RESUMO
Influenza A viruses (IAV) are enveloped viruses carrying a single-stranded negative-sense RNA genome. Detection of host proteins having a relationship with IAV and revealing of the role of these proteins in the viral replication are of great importance in keeping IAV infections under control. Consequently, the importance of human DDX56, which is determined to be associated with a viral NS1 with a yeast two-hybrid assay, was investigated for IAV replication. The viral replication in knocked down cells for the DDX56 gene was evaluated. The NS1 was co-precipitated with the DDX56 protein in lysates of cells transiently expressing DDX56 and NS1 or infected with the viruses, showing that NS1 and DDX56 interact in mammalian cells. Viral NS1 showed a tendency to co-localize with DDX56 in the cells, transiently expressing both of these proteins, which supports the IP and two-hybrid assays results. The data obtained with in silico predictions supported the in vitro protein interaction results. The viral replication was significantly reduced in the DDX56-knockdown cells comparing with that in the control cells. In conclusion, human DDX56 protein interacts with the IAV NS1 protein in both yeast and mammalian cells and has a positive regulatory effect on IAV replication. However, the mechanism of DDX56 on IAV replication requires further elucidation.
RESUMO
Influenza A viruses have a single-stranded RNA genome consisting of 8 segments. Each RNA segment associates with the nucleoprotein (NP) and viral RNA polymerase to and from a viral ribonucleoprotein (vRNP) particle. The viral mRNA synthesis is dependent on a capped primer derived from nascent host RNA transcripts. For these processes to take place, vRNPs must pass through the cell nuclear pore complex (NPC) to the nucleus. The influenza A virus NS2 protein, also called the nuclear export protein (NES), has an important role in the nucleocytoplasmic transport of vRNPs. This protein interacts with the host cellular nucleoporins during the nuclear export of vRNPs. In this study, the human nucleoporin 214 (Nup214) was identified as an NS2-binding protein by using a yeast two-hybrid assay. The interaction between NS2 and human Nup214 was confirmed in both yeast and mammalian cells. It has been shown that the NS2 protein interacts with the amino terminal FG domain of the Nup214 protein. The influenza viral replication was suppressed in knockdown cells for the Nup214 protein. It was concluded that the FG domains of nucleoporins have an important role in the interaction of the influenza NS2 protein with host NPC for vRNA export.
