Abstract
BackgroundInfluenza A virus (IVA) exploits diverse cellular gene products to support its replication in the host. The significance of the regulatory (β) subunit of casein kinase 2 (CK2β) in various cellular mechanisms is well established, but less is known about its potential role in IVA replication. We studied the role of CK2β in IVA-infected A549 human epithelial lung cells.ResultsActivation of CK2β was observed in A549 cells during virus binding and internalization but appeared to be constrained as replication began. We used small interfering RNAs (siRNAs) targeting CK2β mRNA to silence CK2β protein expression in A549 cells without affecting expression of the CK2α subunit. CK2β gene silencing led to increased virus titers, consistent with the inhibition of CK2β during IVA replication. Notably, virus titers increased significantly when CK2β siRNA-transfected cells were inoculated at a lower multiplicity of infection. Virus titers also increased in cells treated with a specific CK2 inhibitor but decreased in cells treated with a CK2β stimulator. CK2β absence did not impair nuclear export of viral ribonucleoprotein complexes (6 h and 8 h after inoculation) or viral polymerase activity (analyzed in a minigenome system). The enhancement of virus titers by CK2β siRNA reflects increased cell susceptibility to influenza virus infection resulting in accelerated virus entry and higher viral protein content.ConclusionThis study demonstrates the role of cellular CK2β protein in the viral biology. Our results are the first to demonstrate a functional link between siRNA-mediated inhibition of the CK2β protein and regulation of influenza A virus replication in infected cells. Overall, the data suggest that expression and activation of CK2β inhibits influenza virus replication by regulating the virus entry process and viral protein synthesis.
Highlights
Influenza A virus (IVA) exploits diverse cellular gene products to support its replication in the host
To assess whether CK2β is activated by influenza virus, we incubated A549 cells with medium containing the human A/Puerto Rico/8/34 influenza virus for 1 h at 37°C and analyzed CK2β activation at different time points by using an antibody that detects serine phosphorylation at position 209 of CK2β
Inhibition of CK2β expression accelerates influenza virus internalization Having shown that 1) CK2β activation is inhibited during IVA replication and 2) CK2β silencing results in increased virus titers, we studied the effect of CK2β gene silencing on the initial step of the viral replication cycle: virus entry
Summary
Influenza A virus (IVA) exploits diverse cellular gene products to support its replication in the host. The significance of the regulatory (β) subunit of casein kinase 2 (CK2β) in various cellular mechanisms is well established, but less is known about its potential role in IVA replication. We studied the role of CK2β in IVA-infected A549 human epithelial lung cells. CK2 is involved in many signal transduction pathways and in negative regulation of apoptosis [8,9,10]. It appears to play crucial roles in cell cycle and growth control via phosphorylation of several important substrates, including p53 [11], Cdc2 [12], and BRCA1 [13]. CK2β is essential for cell viability in mice and is required during early embryonic development [6]
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