Rupp Jonathan C, Locatelli Manon, Grieser Alexis, Ramos Andrea, Campbell Patricia J, Yi Hong, Steel John, Burkhead Jason L, Bortz Eric
Department of Biological Sciences, University of Alaska Anchorage, Anchorage, AK, USA.
Present address: Institute Albert Bonniot - INSERM U1209, Université Grenoble Alpes, Grenoble, France.
Virol J. 2017 Jan 23;14(1):11. doi: 10.1186/s12985-016-0671-7.
The essential role of copper in eukaryotic cellular physiology is known, but has not been recognized as important in the context of influenza A virus infection. In this study, we investigated the effect of cellular copper on influenza A virus replication.
Influenza A/WSN/33 (H1N1) virus growth and macromolecule syntheses were assessed in cultured human lung cells (A549) where the copper concentration of the growth medium was modified, or expression of host genes involved in copper homeostasis was targeted by RNA interference.
Exogenously increasing copper concentration, or chelating copper, resulted in moderate defects in viral growth. Nucleoprotein (NP) localization, neuraminidase activity assays and transmission electron microscopy did not reveal significant defects in virion assembly, morphology or release under these conditions. However, RNAi knockdown of the high-affinity copper importer CTR1 resulted in significant viral growth defects (7.3-fold reduced titer at 24 hours post-infection, p = 0.04). Knockdown of CTR1 or the trans-Golgi copper transporter ATP7A significantly reduced polymerase activity in a minigenome assay. Both copper transporters were required for authentic viral RNA synthesis and NP and matrix (M1) protein accumulation in the infected cell.
These results demonstrate that intracellular copper regulates the influenza virus life cycle, with potentially distinct mechanisms in specific cellular compartments. These observations provide a new avenue for drug development and studies of influenza virus pathogenesis.
铜在真核细胞生理学中的重要作用已为人所知，但在甲型流感病毒感染的背景下尚未被视为重要因素。在本研究中，我们调查了细胞内铜对甲型流感病毒复制的影响。
在培养的人肺细胞（A549）中评估甲型流感病毒A/WSN/33（H1N1）的生长及大分子合成，其中生长培养基的铜浓度被改变，或者通过RNA干扰靶向参与铜稳态的宿主基因的表达。
外源增加铜浓度或螯合铜会导致病毒生长出现中度缺陷。在这些条件下，核蛋白（NP）定位、神经氨酸酶活性测定及透射电子显微镜检查未发现病毒粒子组装、形态或释放存在明显缺陷。然而，高亲和力铜转运体CTR1的RNA干扰敲低导致显著的病毒生长缺陷（感染后24小时滴度降低7.3倍，p = 0.04）。在微型基因组试验中，CTR1或反式高尔基体铜转运体ATP7A的敲低显著降低了聚合酶活性。这两种铜转运体对于受感染细胞中真实的病毒RNA合成以及NP和基质（M1）蛋白积累都是必需的。
这些结果表明细胞内铜调节流感病毒生命周期，在特定细胞区室中可能存在不同机制。这些观察结果为流感病毒发病机制的药物开发和研究提供了新途径。
