the PIKfyve-ArPIKfyve-Sac3 cellular complex, which is involved in the metabolism of phosphatidylinositol (3,5) bisphosphate (PtdIns(3,5)P₂), is critical for EBOV infection. Although the expression of all subunits of the complex was required for efficient entry, PIKfyve kinase activity was specifically critical for entry by all pathogenic filoviruses. Inhibition of PIKfyve prevented colocalization of EBOV with NPC1 and led to virus accumulation in intracellular vesicles with characteristics of early endosomes. Importantly, genetically-encoded phosphoinositide probes revealed an increase in PtdIns(3,5)P₂-positive vesicles in cells during EBOV entry. Taken together, our studies suggest that EBOV requires PtdIns(3,5)P₂ production in cells to promote efficient delivery to NPC1.

为了进入，埃博拉病毒（EBOV）需要其病毒糖蛋白与驻留在晚期内体和溶酶体中的细胞蛋白Niemann-Pick C1（NPC1）相互作用。目前尚不清楚如何传播EBOV并将其传递给NPC1，以及在进入过程中是否对其进行了积极的监管。在这里，我们显示PIKfyve-ArPIKfyve-Sac3细胞复合物，参与磷脂酰肌醇（3,5）双磷酸（PtdIns（3,5）P ₂的代谢），对于EBOV感染至关重要。尽管复合物的所有亚基的表达是有效进入所必需的，但PIKfyve激酶活性对于所有病原性丝状病毒的进入特别重要。抑制PIKfyve可以阻止EBOV与NPC1的共定位，并导致病毒在具有早期内体特征的细胞内囊泡中积聚。重要的是，基因编码的磷酸肌醇探针揭示了EBOV进入过程中细胞中PtdIns（3,5）P ₂阳性囊泡的增加。两者合计，我们的研究表明，EBOV需要在细胞中产生PtdIns（3,5）P ₂才能促进向NPC1的有效传递。