Inactivation of the Host Lipin Gene Accelerates RNA Virus Replication through Viral Exploitation of the Expanded Endoplasmic Reticulum Membrane

RNA viruses take advantage of cellular resources, such as membranes and lipids, to assemble viral replicase complexes (VRCs) that drive viral replication. The host lipins (phosphatidate phosphatases) are particularly interesting because these proteins play key roles in cellular decisions about membrane biogenesis versus lipid storage. Therefore, we examined the relationship between host lipins and tombusviruses, based on yeast model host. We show that deletion of PAH1 (phosphatidic acid phosphohydrolase), which is the single yeast homolog of the lipin gene family of phosphatidate phosphatases, whose inactivation is responsible for proliferation and expansion of the endoplasmic reticulum (ER) membrane, facilitates robust RNA virus replication in yeast. We document increased tombusvirus replicase activity in pah1Δ yeast due to the efficient assembly of VRCs. We show that the ER membranes generated in pah1Δ yeast is efficiently subverted by this RNA virus, thus emphasizing the connection between host lipins and RNA viruses. Thus, instead of utilizing the peroxisomal membranes as observed in wt yeast and plants, TBSV readily switches to the vastly expanded ER membranes in lipin-deficient cells to build VRCs and support increased level of viral replication. Over-expression of the Arabidopsis Pah2p in Nicotiana benthamiana decreased tombusvirus accumulation, validating that our findings are also relevant in a plant host. Over-expression of AtPah2p also inhibited the ER-based replication of another plant RNA virus, suggesting that the role of lipins in RNA virus replication might include several more eukaryotic viruses.

RNA病毒可利用细胞膜与脂质等细胞资源，组装驱动病毒复制的病毒复制酶复合物（viral replicase complexes, VRCs）。宿主脂素（host lipins）尤为值得关注，这类蛋白即磷脂酸磷酸酶，在细胞关于膜生物发生与脂质储存的决策过程中发挥关键调控作用。为此，我们以酵母为模式宿主，探究了宿主脂素与丛矮病毒（tombusviruses）之间的功能关联。研究表明，敲除PAH1基因（PAH1，编码磷脂酸磷酸水解酶，是脂素家族磷脂酸磷酸酶的单一酵母同源基因）可引发内质网（endoplasmic reticulum, ER）膜的增殖与扩张，进而显著增强酵母内RNA病毒的复制效率。我们证实，pah1Δ突变体酵母中，因病毒复制酶复合物的高效组装，丛矮病毒的复制酶活性显著提升。进一步研究发现，该RNA病毒可有效利用pah1Δ突变体酵母中扩增的内质网膜，进一步印证了宿主脂素与RNA病毒之间的功能联系。与野生型酵母及植物中依赖过氧化物酶体膜（peroxisomal membranes）的复制模式不同，TBSV（番茄丛矮病毒）可轻松切换至脂素缺陷细胞中大幅扩张的内质网膜，以此组装病毒复制酶复合物并支持病毒复制水平的提升。在本氏烟草（Nicotiana benthamiana）中过表达拟南芥Pah2p（Arabidopsis Pah2p）可降低丛矮病毒的积累量，验证了我们的发现同样适用于植物宿主。此外，过表达拟南芥AtPah2p（AtPah2p）还可抑制另一种以内质网为复制位点的植物RNA病毒的复制，这提示脂素在RNA病毒复制中的作用可能涵盖更多真核病毒类群。