Multiple poliovirus-induced organelles suggested by comparison of spatiotemporal dynamics of membranous structures and phosphoinositides.
Multiple poliovirus-induced organelles suggested by comparison of spatiotemporal dynamics of membranous structures and phosphoinositides.
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DOI：
10.1371/journal.ppat.1007036
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发表时间：
2018-04
期刊：
影响因子：
6.7
通讯作者：
Cameron CE
中科院分区：
文献类型：
--
作者：
Oh HS;Banerjee S;Aponte-Diaz D;Sharma SD;Aligo J;Lodeiro MF;Ning G;Sharma R;Arnold JJ;Cameron CE
在脊髓灰质炎病毒（PV）增殖达到顶点时，观察到含有磷脂酰肌醇-4-磷酸（PI 4P）的膜，并在横截面中显示为囊泡簇。在基因组复制之前或与基因组复制同时的PI 4P和膜的诱导和重塑尚未得到充分研究。在这里，我们利用两个PV突变体，称为EG和GG，表现出异常的P3前体蛋白水解处理，大大延迟了基因组复制和/或损害病毒组装的发生，照亮PV诱导的膜结构形成的途径。对于WT PV，早在感染后30 min就观察到PI 4P池的变化。甚至在盐酸胍（一种复制抑制剂）存在下也发生PI 4P重塑，并伴随着整个细胞质中膜小管的形成。囊泡簇出现在细胞的核周区域在3小时感染后，时间太慢，这些结构负责基因组复制。在EG和GG PV中观察到的基因组复制开始延迟与病毒诱导的PI 4P池重塑延迟相似，与PI 4P作为基因组复制细胞器的标志物一致。GG PV不能将病毒诱导的小管转化为囊泡簇，这可能解释了该突变体产生的感染性病毒减少近5个对数。我们的研究结果是一致的PV诱导时间不同的膜结构（细胞器）的基因组复制（小管）和病毒组装（泡状簇）。我们认为，形成的速度，时空动态，和复制到组装细胞器转换的效率可以设置由P3多蛋白加工的速率和P3加工产生3AB和/或3CD蛋白的能力。所有的正链RNA病毒复制其基因组与宿主细胞膜。PV不仅重塑现有的膜，而且诱导具有独特结构和脂质组成的膜。有人认为，在感染过程中观察到的PV诱导结构的功能可能不是执行基因组复制的功能。本研究采用动力学分析和动力学陷阱的病毒诱导的膜形成/转化和PI 4P诱导WT PV和两个PV突变体的存在提供证据的病毒诱导的基因组复制细胞器的时间和空间上不同的病毒组装细胞器。此外，我们的研究表明，这两种细胞器的形成可能需要病毒蛋白，3AB和/或3CD的参与。因此，这项研究提供了一个新的视角PV感染的细胞生物学，并应激发一个新的面貌小RNA病毒诱导的细胞器，其功能和P3蛋白在其形成和相互转换的作用。
At the culmination of poliovirus (PV) multiplication, membranes are observed that contain phosphatidylinositol-4-phosphate (PI4P) and appear as vesicular clusters in cross section. Induction and remodeling of PI4P and membranes prior to or concurrent with genome replication has not been well studied. Here, we exploit two PV mutants, termed EG and GG, which exhibit aberrant proteolytic processing of the P3 precursor that substantially delays the onset of genome replication and/or impairs virus assembly, to illuminate the pathway of formation of PV-induced membranous structures. For WT PV, changes to the PI4P pool were observed as early as 30 min post-infection. PI4P remodeling occurred even in the presence of guanidine hydrochloride, a replication inhibitor, and was accompanied by formation of membrane tubules throughout the cytoplasm. Vesicular clusters appeared in the perinuclear region of the cell at 3 h post-infection, a time too slow for these structures to be responsible for genome replication. Delays in the onset of genome replication observed for EG and GG PVs were similar to the delays in virus-induced remodeling of PI4P pools, consistent with PI4P serving as a marker of the genome-replication organelle. GG PV was unable to convert virus-induced tubules into vesicular clusters, perhaps explaining the nearly 5-log reduction in infectious virus produced by this mutant. Our results are consistent with PV inducing temporally distinct membranous structures (organelles) for genome replication (tubules) and virus assembly (vesicular clusters). We suggest that the pace of formation, spatiotemporal dynamics, and the efficiency of the replication-to-assembly-organelle conversion may be set by both the rate of P3 polyprotein processing and the capacity for P3 processing to yield 3AB and/or 3CD proteins. All positive-strand RNA viruses replicate their genomes in association with host cell membranes. PV does not just remodel existing membranes, but induces membranes with unique structure and lipid composition. There has been some suggestion that the functions of the PV-induced structures observed during infection may not be those that perform genome replication. This study uses kinetic analysis and kinetic traps of virus-induced membrane formation/transformation and PI4P induction by WT PV and two PV mutants to provide evidence for the existence of a virus-induced genome-replication organelle temporally and spatially distinct from a virus-assembly organelle. In addition, our studies suggest that formation of both organelles may require participation of viral proteins, 3AB and/or 3CD. Therefore, this study provides a new perspective on the cell biology of PV infection and should inspire a fresh look at picornavirus-induced organelles, their functions and the role of P3 proteins in their formation and interconversion.
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